Links to Exploration step
Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Platelets modulate multiple markers of neutrophil function in response to <italic>in vitro</italic> Toll-like receptor stimulation</title><author><name sortKey="Hally, Kathryn E" sort="Hally, Kathryn E" uniqKey="Hally K" first="Kathryn E." last="Hally">Kathryn E. Hally</name><affiliation><nlm:aff id="aff001"><addr-line>Department of Surgery and Anaesthesia, University of Otago, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Bird, Georgina K" sort="Bird, Georgina K" uniqKey="Bird G" first="Georgina K." last="Bird">Georgina K. Bird</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="La Flamme, Anne C" sort="La Flamme, Anne C" uniqKey="La Flamme A" first="Anne C." last="La Flamme">Anne C. La Flamme</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Harding, Scott A" sort="Harding, Scott A" uniqKey="Harding S" first="Scott A." last="Harding">Scott A. Harding</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff004"><addr-line>Department of Cardiology, Wellington Hospital, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Larsen, Peter D" sort="Larsen, Peter D" uniqKey="Larsen P" first="Peter D." last="Larsen">Peter D. Larsen</name><affiliation><nlm:aff id="aff001"><addr-line>Department of Surgery and Anaesthesia, University of Otago, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">31581214</idno><idno type="pmc">6776355</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6776355</idno><idno type="RBID">PMC:6776355</idno><idno type="doi">10.1371/journal.pone.0223444</idno><date when="2019">2019</date><idno type="wicri:Area/Pmc/Corpus">000776</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000776</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Platelets modulate multiple markers of neutrophil function in response to <italic>in vitro</italic> Toll-like receptor stimulation</title><author><name sortKey="Hally, Kathryn E" sort="Hally, Kathryn E" uniqKey="Hally K" first="Kathryn E." last="Hally">Kathryn E. Hally</name><affiliation><nlm:aff id="aff001"><addr-line>Department of Surgery and Anaesthesia, University of Otago, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Bird, Georgina K" sort="Bird, Georgina K" uniqKey="Bird G" first="Georgina K." last="Bird">Georgina K. Bird</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="La Flamme, Anne C" sort="La Flamme, Anne C" uniqKey="La Flamme A" first="Anne C." last="La Flamme">Anne C. La Flamme</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Harding, Scott A" sort="Harding, Scott A" uniqKey="Harding S" first="Scott A." last="Harding">Scott A. Harding</name><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff004"><addr-line>Department of Cardiology, Wellington Hospital, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author><author><name sortKey="Larsen, Peter D" sort="Larsen, Peter D" uniqKey="Larsen P" first="Peter D." last="Larsen">Peter D. Larsen</name><affiliation><nlm:aff id="aff001"><addr-line>Department of Surgery and Anaesthesia, University of Otago, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff002"><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></nlm:aff></affiliation><affiliation><nlm:aff id="aff003"><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></nlm:aff></affiliation></author></analytic><series><title level="j">PLoS ONE</title><idno type="eISSN">1932-6203</idno><imprint><date when="2019">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><sec id="sec001"><title>Introduction</title><p>In addition to their role in facilitating leukocyte-mediated inflammation, platelets can dampen leukocyte pro-inflammatory responses in some contexts. Consequently, platelets are increasingly appreciated as regulators of inflammation. Together, platelets and neutrophils play a role in inflammation through Toll-like receptor (TLR) expression, although we do not fully understand how platelets shape neutrophil responses to TLR stimulation. Here, we aimed to determine the extent to which platelets can modulate neutrophil function in response to <italic>in vitro</italic> stimulation with TLR4, TLR2/1, and TLR2/6 agonists.</p></sec><sec id="sec002"><title>Methods</title><p>Neutrophils from 10 healthy individuals were cultured alone or with autologous platelets. Neutrophils ± platelets were left unstimulated or were stimulated with 1 or 100 ng/mL lipopolysaccharide (LPS; a TLR4 agonist), Pam3CSK4 (a TLR2/1 agonist) and fibroblast-stimulating lipopeptide (FSL)-1 (a TLR2/6 agonist). Neutrophil activation and phagocytic activity were assessed by flow cytometry, and elastase and interleukin-8 secretion were assessed by ELISA.</p></sec><sec id="sec003"><title>Results</title><p>The addition of platelets attenuated neutrophil CD66b and CD11b expression in response to various doses of Pam3CSK4 and FSL-1. Furthermore, platelet co-culture was associated with higher CD62L expression (indicating reduced CD62L shedding) in response to these TLR agonists. Platelets also reduced elastase secretion in unstimulated cultures and in response to low-dose TLR stimulation. Conversely, platelet co-culture increased neutrophil phagocytosis in unstimulated cultures and in response to low-dose Pam3CSK4 and FSL-1. Platelets also increased IL-8 secretion in response to low-dose LPS.</p></sec><sec id="sec004"><title>Conclusion</title><p>Platelets are complex immunomodulators that can attenuate some, and simultaneously augment other, neutrophil functions. This modulation can occur both in the absence and presence of TLR stimulation.</p></sec></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Yeaman, Mr" uniqKey="Yeaman M">MR Yeaman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Clark, Sr" uniqKey="Clark S">SR Clark</name></author><author><name sortKey="Ma, Ac" uniqKey="Ma A">AC Ma</name></author><author><name sortKey="Tavener, Sa" uniqKey="Tavener S">SA Tavener</name></author><author><name sortKey="Mcdonald, B" uniqKey="Mcdonald B">B McDonald</name></author><author><name sortKey="Goodarzi, Z" uniqKey="Goodarzi Z">Z Goodarzi</name></author><author><name sortKey="Kelly, Mm" uniqKey="Kelly M">MM Kelly</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Verschoor, A" uniqKey="Verschoor A">A Verschoor</name></author><author><name sortKey="Neuenhahn, M" uniqKey="Neuenhahn M">M Neuenhahn</name></author><author><name sortKey="Navarini, Aa" uniqKey="Navarini A">AA Navarini</name></author><author><name sortKey="Graef, P" uniqKey="Graef P">P Graef</name></author><author><name sortKey="Plaumann, A" uniqKey="Plaumann A">A Plaumann</name></author><author><name sortKey="Seidlmeier, A" uniqKey="Seidlmeier A">A Seidlmeier</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Cognasse, F" uniqKey="Cognasse F">F Cognasse</name></author><author><name sortKey="Hamzeh, H" uniqKey="Hamzeh H">H Hamzeh</name></author><author><name sortKey="Chavarin, P" uniqKey="Chavarin P">P Chavarin</name></author><author><name sortKey="Acquart, S" uniqKey="Acquart S">S Acquart</name></author><author><name sortKey="Genin, C" uniqKey="Genin C">C Genin</name></author><author><name sortKey="Garraud, O" uniqKey="Garraud O">O Garraud</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Shiraki, R" uniqKey="Shiraki R">R Shiraki</name></author><author><name sortKey="Inoue, N" uniqKey="Inoue N">N Inoue</name></author><author><name sortKey="Kawasaki, S" uniqKey="Kawasaki S">S Kawasaki</name></author><author><name sortKey="Takei, A" uniqKey="Takei A">A Takei</name></author><author><name sortKey="Kadotani, M" uniqKey="Kadotani M">M Kadotani</name></author><author><name sortKey="Ohnishi, Y" uniqKey="Ohnishi Y">Y Ohnishi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hally, Ke" uniqKey="Hally K">KE Hally</name></author><author><name sortKey="La Flamme, Ac" uniqKey="La Flamme A">AC La Flamme</name></author><author><name sortKey="Larsen, Pd" uniqKey="Larsen P">PD Larsen</name></author><author><name sortKey="Harding, Sa" uniqKey="Harding S">SA Harding</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Blair, P" uniqKey="Blair P">P Blair</name></author><author><name sortKey="Rex, S" uniqKey="Rex S">S Rex</name></author><author><name sortKey="Vitseva, O" uniqKey="Vitseva O">O Vitseva</name></author><author><name sortKey="Beaulieu, L" uniqKey="Beaulieu L">L Beaulieu</name></author><author><name sortKey="Tanriverdi, K" uniqKey="Tanriverdi K">K Tanriverdi</name></author><author><name sortKey="Chakrabarti, S" uniqKey="Chakrabarti S">S Chakrabarti</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Rivadeneyra, L" uniqKey="Rivadeneyra L">L Rivadeneyra</name></author><author><name sortKey="Carestia, A" uniqKey="Carestia A">A Carestia</name></author><author><name sortKey="Etulain, J" uniqKey="Etulain J">J Etulain</name></author><author><name sortKey="Pozner, Rg" uniqKey="Pozner R">RG Pozner</name></author><author><name sortKey="Fondevila, C" uniqKey="Fondevila C">C Fondevila</name></author><author><name sortKey="Negrotto, S" uniqKey="Negrotto S">S Negrotto</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zhang, G" uniqKey="Zhang G">G Zhang</name></author><author><name sortKey="Han, J" uniqKey="Han J">J Han</name></author><author><name sortKey="Welch, Ej" uniqKey="Welch E">EJ Welch</name></author><author><name sortKey="Ye, Rd" uniqKey="Ye R">RD Ye</name></author><author><name sortKey="Voyno Yasenetskaya, Ta" uniqKey="Voyno Yasenetskaya T">TA Voyno-Yasenetskaya</name></author><author><name sortKey="Malik, Ab" uniqKey="Malik A">AB Malik</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hally, Ke" uniqKey="Hally K">KE Hally</name></author><author><name sortKey="La Flamme, Ac" uniqKey="La Flamme A">AC La Flamme</name></author><author><name sortKey="Harding, Sa" uniqKey="Harding S">SA Harding</name></author><author><name sortKey="Larsen, Pd" uniqKey="Larsen P">PD Larsen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Stocker, Tj" uniqKey="Stocker T">TJ Stocker</name></author><author><name sortKey="Ishikawa Ankerhold, H" uniqKey="Ishikawa Ankerhold H">H Ishikawa-Ankerhold</name></author><author><name sortKey="Massberg, S" uniqKey="Massberg S">S Massberg</name></author><author><name sortKey="Schulz, C" uniqKey="Schulz C">C Schulz</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="St Hl, A L" uniqKey="St Hl A">A-l Ståhl</name></author><author><name sortKey="Svensson, M" uniqKey="Svensson M">M Svensson</name></author><author><name sortKey="Morgelin, M" uniqKey="Morgelin M">M Mörgelin</name></author><author><name sortKey="Svanborg, C" uniqKey="Svanborg C">C Svanborg</name></author><author><name sortKey="Tarr, Pi" uniqKey="Tarr P">PI Tarr</name></author><author><name sortKey="Mooney, Jc" uniqKey="Mooney J">JC Mooney</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Linke, B" uniqKey="Linke B">B Linke</name></author><author><name sortKey="Schreiber, Y" uniqKey="Schreiber Y">Y Schreiber</name></author><author><name sortKey="Picard Willems, B" uniqKey="Picard Willems B">B Picard-Willems</name></author><author><name sortKey="Slattery, P" uniqKey="Slattery P">P Slattery</name></author><author><name sortKey="Nusing, Rm" uniqKey="Nusing R">RM Nusing</name></author><author><name sortKey="Harder, S" uniqKey="Harder S">S Harder</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jancinova, V" uniqKey="Jancinova V">V Jancinova</name></author><author><name sortKey="Drabikova, K" uniqKey="Drabikova K">K Drabikova</name></author><author><name sortKey="Petrikova, M" uniqKey="Petrikova M">M Petrikova</name></author><author><name sortKey="Nosal, R" uniqKey="Nosal R">R Nosal</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Reinisch, Cm" uniqKey="Reinisch C">CM Reinisch</name></author><author><name sortKey="Dunzendorfer, S" uniqKey="Dunzendorfer S">S Dunzendorfer</name></author><author><name sortKey="Pechlaner, C" uniqKey="Pechlaner C">C Pechlaner</name></author><author><name sortKey="Ricevuti, G" uniqKey="Ricevuti G">G Ricevuti</name></author><author><name sortKey="Wiedermann, Cj" uniqKey="Wiedermann C">CJ Wiedermann</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hally, Ke" uniqKey="Hally K">KE Hally</name></author><author><name sortKey="La Flamme, Ac" uniqKey="La Flamme A">AC La Flamme</name></author><author><name sortKey="Harding, Sa" uniqKey="Harding S">SA Harding</name></author><author><name sortKey="Larsen, Pd" uniqKey="Larsen P">PD Larsen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ziegler, M" uniqKey="Ziegler M">M Ziegler</name></author><author><name sortKey="Wang, X" uniqKey="Wang X">X Wang</name></author><author><name sortKey="Peter, K" uniqKey="Peter K">K Peter</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Xiang, B" uniqKey="Xiang B">B Xiang</name></author><author><name sortKey="Zhang, G" uniqKey="Zhang G">G Zhang</name></author><author><name sortKey="Guo, L" uniqKey="Guo L">L Guo</name></author><author><name sortKey="Li, Xa" uniqKey="Li X">XA Li</name></author><author><name sortKey="Morris, Aj" uniqKey="Morris A">AJ Morris</name></author><author><name sortKey="Daugherty, A" uniqKey="Daugherty A">A Daugherty</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Bain, W" uniqKey="Bain W">W Bain</name></author><author><name sortKey="Olonisakin, T" uniqKey="Olonisakin T">T Olonisakin</name></author><author><name sortKey="Yu, M" uniqKey="Yu M">M Yu</name></author><author><name sortKey="Qu, Y" uniqKey="Qu Y">Y Qu</name></author><author><name sortKey="Hulver, M" uniqKey="Hulver M">M Hulver</name></author><author><name sortKey="Xiong, Z" uniqKey="Xiong Z">Z Xiong</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Walsh, Tg" uniqKey="Walsh T">TG Walsh</name></author><author><name sortKey="Poole, Aw" uniqKey="Poole A">AW Poole</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gros, A" uniqKey="Gros A">A Gros</name></author><author><name sortKey="Ollivier, V" uniqKey="Ollivier V">V Ollivier</name></author><author><name sortKey="Ho Tin Noe, B" uniqKey="Ho Tin Noe B">B Ho-Tin-Noe</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Assinger, A" uniqKey="Assinger A">A Assinger</name></author><author><name sortKey="Laky, M" uniqKey="Laky M">M Laky</name></author><author><name sortKey="Badrnya, S" uniqKey="Badrnya S">S Badrnya</name></author><author><name sortKey="Esfandeyari, A" uniqKey="Esfandeyari A">A Esfandeyari</name></author><author><name sortKey="Volf, I" uniqKey="Volf I">I Volf</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zarbock, A" uniqKey="Zarbock A">A Zarbock</name></author><author><name sortKey="Singbartl, K" uniqKey="Singbartl K">K Singbartl</name></author><author><name sortKey="Ley, K" uniqKey="Ley K">K Ley</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Mcdonald, B" uniqKey="Mcdonald B">B McDonald</name></author><author><name sortKey="Davis, Rp" uniqKey="Davis R">RP Davis</name></author><author><name sortKey="Kim, Sj" uniqKey="Kim S">SJ Kim</name></author><author><name sortKey="Tse, M" uniqKey="Tse M">M Tse</name></author><author><name sortKey="Esmon, Ct" uniqKey="Esmon C">CT Esmon</name></author><author><name sortKey="Kolaczkowska, E" uniqKey="Kolaczkowska E">E Kolaczkowska</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Herbertsson, H" uniqKey="Herbertsson H">H Herbertsson</name></author><author><name sortKey="Bengtsson, T" uniqKey="Bengtsson T">T Bengtsson</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Losche, W" uniqKey="Losche W">W Losche</name></author><author><name sortKey="Dressel, M" uniqKey="Dressel M">M Dressel</name></author><author><name sortKey="Krause, S" uniqKey="Krause S">S Krause</name></author><author><name sortKey="Redlich, H" uniqKey="Redlich H">H Redlich</name></author><author><name sortKey="Spangenberg, P" uniqKey="Spangenberg P">P Spangenberg</name></author><author><name sortKey="Heptinstall, S" uniqKey="Heptinstall S">S Heptinstall</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Del Principe, D" uniqKey="Del Principe D">D Del Principe</name></author><author><name sortKey="Menichelli, A" uniqKey="Menichelli A">A Menichelli</name></author><author><name sortKey="Di Giulio, S" uniqKey="Di Giulio S">S Di Giulio</name></author><author><name sortKey="De Matteis, W" uniqKey="De Matteis W">W De Matteis</name></author><author><name sortKey="Giordani, M" uniqKey="Giordani M">M Giordani</name></author><author><name sortKey="Pentassuglio, Am" uniqKey="Pentassuglio A">AM Pentassuglio</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Fortunati, E" uniqKey="Fortunati E">E Fortunati</name></author><author><name sortKey="Kazemier, Km" uniqKey="Kazemier K">KM Kazemier</name></author><author><name sortKey="Grutters, Jc" uniqKey="Grutters J">JC Grutters</name></author><author><name sortKey="Koenderman, L" uniqKey="Koenderman L">L Koenderman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Van Oostrom, Aj" uniqKey="Van Oostrom A">AJ van Oostrom</name></author><author><name sortKey="Van Wijk, Jp" uniqKey="Van Wijk J">JP van Wijk</name></author><author><name sortKey="Sijmonsma, Tp" uniqKey="Sijmonsma T">TP Sijmonsma</name></author><author><name sortKey="Rabelink, Tj" uniqKey="Rabelink T">TJ Rabelink</name></author><author><name sortKey="Castro Cabezas, M" uniqKey="Castro Cabezas M">M Castro Cabezas</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Stokes, Ky" uniqKey="Stokes K">KY Stokes</name></author><author><name sortKey="Granger, Dn" uniqKey="Granger D">DN Granger</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gudbrandsdottir, S" uniqKey="Gudbrandsdottir S">S Gudbrandsdottir</name></author><author><name sortKey="Hasselbalch, Hc" uniqKey="Hasselbalch H">HC Hasselbalch</name></author><author><name sortKey="Nielsen, Ch" uniqKey="Nielsen C">CH Nielsen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Nami, N" uniqKey="Nami N">N Nami</name></author><author><name sortKey="Feci, L" uniqKey="Feci L">L Feci</name></author><author><name sortKey="Napoliello, L" uniqKey="Napoliello L">L Napoliello</name></author><author><name sortKey="Giordano, A" uniqKey="Giordano A">A Giordano</name></author><author><name sortKey="Lorenzini, S" uniqKey="Lorenzini S">S Lorenzini</name></author><author><name sortKey="Galeazzi, M" uniqKey="Galeazzi M">M Galeazzi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Sadallah, S" uniqKey="Sadallah S">S Sadallah</name></author><author><name sortKey="Eken, C" uniqKey="Eken C">C Eken</name></author><author><name sortKey="Martin, Pj" uniqKey="Martin P">PJ Martin</name></author><author><name sortKey="Schifferli, Ja" uniqKey="Schifferli J">JA Schifferli</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jancinova, V" uniqKey="Jancinova V">V Jancinova</name></author><author><name sortKey="Drabikova, K" uniqKey="Drabikova K">K Drabikova</name></author><author><name sortKey="Nosal, R" uniqKey="Nosal R">R Nosal</name></author><author><name sortKey="Petrikova, M" uniqKey="Petrikova M">M Petrikova</name></author><author><name sortKey="Ciz, M" uniqKey="Ciz M">M Ciz</name></author><author><name sortKey="Lojek, A" uniqKey="Lojek A">A Lojek</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Deree, J" uniqKey="Deree J">J Deree</name></author><author><name sortKey="Lall, R" uniqKey="Lall R">R Lall</name></author><author><name sortKey="Melbostad, H" uniqKey="Melbostad H">H Melbostad</name></author><author><name sortKey="Grant, M" uniqKey="Grant M">M Grant</name></author><author><name sortKey="Hoyt, Db" uniqKey="Hoyt D">DB Hoyt</name></author><author><name sortKey="Coimbra, R" uniqKey="Coimbra R">R Coimbra</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lacy, P" uniqKey="Lacy P">P Lacy</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Le Cabec, V" uniqKey="Le Cabec V">V Le Cabec</name></author><author><name sortKey="Carreno, S" uniqKey="Carreno S">S Carreno</name></author><author><name sortKey="Moisand, A" uniqKey="Moisand A">A Moisand</name></author><author><name sortKey="Bordier, C" uniqKey="Bordier C">C Bordier</name></author><author><name sortKey="Maridonneau Parini, I" uniqKey="Maridonneau Parini I">I Maridonneau-Parini</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Diacovo, Tg" uniqKey="Diacovo T">TG Diacovo</name></author><author><name sortKey="Roth, Sj" uniqKey="Roth S">SJ Roth</name></author><author><name sortKey="Buccola, Jm" uniqKey="Buccola J">JM Buccola</name></author><author><name sortKey="Bainton, Df" uniqKey="Bainton D">DF Bainton</name></author><author><name sortKey="Springer, Ta" uniqKey="Springer T">TA Springer</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Corken, A" uniqKey="Corken A">A Corken</name></author><author><name sortKey="Russell, S" uniqKey="Russell S">S Russell</name></author><author><name sortKey="Dent, J" uniqKey="Dent J">J Dent</name></author><author><name sortKey="Post, Sr" uniqKey="Post S">SR Post</name></author><author><name sortKey="Ware, J" uniqKey="Ware J">J Ware</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Klarstrom Engstrom, K" uniqKey="Klarstrom Engstrom K">K Klarstrom Engstrom</name></author><author><name sortKey="Brommesson, C" uniqKey="Brommesson C">C Brommesson</name></author><author><name sortKey="Kalvegren, H" uniqKey="Kalvegren H">H Kalvegren</name></author><author><name sortKey="Bengtsson, T" uniqKey="Bengtsson T">T Bengtsson</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Falker, K" uniqKey="Falker K">K Falker</name></author><author><name sortKey="Klarstrom Engstrom, K" uniqKey="Klarstrom Engstrom K">K Klarstrom-Engstrom</name></author><author><name sortKey="Bengtsson, T" uniqKey="Bengtsson T">T Bengtsson</name></author><author><name sortKey="Lindahl, Tl" uniqKey="Lindahl T">TL Lindahl</name></author><author><name sortKey="Grenegard, M" uniqKey="Grenegard M">M Grenegard</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Senis, Ya" uniqKey="Senis Y">YA Senis</name></author><author><name sortKey="Mazharian, A" uniqKey="Mazharian A">A Mazharian</name></author><author><name sortKey="Mori, J" uniqKey="Mori J">J Mori</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Assinger, A" uniqKey="Assinger A">A Assinger</name></author><author><name sortKey="Laky, M" uniqKey="Laky M">M Laky</name></author><author><name sortKey="Schabbauer, G" uniqKey="Schabbauer G">G Schabbauer</name></author><author><name sortKey="Hirschl, Am" uniqKey="Hirschl A">AM Hirschl</name></author><author><name sortKey="Buchberger, E" uniqKey="Buchberger E">E Buchberger</name></author><author><name sortKey="Binder, Br" uniqKey="Binder B">BR Binder</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Belaaouaj, A" uniqKey="Belaaouaj A">A Belaaouaj</name></author><author><name sortKey="Kim, Ks" uniqKey="Kim K">KS Kim</name></author><author><name sortKey="Shapiro, Sd" uniqKey="Shapiro S">SD Shapiro</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kawabata, K" uniqKey="Kawabata K">K Kawabata</name></author><author><name sortKey="Hagio, T" uniqKey="Hagio T">T Hagio</name></author><author><name sortKey="Matsuoka, S" uniqKey="Matsuoka S">S Matsuoka</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Doring, G" uniqKey="Doring G">G Doring</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hammond, Me" uniqKey="Hammond M">ME Hammond</name></author><author><name sortKey="Lapointe, Gr" uniqKey="Lapointe G">GR Lapointe</name></author><author><name sortKey="Feucht, Ph" uniqKey="Feucht P">PH Feucht</name></author><author><name sortKey="Hilt, S" uniqKey="Hilt S">S Hilt</name></author><author><name sortKey="Gallegos, Ca" uniqKey="Gallegos C">CA Gallegos</name></author><author><name sortKey="Gordon, Ca" uniqKey="Gordon C">CA Gordon</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Sahoo, M" uniqKey="Sahoo M">M Sahoo</name></author><author><name sortKey="Del Barrio, L" uniqKey="Del Barrio L">L del Barrio</name></author><author><name sortKey="Miller, Ma" uniqKey="Miller M">MA Miller</name></author><author><name sortKey="Re, F" uniqKey="Re F">F Re</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Bardoel Bart, W" uniqKey="Bardoel Bart W">W Bardoel Bart</name></author><author><name sortKey="Kenny Elaine, F" uniqKey="Kenny Elaine F">F Kenny Elaine</name></author><author><name sortKey="Sollberger, G" uniqKey="Sollberger G">G Sollberger</name></author><author><name sortKey="Zychlinsky, A" uniqKey="Zychlinsky A">A Zychlinsky</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Middleton, Ea" uniqKey="Middleton E">EA Middleton</name></author><author><name sortKey="Rondina, Mt" uniqKey="Rondina M">MT Rondina</name></author><author><name sortKey="Schwertz, H" uniqKey="Schwertz H">H Schwertz</name></author><author><name sortKey="Zimmerman, Ga" uniqKey="Zimmerman G">GA Zimmerman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lee, Kh" uniqKey="Lee K">KH Lee</name></author><author><name sortKey="Hui, Kp" uniqKey="Hui K">KP Hui</name></author><author><name sortKey="Tan, Wc" uniqKey="Tan W">WC Tan</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wuescher, Lm" uniqKey="Wuescher L">LM Wuescher</name></author><author><name sortKey="Takashima, A" uniqKey="Takashima A">A Takashima</name></author><author><name sortKey="Worth, Rg" uniqKey="Worth R">RG Worth</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="De Stoppelaar, Sf" uniqKey="De Stoppelaar S">SF de Stoppelaar</name></author><author><name sortKey="Van T Veer, C" uniqKey="Van T Veer C">C van 't Veer</name></author><author><name sortKey="Claushuis, Ta" uniqKey="Claushuis T">TA Claushuis</name></author><author><name sortKey="Albersen, Bj" uniqKey="Albersen B">BJ Albersen</name></author><author><name sortKey="Roelofs, Jj" uniqKey="Roelofs J">JJ Roelofs</name></author><author><name sortKey="Van Der Poll, T" uniqKey="Van Der Poll T">T van der Poll</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hechler, B" uniqKey="Hechler B">B Hechler</name></author><author><name sortKey="Zimmermann, C" uniqKey="Zimmermann C">C Zimmermann</name></author><author><name sortKey="Rabouel, Y" uniqKey="Rabouel Y">Y Rabouel</name></author><author><name sortKey="Magnenat, S" uniqKey="Magnenat S">S Magnenat</name></author><author><name sortKey="Burban, M" uniqKey="Burban M">M Burban</name></author><author><name sortKey="Boisrame Helms, J" uniqKey="Boisrame Helms J">J Boisramé-Helms</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Martinod, K" uniqKey="Martinod K">K Martinod</name></author><author><name sortKey="Wagner, Dd" uniqKey="Wagner D">DD Wagner</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Schauer, C" uniqKey="Schauer C">C Schauer</name></author><author><name sortKey="Janko, C" uniqKey="Janko C">C Janko</name></author><author><name sortKey="Munoz, Le" uniqKey="Munoz L">LE Munoz</name></author><author><name sortKey="Zhao, Y" uniqKey="Zhao Y">Y Zhao</name></author><author><name sortKey="Kienhofer, D" uniqKey="Kienhofer D">D Kienhofer</name></author><author><name sortKey="Frey, B" uniqKey="Frey B">B Frey</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Looney, Mr" uniqKey="Looney M">MR Looney</name></author><author><name sortKey="Nguyen, Jx" uniqKey="Nguyen J">JX Nguyen</name></author><author><name sortKey="Hu, Y" uniqKey="Hu Y">Y Hu</name></author><author><name sortKey="Van Ziffle, Ja" uniqKey="Van Ziffle J">JA Van Ziffle</name></author><author><name sortKey="Lowell, Ca" uniqKey="Lowell C">CA Lowell</name></author><author><name sortKey="Matthay, Ma" uniqKey="Matthay M">MA Matthay</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Luo, S" uniqKey="Luo S">S Luo</name></author><author><name sortKey="Wang, Y" uniqKey="Wang Y">Y Wang</name></author><author><name sortKey="An, Q" uniqKey="An Q">Q An</name></author><author><name sortKey="Chen, H" uniqKey="Chen H">H Chen</name></author><author><name sortKey="Zhao, J" uniqKey="Zhao J">J Zhao</name></author><author><name sortKey="Zhang, J" uniqKey="Zhang J">J Zhang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Abdulnour, Re" uniqKey="Abdulnour R">RE Abdulnour</name></author><author><name sortKey="Dalli, J" uniqKey="Dalli J">J Dalli</name></author><author><name sortKey="Colby, Jk" uniqKey="Colby J">JK Colby</name></author><author><name sortKey="Krishnamoorthy, N" uniqKey="Krishnamoorthy N">N Krishnamoorthy</name></author><author><name sortKey="Timmons, Jy" uniqKey="Timmons J">JY Timmons</name></author><author><name sortKey="Tan, Sh" uniqKey="Tan S">SH Tan</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Frangogiannis, Ng" uniqKey="Frangogiannis N">NG Frangogiannis</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lefer, Am" uniqKey="Lefer A">AM Lefer</name></author><author><name sortKey="Campbell, B" uniqKey="Campbell B">B Campbell</name></author><author><name sortKey="Scalia, R" uniqKey="Scalia R">R Scalia</name></author><author><name sortKey="Lefer, Dj" uniqKey="Lefer D">DJ Lefer</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Hargrave, B" uniqKey="Hargrave B">B Hargrave</name></author><author><name sortKey="Li, F" uniqKey="Li F">F Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Milioli, M" uniqKey="Milioli M">M Milioli</name></author><author><name sortKey="Ibanez Vea, M" uniqKey="Ibanez Vea M">M Ibanez-Vea</name></author><author><name sortKey="Sidoli, S" uniqKey="Sidoli S">S Sidoli</name></author><author><name sortKey="Palmisano, G" uniqKey="Palmisano G">G Palmisano</name></author><author><name sortKey="Careri, M" uniqKey="Careri M">M Careri</name></author><author><name sortKey="Larsen, Mr" uniqKey="Larsen M">MR Larsen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Velez, P" uniqKey="Velez P">P Vélez</name></author><author><name sortKey="Izquierdo, I" uniqKey="Izquierdo I">I Izquierdo</name></author><author><name sortKey="Rosa, I" uniqKey="Rosa I">I Rosa</name></author><author><name sortKey="Garcia, A" uniqKey="Garcia A">Á García</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chatterjee, M" uniqKey="Chatterjee M">M Chatterjee</name></author><author><name sortKey="Huang, Z" uniqKey="Huang Z">Z Huang</name></author><author><name sortKey="Zhang, W" uniqKey="Zhang W">W Zhang</name></author><author><name sortKey="Jiang, L" uniqKey="Jiang L">L Jiang</name></author><author><name sortKey="Hultenby, K" uniqKey="Hultenby K">K Hultenby</name></author><author><name sortKey="Zhu, L" uniqKey="Zhu L">L Zhu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Pokrovskaya, Id" uniqKey="Pokrovskaya I">ID Pokrovskaya</name></author><author><name sortKey="Aronova, Ma" uniqKey="Aronova M">MA Aronova</name></author><author><name sortKey="Kamykowski, Ja" uniqKey="Kamykowski J">JA Kamykowski</name></author><author><name sortKey="Prince, Aa" uniqKey="Prince A">AA Prince</name></author><author><name sortKey="Hoyne, Jd" uniqKey="Hoyne J">JD Hoyne</name></author><author><name sortKey="Calco, Gn" uniqKey="Calco G">GN Calco</name></author></analytic></biblStruct></listBibl></div1></back></TEI><pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">PLoS One</journal-id><journal-id journal-id-type="iso-abbrev">PLoS ONE</journal-id><journal-id journal-id-type="publisher-id">plos</journal-id><journal-id journal-id-type="pmc">plosone</journal-id><journal-title-group><journal-title>PLoS ONE</journal-title></journal-title-group><issn pub-type="epub">1932-6203</issn><publisher><publisher-name>Public Library of Science</publisher-name><publisher-loc>San Francisco, CA USA</publisher-loc></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">31581214</article-id><article-id pub-id-type="pmc">6776355</article-id><article-id pub-id-type="doi">10.1371/journal.pone.0223444</article-id><article-id pub-id-type="publisher-id">PONE-D-19-17079</article-id><article-categories><subj-group subj-group-type="heading"><subject>Research Article</subject></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Cell Biology</subject><subj-group><subject>Cellular Types</subject><subj-group><subject>Animal Cells</subject><subj-group><subject>Blood Cells</subject><subj-group><subject>White Blood Cells</subject><subj-group><subject>Neutrophils</subject></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Cell Biology</subject><subj-group><subject>Cellular Types</subject><subj-group><subject>Animal Cells</subject><subj-group><subject>Immune Cells</subject><subj-group><subject>White Blood Cells</subject><subj-group><subject>Neutrophils</subject></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune Cells</subject><subj-group><subject>White Blood Cells</subject><subj-group><subject>Neutrophils</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune Cells</subject><subj-group><subject>White Blood Cells</subject><subj-group><subject>Neutrophils</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Anatomy</subject><subj-group><subject>Body Fluids</subject><subj-group><subject>Blood</subject><subj-group><subject>Platelets</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Anatomy</subject><subj-group><subject>Body Fluids</subject><subj-group><subject>Blood</subject><subj-group><subject>Platelets</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Physiology</subject><subj-group><subject>Body Fluids</subject><subj-group><subject>Blood</subject><subj-group><subject>Platelets</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Physiology</subject><subj-group><subject>Body Fluids</subject><subj-group><subject>Blood</subject><subj-group><subject>Platelets</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Cell Biology</subject><subj-group><subject>Cellular Types</subject><subj-group><subject>Animal Cells</subject><subj-group><subject>Blood Cells</subject><subj-group><subject>Platelets</subject></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Cell Biology</subject><subj-group><subject>Cell Processes</subject><subj-group><subject>Phagocytosis</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Hematology</subject><subj-group><subject>Blood Coagulation</subject><subj-group><subject>Platelet Activation</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune Response</subject><subj-group><subject>Inflammation</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune Response</subject><subj-group><subject>Inflammation</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Diagnostic Medicine</subject><subj-group><subject>Signs and Symptoms</subject><subj-group><subject>Inflammation</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Pathology and Laboratory Medicine</subject><subj-group><subject>Signs and Symptoms</subject><subj-group><subject>Inflammation</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Physiology</subject><subj-group><subject>Physiological Processes</subject><subj-group><subject>Secretion</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Physiology</subject><subj-group><subject>Physiological Processes</subject><subj-group><subject>Secretion</subject></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune System Proteins</subject><subj-group><subject>Immune Receptors</subject><subj-group><subject>Toll-like Receptors</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Immunology</subject><subj-group><subject>Immune System Proteins</subject><subj-group><subject>Immune Receptors</subject><subj-group><subject>Toll-like Receptors</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Biochemistry</subject><subj-group><subject>Proteins</subject><subj-group><subject>Immune System Proteins</subject><subj-group><subject>Immune Receptors</subject><subj-group><subject>Toll-like Receptors</subject></subj-group></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Biology and Life Sciences</subject><subj-group><subject>Cell Biology</subject><subj-group><subject>Signal Transduction</subject><subj-group><subject>Immune Receptors</subject><subj-group><subject>Toll-like Receptors</subject></subj-group></subj-group></subj-group></subj-group></subj-group><subj-group subj-group-type="Discipline-v3"><subject>Medicine and Health Sciences</subject><subj-group><subject>Inflammatory Diseases</subject></subj-group></subj-group></article-categories><title-group><article-title>Platelets modulate multiple markers of neutrophil function in response to <italic>in vitro</italic> Toll-like receptor stimulation</article-title><alt-title alt-title-type="running-head">Platelets modulate neutrophil function</alt-title></title-group><contrib-group><contrib contrib-type="author"><contrib-id authenticated="true" contrib-id-type="orcid">http://orcid.org/0000-0003-3608-3795</contrib-id><name><surname>Hally</surname><given-names>Kathryn E.</given-names></name><role content-type="http://credit.casrai.org/">Conceptualization</role><role content-type="http://credit.casrai.org/">Data curation</role><role content-type="http://credit.casrai.org/">Formal analysis</role><role content-type="http://credit.casrai.org/">Funding acquisition</role><role content-type="http://credit.casrai.org/">Writing – original draft</role><role content-type="http://credit.casrai.org/">Writing – review & editing</role><xref ref-type="aff" rid="aff001"><sup>1</sup></xref><xref ref-type="aff" rid="aff002"><sup>2</sup></xref><xref ref-type="aff" rid="aff003"><sup>3</sup></xref><xref ref-type="corresp" rid="cor001">*</xref></contrib><contrib contrib-type="author"><name><surname>Bird</surname><given-names>Georgina K.</given-names></name><role content-type="http://credit.casrai.org/">Data curation</role><role content-type="http://credit.casrai.org/">Writing – review & editing</role><xref ref-type="aff" rid="aff002"><sup>2</sup></xref><xref ref-type="aff" rid="aff003"><sup>3</sup></xref></contrib><contrib contrib-type="author"><contrib-id authenticated="true" contrib-id-type="orcid">http://orcid.org/0000-0002-2344-7281</contrib-id><name><surname>La Flamme</surname><given-names>Anne C.</given-names></name><role content-type="http://credit.casrai.org/">Conceptualization</role><role content-type="http://credit.casrai.org/">Funding acquisition</role><role content-type="http://credit.casrai.org/">Supervision</role><role content-type="http://credit.casrai.org/">Writing – review & editing</role><xref ref-type="aff" rid="aff002"><sup>2</sup></xref><xref ref-type="aff" rid="aff003"><sup>3</sup></xref></contrib><contrib contrib-type="author"><name><surname>Harding</surname><given-names>Scott A.</given-names></name><role content-type="http://credit.casrai.org/">Conceptualization</role><role content-type="http://credit.casrai.org/">Funding acquisition</role><role content-type="http://credit.casrai.org/">Supervision</role><role content-type="http://credit.casrai.org/">Writing – review & editing</role><xref ref-type="aff" rid="aff002"><sup>2</sup></xref><xref ref-type="aff" rid="aff003"><sup>3</sup></xref><xref ref-type="aff" rid="aff004"><sup>4</sup></xref></contrib><contrib contrib-type="author"><name><surname>Larsen</surname><given-names>Peter D.</given-names></name><role content-type="http://credit.casrai.org/">Conceptualization</role><role content-type="http://credit.casrai.org/">Formal analysis</role><role content-type="http://credit.casrai.org/">Funding acquisition</role><role content-type="http://credit.casrai.org/">Supervision</role><role content-type="http://credit.casrai.org/">Writing – review & editing</role><xref ref-type="aff" rid="aff001"><sup>1</sup></xref><xref ref-type="aff" rid="aff002"><sup>2</sup></xref><xref ref-type="aff" rid="aff003"><sup>3</sup></xref></contrib></contrib-group><aff id="aff001"><label>1</label><addr-line>Department of Surgery and Anaesthesia, University of Otago, Wellington, New Zealand</addr-line></aff><aff id="aff002"><label>2</label><addr-line>School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand</addr-line></aff><aff id="aff003"><label>3</label><addr-line>Wellington Cardiovascular Research Group, Wellington, New Zealand</addr-line></aff><aff id="aff004"><label>4</label><addr-line>Department of Cardiology, Wellington Hospital, Wellington, New Zealand</addr-line></aff><contrib-group><contrib contrib-type="editor"><name><surname>Gay</surname><given-names>Nick</given-names></name><role>Editor</role><xref ref-type="aff" rid="edit1"></xref></contrib></contrib-group><aff id="edit1"><addr-line>University of Cambridge, UNITED KINGDOM</addr-line></aff><author-notes><fn fn-type="COI-statement" id="coi001"><p><bold>Competing Interests: </bold>The authors have declared that no competing interests exist.</p></fn><corresp id="cor001">* E-mail: <email>kathryn.hally@otago.ac.nz</email></corresp></author-notes><pub-date pub-type="epub"><day>3</day><month>10</month><year>2019</year></pub-date><pub-date pub-type="collection"><year>2019</year></pub-date><volume>14</volume><issue>10</issue><elocation-id>e0223444</elocation-id><history><date date-type="received"><day>18</day><month>6</month><year>2019</year></date><date date-type="accepted"><day>20</day><month>9</month><year>2019</year></date></history><permissions><copyright-statement>© 2019 Hally et al</copyright-statement><copyright-year>2019</copyright-year><copyright-holder>Hally et al</copyright-holder><license xlink:href="http://creativecommons.org/licenses/by/4.0/"><license-p>This is an open access article distributed under the terms of the <ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">Creative Commons Attribution License</ext-link>, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.</license-p></license></permissions><self-uri content-type="pdf" xlink:href="pone.0223444.pdf"></self-uri><abstract><sec id="sec001"><title>Introduction</title><p>In addition to their role in facilitating leukocyte-mediated inflammation, platelets can dampen leukocyte pro-inflammatory responses in some contexts. Consequently, platelets are increasingly appreciated as regulators of inflammation. Together, platelets and neutrophils play a role in inflammation through Toll-like receptor (TLR) expression, although we do not fully understand how platelets shape neutrophil responses to TLR stimulation. Here, we aimed to determine the extent to which platelets can modulate neutrophil function in response to <italic>in vitro</italic> stimulation with TLR4, TLR2/1, and TLR2/6 agonists.</p></sec><sec id="sec002"><title>Methods</title><p>Neutrophils from 10 healthy individuals were cultured alone or with autologous platelets. Neutrophils ± platelets were left unstimulated or were stimulated with 1 or 100 ng/mL lipopolysaccharide (LPS; a TLR4 agonist), Pam3CSK4 (a TLR2/1 agonist) and fibroblast-stimulating lipopeptide (FSL)-1 (a TLR2/6 agonist). Neutrophil activation and phagocytic activity were assessed by flow cytometry, and elastase and interleukin-8 secretion were assessed by ELISA.</p></sec><sec id="sec003"><title>Results</title><p>The addition of platelets attenuated neutrophil CD66b and CD11b expression in response to various doses of Pam3CSK4 and FSL-1. Furthermore, platelet co-culture was associated with higher CD62L expression (indicating reduced CD62L shedding) in response to these TLR agonists. Platelets also reduced elastase secretion in unstimulated cultures and in response to low-dose TLR stimulation. Conversely, platelet co-culture increased neutrophil phagocytosis in unstimulated cultures and in response to low-dose Pam3CSK4 and FSL-1. Platelets also increased IL-8 secretion in response to low-dose LPS.</p></sec><sec id="sec004"><title>Conclusion</title><p>Platelets are complex immunomodulators that can attenuate some, and simultaneously augment other, neutrophil functions. This modulation can occur both in the absence and presence of TLR stimulation.</p></sec></abstract><funding-group><funding-statement>K.E.H. was supported by a Doctoral Scholarship from Victoria University of Wellington, New Zealand (<ext-link ext-link-type="uri" xlink:href="http://www.victoria.ac.nz">www.victoria.ac.nz</ext-link>) and is currently supported by a Postdoctoral Fellowship from the Heart Foundation, New Zealand (<ext-link ext-link-type="uri" xlink:href="http://www.heartfoundation.org.nz">www.heartfoundation.org.nz</ext-link>). G.K.B. is currently supported by a Doctoral Scholarship from Victoria University of Wellington. This study was funded by the Wellington Medical Research Foundation, Research For Life (2016/279; <ext-link ext-link-type="uri" xlink:href="http://www.researchforlife.org.nz">www.researchforlife.org.nz</ext-link>). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.</funding-statement></funding-group><counts><fig-count count="8"></fig-count><table-count count="3"></table-count><page-count count="18"></page-count></counts><custom-meta-group><custom-meta id="data-availability"><meta-name>Data Availability</meta-name><meta-value>All relevant data are within the manuscript and its Supporting Information files.</meta-value></custom-meta></custom-meta-group></article-meta><notes><title>Data Availability</title><p>All relevant data are within the manuscript and its Supporting Information files.</p></notes></front><body><sec sec-type="intro" id="sec005"><title>Introduction</title><p>Alongside their roles in hemostasis and thrombosis, platelets have emerged as key effectors of host-defense [<xref rid="pone.0223444.ref001" ref-type="bibr">1</xref>]. Platelets participate in this process principally via cross-talk with leukocytes, where platelets enhance a number of host-defense functions including neutrophil extracellular trap (NET) formation [<xref rid="pone.0223444.ref002" ref-type="bibr">2</xref>] and effective antigen presentation [<xref rid="pone.0223444.ref003" ref-type="bibr">3</xref>]. Platelets are able to mediate these inflammatory responses partly via expression of Toll-like receptors (TLRs) [<xref rid="pone.0223444.ref004" ref-type="bibr">4</xref>–<xref rid="pone.0223444.ref006" ref-type="bibr">6</xref>]. TLRs are crucial host-defense mechanisms and some TLRs are important in eliciting platelet-mediated inflammation [<xref rid="pone.0223444.ref006" ref-type="bibr">6</xref>–<xref rid="pone.0223444.ref010" ref-type="bibr">10</xref>].</p><p>The involvement of platelets in shaping inflammation is complex. Recent evidence suggests that, in addition to their role in promoting inflammation, platelets provide anti-inflammatory cues to dampen leukocyte responses during excessive inflammation [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>]. For example, platelets can attenuate the production of pro-inflammatory cytokines [<xref rid="pone.0223444.ref012" ref-type="bibr">12</xref>, <xref rid="pone.0223444.ref013" ref-type="bibr">13</xref>] and reactive oxygen species (ROS) [<xref rid="pone.0223444.ref014" ref-type="bibr">14</xref>, <xref rid="pone.0223444.ref015" ref-type="bibr">15</xref>], and attenuate expression of activation markers [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>] by immune cells in response to inflammatory stimuli. In light of these findings, platelets are increasingly appreciated as immune regulators, rather than purely pro-inflammatory cells. Interestingly, this regulatory response has been characterized in a number of inflammatory diseases where platelets are also known to be predominant drivers of leukocyte infiltration and leukocyte-mediated inflammation [<xref rid="pone.0223444.ref017" ref-type="bibr">17</xref>–<xref rid="pone.0223444.ref019" ref-type="bibr">19</xref>]. The dynamicity in platelet responses may be a mechanism to enhance, and then temper, an inflammatory response [<xref rid="pone.0223444.ref020" ref-type="bibr">20</xref>] to prevent rampant host damage [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>] and is also both context- and stimulus-dependent [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>, <xref rid="pone.0223444.ref021" ref-type="bibr">21</xref>].</p><p>We have previously investigated the role of platelets in regulating leukocyte-mediated inflammation in response to TLR stimulation [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>]. In addition to examining other leukocyte populations, we showed that platelets attenuated neutrophil elastase secretion and expression of the activation marker, CD66b, in response to <italic>in vitro</italic> stimulation with lipopolysaccharide (LPS, a TLR4 agonist), Pam3CSK4 (a TLR2/1 agonist) and fibroblast-stimulating lipopeptide (FSL)-1 (a TLR2/6 agonist). We examined this particular subset of platelet-TLRs as we have previously shown that platelets elicit very different patterns of activation in response to these prototypical TLR agonists. We [<xref rid="pone.0223444.ref006" ref-type="bibr">6</xref>] and others [<xref rid="pone.0223444.ref007" ref-type="bibr">7</xref>, <xref rid="pone.0223444.ref008" ref-type="bibr">8</xref>, <xref rid="pone.0223444.ref022" ref-type="bibr">22</xref>] have consistently shown that platelets become directly activated in response to Pam3CSK4, but platelets show minimal activation following incubation with either LPS or FSL-1 [<xref rid="pone.0223444.ref002" ref-type="bibr">2</xref>, <xref rid="pone.0223444.ref006" ref-type="bibr">6</xref>, <xref rid="pone.0223444.ref008" ref-type="bibr">8</xref>]. However, although not able to induce platelet activation or aggregation, LPS has been shown to facilitate platelet-neutrophil aggregation and subsequent robust production of neutrophil extracellular traps (NETs) [<xref rid="pone.0223444.ref002" ref-type="bibr">2</xref>]. These results suggest that platelets may mediate a more complex effect to LPS and FSL-1 via their interaction with leukocytes.</p><p>Due to the broad nature of our previous study, where we aimed to assess how platelets modulated the function of various leukocyte subsets, we did not examine neutrophil function beyond these two measurements. Thus, a detailed description of the platelet effect on neutrophil responses to TLR stimulation has yet to be conducted. This is particularly important given that platelets can exert both pro-inflammatory [<xref rid="pone.0223444.ref002" ref-type="bibr">2</xref>, <xref rid="pone.0223444.ref023" ref-type="bibr">23</xref>, <xref rid="pone.0223444.ref024" ref-type="bibr">24</xref>] and anti-inflammatory [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>, <xref rid="pone.0223444.ref025" ref-type="bibr">25</xref>–<xref rid="pone.0223444.ref027" ref-type="bibr">27</xref>] effects on neutrophils in response to various other stimuli. We suggest that different platelet phenotypes can be triggered in a stimulus-specific manner, and this has yet to be fully examined in response to TLR stimulation. Here, we aimed to determine the extent to which platelets can modulate neutrophil responses to <italic>in vitro</italic> stimulation with TLR4, TLR2/1, and TLR2/6 agonists.</p></sec><sec sec-type="materials|methods" id="sec006"><title>Materials and methods</title><sec id="sec007"><title>Neutrophil and platelet isolation</title><p>Ten healthy subjects (5 male, mean age 26 ± 3 years) were recruited into this study. Ethical approval was granted by the University of Otago Human Ethics Committee (HE16/004). Ten mL blood was drawn by venipuncture from each subject. Neutrophils were isolated from EDTA-anticoagulated blood by magnetic negative selection as per manufacturer’s instructions (Miltenyi Biotec, Bergish Gladbach, Germany). Neutrophils were then washed and resuspended in culture media (10% FCS, 2 mM L-glutamate, 100 U/mL penicillin, 100 μg/mL streptomycin, 0.01M HEPES buffer, 0.1% β-mercaptoethanol, 0.01 nM non-essential amino acids) to 10<sup>6</sup> neutrophils/mL. Platelet-rich plasma (PRP) and platelet-poor plasma (PPP) were isolated from hirudin-anticoagulated blood by centrifugation at 200 x g for 12 minutes or at 1500 x g for 12 minutes, respectively. PRP was adjusted to 2.5x10<sup>8</sup> platelets/mL in phosphate-buffered saline (PBS; 145 mM NaCl, 8.7 mM Na<sub>2</sub>HPO<sub>4</sub>, 1.3 mM NaH<sub>2</sub>PO<sub>4</sub>). PPP was similarly diluted in PBS. PRP and PPP were used to assess the platelet effect on markers of neutrophil activation by flow cytometry. PRP was also used to produce washed platelets (WPs). Briefly, PRP was diluted in PBS with 1 μM prostaglandin E1, pelleted, and platelets were resuspended to 2.5x10<sup>8</sup> platelets/mL in culture media. WPs were used to assess the platelet effect on neutrophil phagocytosis. Platelets are quiescent during isolation from whole blood and prior to culture, as platelets did not stain positively for PAC1, a platelet activation marker, by flow cytometry (<xref ref-type="supplementary-material" rid="pone.0223444.s001">S1 Fig</xref>). Platelets also remain quiescent when cultured alone (in the absence of TLR stimulation and neutrophil co-culture) for 4 hours (also <xref ref-type="supplementary-material" rid="pone.0223444.s001">S1 Fig</xref>).</p></sec><sec id="sec008"><title><italic>In vitro</italic> TLR stimulation</title><p>To assess markers of neutrophil activation by flow cytometry, neutrophils were cultured with PRP in a ratio of 1:250 neutrophils: platelets (+ platelets). An equal amount of PPP was added to neutrophil-only cultures (- platelets). We have previously demonstrated that platelets exert their effect on leukocytes in a dose-dependent manner, and that this effect was most apparent at a neutrophil: platelet ratio of 1:250 [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>]. In observance to this previous finding, we have employed the same neutrophil: platelet ratio in this study. To assess neutrophil phagocytosis, neutrophils were cultured either with WPs (+ platelets) or culture media (-platelets) and incubated with FITC-labelled rabbit IgG-coated latex beads (Cayman Chemicals, Ann Arbor, MI, USA) in a ratio of 1 μL of latex beads to every 200 μL culture media. For both neutrophil activation and phagocytosis, neutrophils ± platelets were left unstimulated or stimulated with 1 and 100 ng/mL of the following for 4 hours at 37°C/5% CO<sub>2</sub>: LPS from <italic>Escherichia coli</italic> serotype R515 (a TLR4 agonist; Enzo Life Sciences, Farmingdale, NY, USA); Pam3CSK4 (a TLR2/1 agonist; Tocris Bioscience, Bristol, UK) and FSL-1 (a TLR2/6 agonist; Santa Cruz Biotechnology, Santa Cruz, CA, USA). LPS and FSL-1 were guaranteed by the respective manufacturers to be free of any contaminants that have agonist TLR activity.</p></sec><sec id="sec009"><title>Assessing neutrophil function</title><p>To assess neutrophil activation, neutrophils ± platelets were incubated with anti-CD16-BV421 (clone 3G8), anti-CD66b-BB515 (clone G10F5), anti-CD11b-BV510 (clone ICRF44), all sourced from Becton Dickinson, and anti-CD62L-APC (DREG-56; BioLegend, San Diego, CA, USA), or the appropriate isotype control for 50 minutes at 4°C, followed by fixation. To assess neutrophil phagocytosis, cells were incubated with Trypan blue to quench fluorescence from surface-bound latex beads-FITC as recommended by the manufacturer, and were then washed and fixed. All samples were run on a FACSCanto II flow cytometer (Becton Dickinson), and data were analyzed using FlowJo software (v10.0.7, Tree star).</p><p>For measuring neutrophil activation, neutrophils were identified firstly by doublet exclusion and secondly by high expression of CD16, as outlined in <xref ref-type="supplementary-material" rid="pone.0223444.s002">S2 Fig</xref>. Representative plots of each flow cytometry activation marker (CD66b, CD62L and CD11b) in unstimulated and 100 ng/mL FSL-1 stimulated cultures are given in <xref ref-type="fig" rid="pone.0223444.g001">Fig 1</xref>. Increased cell-surface CD66b and CD11b expression, and increased CD62L shedding (reduced CD62L expression) are all established markers of neutrophil activation [<xref rid="pone.0223444.ref028" ref-type="bibr">28</xref>, <xref rid="pone.0223444.ref029" ref-type="bibr">29</xref>]. For reporting on the expression of CD11b, CD66b and CD62L, delta geometric mean fluorescence intensity (ΔgMFI) was calculated by subtracting the gMFI of the isotype control from the gMFI of each sample for each antibody used. For measuring neutrophil phagocytosis, neutrophils were identified by doublet exclusion. The percentage of neutrophils that were positive for FITC fluorescence (positive for internalizing latex beads conjugated to FITC) was reported. A representative plot of neutrophil phagocytosis is given in <xref ref-type="fig" rid="pone.0223444.g001">Fig 1</xref>. Representative plots of these flow cytometry markers (CD11b, CD66b, CD62L, phagocytosis) with and without the addition of platelets in 100 ng/mL FSL-1 stimulated cultures are given in <xref ref-type="fig" rid="pone.0223444.g002">Fig 2</xref>. Platelets become dimly positive for PAC1, a platelet activation marker, following incubation with these TLR agonists (without the addition of neutrophils), and expression increases when platelets and neutrophils are co-cultured together (<xref ref-type="supplementary-material" rid="pone.0223444.s003">S3 Fig</xref>).</p><fig id="pone.0223444.g001" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g001</object-id><label>Fig 1</label><caption><title>Detection of neutrophil responses to <italic>in vitro</italic> TLR stimulation by flow cytometry.</title><p>Highly CD16-positive neutrophils were identified as outlined in <xref ref-type="supplementary-material" rid="pone.0223444.s002">S2 Fig</xref>. Representative plots of neutrophil CD66b expression (A), CD62L expression (B) and CD11b expression (C) are shown for unstimulated and FSL-1 (100 ng/mL)-stimulated neutrophils following 4 hours of culture. CD66b and CD11b expression levels increase following neutrophil activation, while CD62L levels decrease (CD62L is shed from the neutrophil surface). A representative plot of neutrophil phagocytic activity, as measured by the percentage of latex bead-FITC-positive neutrophils (indicating % of neutrophils that have internalized latex beads conjugated to FITC), is shown for unstimulated and FSL-1-stimulated (100 ng/mL) conditions (D).</p></caption><graphic xlink:href="pone.0223444.g001"></graphic></fig><fig id="pone.0223444.g002" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g002</object-id><label>Fig 2</label><caption><title>Platelets modulate neutrophil responses to <italic>in vitro</italic> TLR stimulation.</title><p>Representative plots of neutrophil CD66b expression (A), CD62L expression (B), CD11b expression (C), and neutrophil phagocytic activity (D) are shown for neutrophils stimulated with 100 ng/mL FSL-1, with and without the addition of platelets.</p></caption><graphic xlink:href="pone.0223444.g002"></graphic></fig><p>Elastase secretion (Abcam, Cambridge, United Kingdom) and IL-8 secretion (Thermo Fisher Scientific, Waltham, MA, USA) was measured by ELISA, as per the manufacturers’ instructions.</p></sec><sec id="sec010"><title>Statistical analysis</title><p>Continuous variables are expressed as mean ± standard deviation. In the absence of platelets, the changes in markers of neutrophil function in response to TLR stimulation were examined using one-way ANOVA with post-hoc Dunnett multiple comparison tests. To examine the effect of platelets on the neutrophil response to TLR stimulation, each baseline (- platelets) neutrophil only measurement was normalized to 1 and each platelet co-culture (+ platelets) measurement was reported as a relative change. Differences in these relative changes (- platelets vs. + platelets) were examined using a repeated measures two-way ANOVA (row factor as culture condition, column factor as ± platelets) with post-hoc Sidak multiple comparison tests (to compare row means across the column factor) using GraphPad Prism 7 (GraphPad Software Inc.).</p></sec></sec><sec sec-type="results" id="sec011"><title>Results</title><sec id="sec012"><title>Platelets differentially modulate markers of neutrophil activation in response to TLR stimulation</title><p>In the absence of platelets, stimulation with each TLR agonist induced a significant increase in neutrophil CD66b and CD11b expression and a significant decrease in CD62L expression (indicating increased CD62L shedding). Expression of these markers under each culture condition is shown in Figs <xref ref-type="fig" rid="pone.0223444.g003">3</xref>–<xref ref-type="fig" rid="pone.0223444.g005">5(A)</xref>, and is tabulated in <xref ref-type="supplementary-material" rid="pone.0223444.s004">S1</xref>, <xref ref-type="supplementary-material" rid="pone.0223444.s005">S2</xref> and <xref ref-type="supplementary-material" rid="pone.0223444.s006">S3</xref> Tables. Incubation with 1 and 100 ng/mL of each TLR agonist represented sub-maximal and maximal stimulation, respectively. In response to TLR stimulation, the addition of platelets differentially affected these expression levels (Figs <xref ref-type="fig" rid="pone.0223444.g003">3</xref>–<xref ref-type="fig" rid="pone.0223444.g005">5(B)</xref>, <xref rid="pone.0223444.t001" ref-type="table">Table 1</xref>). All three markers of neutrophil activation (CD66b and CD11b expression, CD62L shedding) measured in this study were attenuated by platelets, and were sensitive to modulation by platelets only in response to Pam3CSK4 and FSL-1, but not in response to LPS.</p><fig id="pone.0223444.g003" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g003</object-id><label>Fig 3</label><caption><title>Neutrophil CD66b expression ± platelet co-culture.</title><p>Raw CD66b expression results are shown in (A). CD66b was measured as ΔgMFI without platelets (- platelets) and with platelets (+ platelets) in unstimulated or TLR-stimulated cultures. For each culture condition, paired measurements (- platelets vs. + platelets) are linked with a solid black line for n = 10 healthy subjects. Relative change in the presence of platelets is shown in (B). ΔgMFI in neutrophil only cultures (- platelets) was normalized to 1 (ble) and each co-culture measurement (+ platelets) was compared to this normalized response. Differences between expression with and without platelets were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. ***p<0.001, ****p<0.0001.</p></caption><graphic xlink:href="pone.0223444.g003"></graphic></fig><fig id="pone.0223444.g004" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g004</object-id><label>Fig 4</label><caption><title>Neutrophil CD62L expression ± platelet co-culture.</title><p>Raw CD62L expression results are shown in (A) for each culture condition, with each paired measurement (- platelets vs. + platelets) linked with a solid black line. CD62L expression is expressed on resting neutrophils, and is shed from the neutrophil surface in response to stimulation. Relative change in the presence of platelets is shown in (B), compared against measurements from neutrophil-only cultures, which were normalized to 1 (ble). Differences in expression (- platelets vs. + platelets) were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. **p<0.01, ****p<0.0001.</p></caption><graphic xlink:href="pone.0223444.g004"></graphic></fig><fig id="pone.0223444.g005" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g005</object-id><label>Fig 5</label><caption><title>Neutrophil CD11b expression ± platelet co-culture.</title><p>Raw CD11b expression results are shown in (A) for each culture condition, with each paired measurement (- platelets vs. + platelets) linked with a solid black line. Relative change in the presence of platelets is shown in (B), compared against measurements from neutrophil-only cultures normalized to 1 (ble). Differences in expression (- platelets vs. + platelets) were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. **p<0.01, ***p<0.001, ****p<0.0001.</p></caption><graphic xlink:href="pone.0223444.g005"></graphic></fig><table-wrap id="pone.0223444.t001" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.t001</object-id><label>Table 1</label><caption><title>Relative change in neutrophil markers measured by flow cytometry in neutrophil-platelet co-culture.</title></caption><alternatives><graphic id="pone.0223444.t001g" xlink:href="pone.0223444.t001"></graphic><table frame="hsides" rules="groups"><colgroup span="1"><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col></colgroup><thead><tr><th align="center" rowspan="1" colspan="1"></th><th align="center" rowspan="1" colspan="1"></th><th align="center" colspan="4" rowspan="1">+ platelets<xref ref-type="table-fn" rid="t001fn001"><sup>1</sup></xref></th></tr><tr><th align="center" rowspan="1" colspan="1">Agonist</th><th align="center" rowspan="1" colspan="1">ng/mL</th><th align="center" rowspan="1" colspan="1">CD66b</th><th align="center" rowspan="1" colspan="1">CD62L</th><th align="center" rowspan="1" colspan="1">CD11b</th><th align="center" rowspan="1" colspan="1">Phagocytosis</th></tr></thead><tbody><tr><td align="center" rowspan="1" colspan="1">Unstimulated</td><td align="center" rowspan="1" colspan="1">--</td><td align="center" rowspan="1" colspan="1">0.97 ± 0.04</td><td align="center" rowspan="1" colspan="1">1.09 ± 0.14</td><td align="center" rowspan="1" colspan="1"><bold>0.85 ± 0.13</bold><xref ref-type="table-fn" rid="t001fn004">***</xref></td><td align="center" rowspan="1" colspan="1"><bold>2.70 ± 1.10</bold><xref ref-type="table-fn" rid="t001fn005">****</xref></td></tr><tr><td align="center" rowspan="1" colspan="1">LPS</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1">0.92 ± 0.17</td><td align="center" rowspan="1" colspan="1">1.06 ± 0.27</td><td align="center" rowspan="1" colspan="1">0.96 ± 0.19</td><td align="center" rowspan="1" colspan="1">1.08 ± 0.15</td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1">1.00 ± 0.12</td><td align="center" rowspan="1" colspan="1">1.00 ± 0.21</td><td align="center" rowspan="1" colspan="1">0.93 ± 0.14</td><td align="center" rowspan="1" colspan="1">1.03 ± 0.12</td></tr><tr><td align="center" rowspan="1" colspan="1">Pam3CSK4</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1">0.94 ± 0.12</td><td align="center" rowspan="1" colspan="1">0.99 ± 0.14</td><td align="center" rowspan="1" colspan="1"><bold>0.88 ± 0.09</bold><xref ref-type="table-fn" rid="t001fn003">**</xref></td><td align="center" rowspan="1" colspan="1"><bold>1.78 ± 0.76</bold><xref ref-type="table-fn" rid="t001fn005">****</xref></td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1"><bold>0.82 ± 0.11</bold><xref ref-type="table-fn" rid="t001fn005">****</xref></td><td align="center" rowspan="1" colspan="1"><bold>1.25 ± 0.42</bold><xref ref-type="table-fn" rid="t001fn005">****</xref></td><td align="center" rowspan="1" colspan="1"><bold>0.85 ± 0.16</bold><xref ref-type="table-fn" rid="t001fn004">***</xref></td><td align="center" rowspan="1" colspan="1">1.17 ± 0.33</td></tr><tr><td align="center" rowspan="1" colspan="1">FSL-1</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"><bold>0.84 ± 0.09</bold><xref ref-type="table-fn" rid="t001fn004">***</xref></td><td align="center" rowspan="1" colspan="1"><bold>1.19 ± 0.13</bold><xref ref-type="table-fn" rid="t001fn003">**</xref></td><td align="center" rowspan="1" colspan="1"><bold>0.81 ± 0.12</bold><xref ref-type="table-fn" rid="t001fn005">****</xref></td><td align="center" rowspan="1" colspan="1"><bold>1.41 ± 0.31</bold><xref ref-type="table-fn" rid="t001fn002">*</xref></td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1">0.94 ± 0.10</td><td align="center" rowspan="1" colspan="1"><bold>1.19 ± 0.35</bold><xref ref-type="table-fn" rid="t001fn003">**</xref></td><td align="center" rowspan="1" colspan="1"><bold>0.88 ± 0.15</bold><xref ref-type="table-fn" rid="t001fn003">**</xref></td><td align="center" rowspan="1" colspan="1">1.11 ± 0.13</td></tr></tbody></table></alternatives><table-wrap-foot><fn id="t001fn001"><p><sup>1</sup> For each subject, all neutrophil only measurements (- platelets) were normalized to 1, and all co-culture measurements (+ platelets) were compared to this normalized response. Relative change in ΔgMFI (CD66b, CD62L, CD11b) or % phagocytosis presented as mean ± standard deviation for n = 10 healthy subjects. Differences (- platelets vs. + platelets) were examined by a repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests.</p></fn><fn id="t001fn002"><p>*p<0.05</p></fn><fn id="t001fn003"><p>**p<0.01</p></fn><fn id="t001fn004"><p>***p<0.001</p></fn><fn id="t001fn005"><p>****p<0.0001.</p></fn></table-wrap-foot></table-wrap><p>Addition of platelets attenuated CD66b expression by 18% in response to high-dose (100 ng/mL) Pam3CSK4, and by 16% in response to low-dose (1 ng/mL) FSL-1 (<xref ref-type="fig" rid="pone.0223444.g003">Fig 3B</xref>, all p<0.01). With platelet co-culture, neutrophil CD62L expression was higher in response to high-dose Pam3CSK4 (25% higher, p<0.0001) and both doses of FSL-1 (19% higher, p<0.01) compared to neutrophil-only cultures (<xref ref-type="fig" rid="pone.0223444.g004">Fig 4B</xref>). These results indicate that platelets attenuate neutrophil CD62L shedding under these conditions.CD11b expression was attenuated in the presence of platelets in response to both Pam3CSK4 and FSL-1 (12% to 19% reduction across these culture conditions, all p<0.01, <xref ref-type="fig" rid="pone.0223444.g005">Fig 5B</xref>). Platelets also modulated CD11b expression in unstimulated cultures (15% reduction, p<0.001).</p></sec><sec id="sec013"><title>Platelets enhance neutrophil phagocytosis in unstimulated cultures and in response to some TLR agonists</title><p>Compared to unstimulated cultures, baseline neutrophil phagocytic activity (in the absence of platelets) was significantly increased in response to all TLR agonist conditions (<xref ref-type="fig" rid="pone.0223444.g006">Fig 6A</xref>, <xref ref-type="supplementary-material" rid="pone.0223444.s007">S4 Table</xref>). In this study, phagocytosis was defined as the percentage of neutrophils that were positive for FITC fluorescence, indicating internalization of latex beads conjugated to FITC that were added to all neutrophil cultures. In unstimulated cultures, the percentage of neutrophils that were positive for phagocytosis was 2.7-fold higher with the addition of platelets, compared to neutrophils alone (p<0.0001, <xref ref-type="fig" rid="pone.0223444.g006">Fig 6B</xref>, <xref rid="pone.0223444.t001" ref-type="table">Table 1</xref>). Platelet co-culture also resulted in a higher rate of phagocytosis seen in response to low-dose Pam3CSK4 (1.78-fold higher) and low-dose FSL-1 (1.41-fold higher, both p<0.05, <xref ref-type="fig" rid="pone.0223444.g006">Fig 6B</xref>).</p><fig id="pone.0223444.g006" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g006</object-id><label>Fig 6</label><caption><title>Neutrophil phagocytosis ± platelet co-culture.</title><p>Phagocytosis was measured as the % of the neutrophil population that was FITC-positive (positive for internalizing latex beads conjugated to FITC) under each culture condition. Raw phagocytosis results are shown in (A). Relative change in the presence of platelets is shown in (B), compared against measurements from neutrophil-only cultures normalized to 1 (ble). Differences in expression (- platelets vs. + platelets) were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. *p <0.05, ****p<0.0001.</p></caption><graphic xlink:href="pone.0223444.g006"></graphic></fig></sec><sec id="sec014"><title>Platelets modulate elastase and IL-8 secretion by neutrophils</title><p>Both elastase and IL-8 secretion were assessed from the supernatant of neutrophil-only and neutrophil-platelet co-cultures. Baseline secretion (-platelets) of both markers increased in response to TLR stimulation (<xref ref-type="fig" rid="pone.0223444.g007">Fig 7A</xref> and <xref ref-type="fig" rid="pone.0223444.g008">Fig 8A</xref>, <xref ref-type="supplementary-material" rid="pone.0223444.s008">S5</xref> and <xref ref-type="supplementary-material" rid="pone.0223444.s009">S6</xref> Tables). For elastase, the increase in secretion was only statistically significant in response to low-dose (1 ng/mL) TLR stimulation.</p><fig id="pone.0223444.g007" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g007</object-id><label>Fig 7</label><caption><title>Neutrophil elastase secretion ± platelet co-culture.</title><p>Raw elastase measurements are shown in (A) for each culture condition. Relative change in the presence of platelets is shown in (B), compared against measurements from neutrophil-only cultures normalized to 1 (ble). Differences in expression (- platelets vs. + platelets) were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. **p<0.01.</p></caption><graphic xlink:href="pone.0223444.g007"></graphic></fig><fig id="pone.0223444.g008" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.g008</object-id><label>Fig 8</label><caption><title>Neutrophil IL-8 secretion ± platelet co-culture.</title><p>Raw IL-8 measurements are shown in (A) for each culture condition. Relative change in the presence of platelets is shown in (B), compared against measurements from neutrophil-only cultures normalized to 1 (ble). Differences in expression (- platelets vs. + platelets) were examined by repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests. ***p<0.001.</p></caption><graphic xlink:href="pone.0223444.g008"></graphic></fig><p>With the addition of platelets, elastase secretion was significantly lowered by 18% in unstimulated cultures (p<0.01, <xref ref-type="fig" rid="pone.0223444.g007">Fig 7B</xref>, <xref rid="pone.0223444.t002" ref-type="table">Table 2</xref>), and the increase in elastase secretion following low-dose TLR stimulation was also attenuated by platelets (17% to 19% reduction; all p<0.01). The presence of platelets significantly altered IL-8 secretion only in response to low-dose LPS: the increase in IL-8 secretion with this dose was 1.28-fold greater in platelet co-culture (p<0.001, <xref ref-type="fig" rid="pone.0223444.g008">Fig 8B</xref>, <xref rid="pone.0223444.t002" ref-type="table">Table 2</xref>).</p><table-wrap id="pone.0223444.t002" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.t002</object-id><label>Table 2</label><caption><title>Relative change in neutrophil secretory products in neutrophil-platelet co-culture.</title></caption><alternatives><graphic id="pone.0223444.t002g" xlink:href="pone.0223444.t002"></graphic><table frame="hsides" rules="groups"><colgroup span="1"><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col></colgroup><thead><tr><th align="center" rowspan="1" colspan="1"></th><th align="center" rowspan="1" colspan="1"></th><th align="center" colspan="2" rowspan="1">+ platelets<xref ref-type="table-fn" rid="t002fn001"><sup>1</sup></xref></th></tr><tr><th align="center" rowspan="1" colspan="1">Agonist</th><th align="center" rowspan="1" colspan="1">ng/mL</th><th align="center" rowspan="1" colspan="1">Elastase</th><th align="center" rowspan="1" colspan="1">IL-8</th></tr></thead><tbody><tr><td align="center" rowspan="1" colspan="1">Unstimulated</td><td align="center" rowspan="1" colspan="1">--</td><td align="center" rowspan="1" colspan="1"><bold>0.82 ± 0.13</bold><xref ref-type="table-fn" rid="t002fn003">**</xref></td><td align="center" rowspan="1" colspan="1">0.94 ± 0.24</td></tr><tr><td align="center" rowspan="1" colspan="1">LPS</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"><bold>0.83 ± 0.14</bold><xref ref-type="table-fn" rid="t002fn003">**</xref></td><td align="center" rowspan="1" colspan="1"><bold>1.28 ± 0.28</bold><xref ref-type="table-fn" rid="t002fn004">***</xref></td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1">0.93 ± 0.31</td><td align="center" rowspan="1" colspan="1">1.00 ± 0.18</td></tr><tr><td align="center" rowspan="1" colspan="1">Pam3CSK4</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"><bold>0.81 ± 0.23</bold><xref ref-type="table-fn" rid="t002fn003">**</xref></td><td align="center" rowspan="1" colspan="1">1.15 ± 0.45</td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1">0.87 ± 0.19</td><td align="center" rowspan="1" colspan="1">0.94 ± 0.30</td></tr><tr><td align="center" rowspan="1" colspan="1">FSL-1</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"><bold>0.81 ± 0.13</bold><xref ref-type="table-fn" rid="t002fn003">**</xref></td><td align="center" rowspan="1" colspan="1">1.05 ± 0.51</td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1">0.89 ± 0.13</td><td align="center" rowspan="1" colspan="1">0.99 ± 0.14</td></tr></tbody></table></alternatives><table-wrap-foot><fn id="t002fn001"><p><sup>1</sup> For each subject, all neutrophil only measurements (- platelets) were normalized to 1, and all co-culture measurements (+ platelets) were compared to this normalized response. Relative change in secretory products presented as mean ± standard deviation for n = 10 healthy subjects. Differences (- platelets vs. + platelets) were examined by a repeated measures two-way ANOVA with post-hoc Sidak multiple comparisons tests.</p></fn><fn id="t002fn002"><p>*p<0.05</p></fn><fn id="t002fn003"><p>**p<0.01</p></fn><fn id="t002fn004"><p>***p<0.001.</p></fn></table-wrap-foot></table-wrap></sec></sec><sec sec-type="conclusions" id="sec015"><title>Discussion</title><p>We demonstrate that platelets differentially modulate several neutrophil functions in a TLR agonist-specific and dose-specific manner. Furthermore, platelets can attenuate some, and simultaneously augment other, neutrophil functions. A summary of the results described in this study is given in <xref rid="pone.0223444.t003" ref-type="table">Table 3</xref>. The influence of platelets on the expression of three well-characterized flow cytometric markers of neutrophil activation were assessed in this study. Platelets attenuated the expression of both CD66b and CD11b on neutrophils stimulated with Pam3CSK4 and FSL-1 and, additionally, CD11b expression was also attenuated in unstimulated cultures. In response to these TLR agonists, neutrophil CD62L expression was higher in the presence of platelets, indicating reduced CD62L shedding under these culture conditions. Conversely, neutrophil phagocytosis was significantly higher in co-culture both without stimulation and with TLR stimulation (Pam3CSK4 and FSL-1). Platelets did not alter the expression of any of these markers in response to LPS. We also assessed the effect of platelets on two neutrophil secretory products. Elastase secretion was attenuated in the presence of platelets in unstimulated cultures as well as in response to low-dose stimulation with all three TLR agonists. We also show that the increase in IL-8 secretion in response to low-dose LPS was further increased in the presence of platelets. In conclusion, platelets modify a number of neutrophil functions in both the absence of stimulation and in response to TLR stimulation. Additionally, it is interesting to note that platelets have a dampening effect on a majority of the markers examined here (CD66b and CD11b expression, CD62L shedding and elastase secretion).</p><table-wrap id="pone.0223444.t003" orientation="portrait" position="float"><object-id pub-id-type="doi">10.1371/journal.pone.0223444.t003</object-id><label>Table 3</label><caption><title>Summary of the effect of platelets on the markers of neutrophil function assessed in this study.</title></caption><alternatives><graphic id="pone.0223444.t003g" xlink:href="pone.0223444.t003"></graphic><table frame="hsides" rules="groups"><colgroup span="1"><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col><col align="left" valign="middle" span="1"></col></colgroup><thead><tr><th align="center" rowspan="1" colspan="1"></th><th align="center" rowspan="1" colspan="1"></th><th align="center" colspan="6" rowspan="1">+ platelets</th></tr><tr><th align="center" rowspan="1" colspan="1">Agonist</th><th align="center" rowspan="1" colspan="1">ng/mL</th><th align="center" rowspan="1" colspan="1">CD66b</th><th align="center" rowspan="1" colspan="1">CD62L</th><th align="center" rowspan="1" colspan="1">CD11b</th><th align="center" rowspan="1" colspan="1">Phagocytosis</th><th align="center" rowspan="1" colspan="1">Elastase</th><th align="center" rowspan="1" colspan="1">IL-8</th></tr></thead><tbody><tr><td align="center" rowspan="1" colspan="1">Unstimulated</td><td align="center" rowspan="1" colspan="1">--</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" rowspan="1" colspan="1"></td></tr><tr><td align="center" rowspan="1" colspan="1">LPS</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td></tr><tr><td align="center" rowspan="1" colspan="1">Pam3CSK4</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" rowspan="1" colspan="1"></td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td></tr><tr><td align="center" rowspan="1" colspan="1">FSL-1</td><td align="center" rowspan="1" colspan="1">1</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" rowspan="1" colspan="1"></td></tr><tr><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1">100</td><td align="center" rowspan="1" colspan="1"></td><td align="center" style="background-color:#8EAADB" rowspan="1" colspan="1">↑</td><td align="center" style="background-color:#FF0000" rowspan="1" colspan="1">↓</td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td><td align="center" rowspan="1" colspan="1"></td></tr></tbody></table></alternatives><table-wrap-foot><fn id="t003fn001"><p>Red shaded/↓, relative change was statistically significantly decreased with platelets; Blue shaded/↑, relative change in marker was statistically significantly increased with the addition of platelets.</p></fn></table-wrap-foot></table-wrap><p>This study was conducted to further investigate the findings of our previous study [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>], where we demonstrated that platelets regulate a number of leukocyte responses to TLR stimulation. Here, we have expanded on the number of neutrophil functions examined, and demonstrate that platelets are able to exert both stimulatory and dampening effects on these functions. These results contribute to a growing body of evidence indicating that platelets play a dual role in inflammation. While platelets have traditionally been considered pro-inflammatory [<xref rid="pone.0223444.ref021" ref-type="bibr">21</xref>, <xref rid="pone.0223444.ref030" ref-type="bibr">30</xref>], a number of studies [<xref rid="pone.0223444.ref015" ref-type="bibr">15</xref>, <xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>, <xref rid="pone.0223444.ref027" ref-type="bibr">27</xref>, <xref rid="pone.0223444.ref031" ref-type="bibr">31</xref>–<xref rid="pone.0223444.ref034" ref-type="bibr">34</xref>] indicate that platelets can limit inflammation by dampening leukocyte-mediated pro-inflammatory processes. Platelets are postulated to provide these cues to limit host damage that can occur during excessive inflammation, such as during sepsis [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>]. Thus, it is likely that the context plays an important role in determining the platelet response during inflammation.</p><p>The activation of neutrophils is widely associated with changes in expression of CD66b, CD11b and CD62L [<xref rid="pone.0223444.ref028" ref-type="bibr">28</xref>, <xref rid="pone.0223444.ref029" ref-type="bibr">29</xref>], and these markers function across a number of neutrophil responses including degranulation, transmigration and phagocytosis [<xref rid="pone.0223444.ref035" ref-type="bibr">35</xref>–<xref rid="pone.0223444.ref038" ref-type="bibr">38</xref>]. We show, here, that platelets can attenuate CD66b expression and this is consistent with our previous work [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>], where we show that platelets reduced CD66b expression (relative reductions of 14–19%) in response to stimulation with Pam3CSK4 and FSL-1. We also show that platelets similarly dampen neutrophil CD11b expression and CD62L shedding. These results are in agreement with the findings of Corken et al. [<xref rid="pone.0223444.ref039" ref-type="bibr">39</xref>], who demonstrated that the loss of a single platelet receptor, GPIb-IX, was sufficient to increase the level of neutrophil Mac-1 (CD11b/CD18) expression that occurs 24 hours post onset of sepsis in a mouse model of polymicrobial sepsis.</p><p>The expression of all three markers remained unchanged by platelets in response to LPS; platelets were only able to significantly alter neutrophil activation in response to Pam3CSK4 and FSL-1. It interesting to consider what drives a difference in the magnitude of the platelet effect in response to these TLR agonists. As both Pam3CSK4 and FSL-1 signal through TLR2 heterodimers, while LPS signals through TLR4, this may be due to differential platelet-TLR signal transduction. Platelet-TLR signal transduction has been most well-studied in response to Pam3CSK4, and platelet-TLR2/1 signalling has shown to induce Src- and Syk- family kinases [<xref rid="pone.0223444.ref040" ref-type="bibr">40</xref>, <xref rid="pone.0223444.ref041" ref-type="bibr">41</xref>], which mimics the signalling pathway of common haemostatic platelet receptors [<xref rid="pone.0223444.ref042" ref-type="bibr">42</xref>]. It may be that platelets can exert greater effects on neutrophils following stimulation with TLR2 agonists due to the ability to recruit or mimic haemostatic receptor pathways. However, comparing the differences in signalling induced by each TLR agonist has yet to be determined, and is an outstanding question in this field.</p><p>We show that platelets can enhance neutrophil phagocytosis both with and without TLR stimulation (low doses of Pam3CSK4 and FSL-1). These observations are consistent with prior reports: phagocytosis of periodontopathogens by neutrophils can be enhanced by platelets [<xref rid="pone.0223444.ref043" ref-type="bibr">43</xref>], and neutrophils within platelet-neutrophil complexes have increased phagocytic activity when compared to neutrophils that did not bind to platelets [<xref rid="pone.0223444.ref026" ref-type="bibr">26</xref>]. In our study, platelets did not further increase phagocytic activity under any other stimulation condition (high-dose TLR stimulation and low-dose LPS stimulation). In the absence of platelets, phagocytosis under these culture conditions was uniformly increased (65–70% of neutrophils were FITC+). One explanation for the absence of a platelet response here may be that the ‘maximum’ phagocytic activity seen under these conditions limits the ability of platelets to further enhance this activity.</p><p>Within this study, we also assessed the effect of platelets on elastase and IL-8 secretion by neutrophils. The platelet effect seen in response to low-dose TLR stimulation that we report in this study mirrors the results from our previous work [<xref rid="pone.0223444.ref016" ref-type="bibr">16</xref>], and is also in agreement with others [<xref rid="pone.0223444.ref026" ref-type="bibr">26</xref>]. Elastase is a potent serine protease which can degrade a multitude of plasma and extracellular matrix proteins. As such, extracellular elastase is a powerful bacterial killing mechanism [<xref rid="pone.0223444.ref044" ref-type="bibr">44</xref>] but has also been implicated in acute and chronic inflammatory host damage [<xref rid="pone.0223444.ref045" ref-type="bibr">45</xref>, <xref rid="pone.0223444.ref046" ref-type="bibr">46</xref>]. Platelets may dampen elastase secretion as a mechanism to prevent inflammatory host damage. Conversely, platelets modulated IL-8 secretion only in response to low-dose LPS. IL-8 is an important pro-inflammatory chemoattractant protein [<xref rid="pone.0223444.ref047" ref-type="bibr">47</xref>] and it may be that, under particular stimulation conditions, platelets can increase neutrophil IL-8 secretion and enhance further neutrophil chemoattraction to the site of inflammation. However, it is unclear why an effect of platelets was only observed under this condition. Examining a larger panel of cytokines and chemokines secreted is required to contextualize the significance of this platelet effect on IL-8 secretion.</p><p>The combination of the platelet effects described in this study suggest that platelets can both augment and attenuate neutrophil functions, and modulation of these functions can occur in both the absence and presence of TLR stimulation. For example, we show that platelets can simultaneously reduce elastase secretion and increase phagocytic activity in unstimulated cultures. Similarly, these opposing effects are seen following stimulation with Pam3CSK4 and FSL-1. The opposing effect of platelets noted here may be a mechanism for enhancing particular anti-microbial functions (for example, phagocytosis) while dampening neutrophil functions that are more likely to lead to host damage, if unchecked (for example, elastase secretion). Overall, we suggest that platelets finely regulate neutrophil functions, rather than providing broad anti-inflammatory cues to these cells, and we postulate that the platelet effect can modulate some aspects of the inflammatory environment to reduce host damage.</p><p>We also note that the platelet effect on these markers of neutrophil function was modulatory, rather than completely inhibitory. Neutrophils are highly reactive and short-lived cells that are crucial to anti-microbial host defence, but have also been implicated in causing inflammatory host damage [<xref rid="pone.0223444.ref048" ref-type="bibr">48</xref>, <xref rid="pone.0223444.ref049" ref-type="bibr">49</xref>]. Therefore, the inflammatory response must be rapid and robust, but also must be tightly controlled to prevent rampant inflammation. We suggest that platelets are regulators of neutrophil function and, within this role, platelets can act as a brake to neutrophil-mediated inflammation. In this regard, the platelet effect can be considered a mechanism for inflammation control.</p><p>We speculate, here, on how these results can be interpreted in the context of clinical pathologies that are characterized by platelet activation and inflammation. The newly-emerging role of platelets as immune regulators in sepsis [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>], cardiac ischaemia/reperfusion (I/R) injury [<xref rid="pone.0223444.ref017" ref-type="bibr">17</xref>, <xref rid="pone.0223444.ref020" ref-type="bibr">20</xref>] and acute lung injury (ALI) [<xref rid="pone.0223444.ref050" ref-type="bibr">50</xref>] is often juxtaposed against their well-established role in promoting leukocyte-mediated inflammation in these inflammatory diseases. An emerging working hypothesis is that platelets can elicit dual, potentially sequential, roles in inflammation [<xref rid="pone.0223444.ref011" ref-type="bibr">11</xref>, <xref rid="pone.0223444.ref020" ref-type="bibr">20</xref>]. Platelets can enhance leukocyte infiltration and inflammation to provide anti-microbial protection during infection or to restore haemostasis during sterile inflammation. Furthermore, platelets can then switch their phenotype to temper this inflammatory response to protect from host damage.</p><p>In the context of sepsis and septic shock, there has been particular focus on how platelets modulate their cytokine environment. Thrombocytopenia in septic patients [<xref rid="pone.0223444.ref051" ref-type="bibr">51</xref>] and platelet depletion in mouse models of sepsis [<xref rid="pone.0223444.ref018" ref-type="bibr">18</xref>, <xref rid="pone.0223444.ref052" ref-type="bibr">52</xref>] is associated with mortality, and the loss of platelets can drive an elevation in plasma pro-inflammatory cytokines (TNFα and IL-6) in septic mice [<xref rid="pone.0223444.ref018" ref-type="bibr">18</xref>, <xref rid="pone.0223444.ref053" ref-type="bibr">53</xref>, <xref rid="pone.0223444.ref054" ref-type="bibr">54</xref>]. We also add that, in this study, platelets can dampen neutrophilic inflammation in a stimulus-specific manner. Conversely, unregulated NET formation, propagated by platelet-neutrophil interactions [<xref rid="pone.0223444.ref002" ref-type="bibr">2</xref>], can cause significant host damage during sepsis by initiating intravascular thrombosis [<xref rid="pone.0223444.ref024" ref-type="bibr">24</xref>, <xref rid="pone.0223444.ref055" ref-type="bibr">55</xref>]. To add complexity, NETs have also been shown to degrade cytokines and chemokines and, consequently, reduce inflammation [<xref rid="pone.0223444.ref056" ref-type="bibr">56</xref>].</p><p>In a similar vein, platelets can exacerbate [<xref rid="pone.0223444.ref023" ref-type="bibr">23</xref>, <xref rid="pone.0223444.ref057" ref-type="bibr">57</xref>] and protect against [<xref rid="pone.0223444.ref058" ref-type="bibr">58</xref>, <xref rid="pone.0223444.ref059" ref-type="bibr">59</xref>] ALI. Platelets have been shown to mediate these opposing effects via their interaction with neutrophils. For example, platelet-neutrophil cross-talk is essential to early inflammatory responses in the lung [<xref rid="pone.0223444.ref023" ref-type="bibr">23</xref>, <xref rid="pone.0223444.ref057" ref-type="bibr">57</xref>], but these aggregates can also specifically produce and process the pro-resolving mediator, Maresin 1, to reduce lung inflammation [<xref rid="pone.0223444.ref059" ref-type="bibr">59</xref>]. The ability of platelets to propagate and resolve inflammation is particularly important in acute myocardial infarction (AMI) and I/R injury, where the resolution of inflammation in a timely manner is required for optimal myocardial healing and long-term repair [<xref rid="pone.0223444.ref060" ref-type="bibr">60</xref>]. In both conditions, platelets are known to have both pro-inflammatory [<xref rid="pone.0223444.ref017" ref-type="bibr">17</xref>, <xref rid="pone.0223444.ref061" ref-type="bibr">61</xref>] and pro-resolving [<xref rid="pone.0223444.ref020" ref-type="bibr">20</xref>, <xref rid="pone.0223444.ref062" ref-type="bibr">62</xref>] effects. In a clinical setting, anti-platelet agents are administered as a cornerstone of intervention in AMI. Although necessary to inhibit the pro-thrombotic effects of platelets during AMI, this treatment strategy may inhibit other, more subtle and perhaps beneficial, platelet immune effects. For example, the ability of platelets to control human neutrophil production of reactive oxygen species is reduced by administration of aspirin or clopidogrel [<xref rid="pone.0223444.ref015" ref-type="bibr">15</xref>]. In a similar vein, the platelet effect on neutrophil function that we report in this study may also be abolished by potent anti-platelet therapy.</p><p>To summarize, the immune functions of platelets are various and diverse. It is likely that the induction of a particular platelet response is a combination of the type of ligand present, the magnitude of the immune response and the type of platelet-leukocyte interaction occurring. In particular, platelets are known to elicit different responses to various agonists [<xref rid="pone.0223444.ref063" ref-type="bibr">63</xref>, <xref rid="pone.0223444.ref064" ref-type="bibr">64</xref>], which includes the differential release of immunomodulators from their stored granules [<xref rid="pone.0223444.ref065" ref-type="bibr">65</xref>, <xref rid="pone.0223444.ref066" ref-type="bibr">66</xref>]. This suggests an ability to elicit distinct pro-inflammatory or anti-inflammatory responses depending on the type of inflammatory stimulus received by the platelet population.</p><p>Our study had a number of limitations. The methodology was designed to minimize neutrophil activation prior to TLR stimulation, but we cannot be certain that the activation state was not altered by the isolation process. Neutrophils and platelets do not act in isolation and it may be that, within whole blood, the modulation of neutrophil function by platelets differs Additionally, although Pam3CSK4 shows 98.1% purity, we cannot guarantee that this reagent did not contain any contaminants with TLR agonist activity. Finally, we did not assess whether the effect of platelets was directly or indirectly mediated, rather we aimed to examine the holistic platelet effects in this study.</p></sec><sec sec-type="conclusions" id="sec016"><title>Conclusion</title><p>In conclusion, the combination of the platelet effects described in this study suggest that platelets can both augment and attenuate neutrophil functions, and modulation of these functions can occur in both the absence and presence of TLR stimulation. We suggest that platelets regulate neutrophil function in a complex manner, rather than providing broad anti-inflammatory cues to these cells. This platelet effect may modulate some aspects of the inflammatory environment to reduce host damage.</p></sec><sec sec-type="supplementary-material" id="sec017"><title>Supporting information</title><supplementary-material content-type="local-data" id="pone.0223444.s001"><label>S1 Fig</label><caption><title>Platelets remain quiescent both pre- and post-culture.</title><p>Platelets were isolated and probed for activation post-isolation and prior to culture (pre-culture; dashed line). Platelets were then cultured alone in the absence of TLR stimulation for 4 hours (post-culture; solid + filled line), and also probed for activation. To detect activation, platelets were incubated with PAC1-FITC (clone PAC-1, Becton Dickinson) for 30 minutes at 4°C, fixed and analysed. PAC1 expression under both conditions was minimal, when compared to unstained platelets (dot-dashed line). PAC1 is a common platelet activation marker that recognizes the activation-dependent glycoprotein IIb/IIIa complex on the platelet surface.</p><p>(TIF)</p></caption><media xlink:href="pone.0223444.s001.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s002"><label>S2 Fig</label><caption><title>Gating strategy to identify neutrophils after 4-hour culture.</title><p>Initially, all samples were visualised by SSC-A vs. time to check the flow of cells during acquisition. Doublet exclusion was performed firstly by examining the SSC-A vs. SSC-H profile (A) and secondly by examining the FSC-A vs. FSC-H profile (B). The resulting SSC-A vs. FSC-A profile of the population is shown in (C). Neutrophils were identified as cells that were highly expressing CD16 (D) against the isotype control (E). These representative plots are from a sample of unstimulated neutrophils, and we did not observe a visual difference between unstimulated and stimulated neutrophils using these plots.</p><p>(TIF)</p></caption><media xlink:href="pone.0223444.s002.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s003"><label>S3 Fig</label><caption><title>Platelets become dimly positive for PAC1 following incubation with TLR agonists.</title><p>Platelets were isolated and cultured alone (dotted line) or with neutrophils (dashed line) for 4 hours in the presence of 100 ng/mL of either LPS (A), Pam3CSK4 (B) or FSL-1 (C). Platelets were stained for PAC1-FITC as described in <xref ref-type="supplementary-material" rid="pone.0223444.s002">S2 Fig</xref>. PAC1 expression under these stimulation conditions was compared to unstimulated platelets (solid + filled line). The shift in PAC1 expression with TLR stimulation was more pronounced for LPS and FSL-1, compared to Pam3CSK4. The same trend was observed when platelets were stimulated with 1 ng/mL of all TLR agonists (data not shown).</p><p>(TIF)</p></caption><media xlink:href="pone.0223444.s003.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s004"><label>S1 Table</label><caption><title>Raw neutrophil CD66b expression ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s004.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s005"><label>S2 Table</label><caption><title>Raw neutrophil CD62L expression ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s005.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s006"><label>S3 Table</label><caption><title>Raw neutrophil CD11b expression ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s006.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s007"><label>S4 Table</label><caption><title>Raw neutrophil phagocytosis ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s007.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s008"><label>S5 Table</label><caption><title>Raw neutrophil elastase secretion ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s008.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material><supplementary-material content-type="local-data" id="pone.0223444.s009"><label>S6 Table</label><caption><title>Raw neutrophil IL-8 secretion ± platelet co-culture.</title><p>(TIF)</p></caption><media xlink:href="pone.0223444.s009.tif"><caption><p>Click here for additional data file.</p></caption></media></supplementary-material></sec></body><back><glossary><title>Abbreviations</title><def-list><def-item><term>FSL-1</term><def><p>Fibroblast-stimulating lipopeptide -1</p></def></def-item><def-item><term>LPS</term><def><p>Lipopolysaccharide</p></def></def-item><def-item><term>NETs</term><def><p>Neutrophil extracellular traps</p></def></def-item><def-item><term>PPP</term><def><p>Platelet-poor plasma</p></def></def-item><def-item><term>PRP</term><def><p>Platelet-rich plasma</p></def></def-item><def-item><term>ROS</term><def><p>Reactive oxygen species</p></def></def-item><def-item><term>TLR</term><def><p>Toll-like receptors</p></def></def-item><def-item><term>WPs</term><def><p>Washed platelets</p></def></def-item></def-list></glossary><ref-list><title>References</title><ref id="pone.0223444.ref001"><label>1</label><mixed-citation publication-type="journal"><name><surname>Yeaman</surname><given-names>MR</given-names></name>. <article-title>Platelets: at the nexus of antimicrobial defence</article-title>. <source><italic toggle="yes">Nature Reviews Microbiology</italic></source>. <year>2014</year>; <volume>12</volume>: <fpage>426</fpage>–<lpage>37</lpage>. <pub-id pub-id-type="doi">10.1038/nrmicro3269</pub-id><pub-id pub-id-type="pmid">24830471</pub-id></mixed-citation></ref><ref id="pone.0223444.ref002"><label>2</label><mixed-citation publication-type="journal"><name><surname>Clark</surname><given-names>SR</given-names></name>, <name><surname>Ma</surname><given-names>AC</given-names></name>, <name><surname>Tavener</surname><given-names>SA</given-names></name>, <name><surname>McDonald</surname><given-names>B</given-names></name>, <name><surname>Goodarzi</surname><given-names>Z</given-names></name>, <name><surname>Kelly</surname><given-names>MM</given-names></name>, <etal>et al</etal><article-title>Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood</article-title>. <source><italic toggle="yes">Nat Med</italic></source>. <year>2007</year>; <volume>13</volume>: <fpage>463</fpage>–<lpage>9</lpage>. <pub-id pub-id-type="doi">10.1038/nm1565</pub-id><pub-id pub-id-type="pmid">17384648</pub-id></mixed-citation></ref><ref id="pone.0223444.ref003"><label>3</label><mixed-citation publication-type="journal"><name><surname>Verschoor</surname><given-names>A</given-names></name>, <name><surname>Neuenhahn</surname><given-names>M</given-names></name>, <name><surname>Navarini</surname><given-names>AA</given-names></name>, <name><surname>Graef</surname><given-names>P</given-names></name>, <name><surname>Plaumann</surname><given-names>A</given-names></name>, <name><surname>Seidlmeier</surname><given-names>A</given-names></name>, <etal>et al</etal><article-title>A platelet-mediated system for shuttling blood-borne bacteria to CD8[alpha]+ dendritic cells depends on glycoprotein GPIb and complement C3</article-title>. <source><italic toggle="yes">Nat Immunol</italic></source>. <year>2011</year>; <volume>12</volume>: <fpage>1194</fpage>–<lpage>201</lpage>. <pub-id pub-id-type="doi">10.1038/ni.2140</pub-id><pub-id pub-id-type="pmid">22037602</pub-id></mixed-citation></ref><ref id="pone.0223444.ref004"><label>4</label><mixed-citation publication-type="journal"><name><surname>Cognasse</surname><given-names>F</given-names></name>, <name><surname>Hamzeh</surname><given-names>H</given-names></name>, <name><surname>Chavarin</surname><given-names>P</given-names></name>, <name><surname>Acquart</surname><given-names>S</given-names></name>, <name><surname>Genin</surname><given-names>C</given-names></name>, <name><surname>Garraud</surname><given-names>O</given-names></name>. <article-title>Evidence of Toll-like receptor molecules on human platelets</article-title>. <source><italic toggle="yes">Immunol Cell Biol</italic></source>. <year>2005</year>; <volume>83</volume>: <fpage>196</fpage>–<lpage>8</lpage>. <pub-id pub-id-type="doi">10.1111/j.1440-1711.2005.01314.x</pub-id><pub-id pub-id-type="pmid">15748217</pub-id></mixed-citation></ref><ref id="pone.0223444.ref005"><label>5</label><mixed-citation publication-type="journal"><name><surname>Shiraki</surname><given-names>R</given-names></name>, <name><surname>Inoue</surname><given-names>N</given-names></name>, <name><surname>Kawasaki</surname><given-names>S</given-names></name>, <name><surname>Takei</surname><given-names>A</given-names></name>, <name><surname>Kadotani</surname><given-names>M</given-names></name>, <name><surname>Ohnishi</surname><given-names>Y</given-names></name>, <etal>et al</etal><article-title>Expression of Toll-like receptors on human platelets</article-title>. <source><italic toggle="yes">Thromb Res</italic></source>. <year>2004</year>; <volume>113</volume>: <fpage>379</fpage>–<lpage>85</lpage>. <pub-id pub-id-type="doi">10.1016/j.thromres.2004.03.023</pub-id><pub-id pub-id-type="pmid">15226092</pub-id></mixed-citation></ref><ref id="pone.0223444.ref006"><label>6</label><mixed-citation publication-type="journal"><name><surname>Hally</surname><given-names>KE</given-names></name>, <name><surname>La Flamme</surname><given-names>AC</given-names></name>, <name><surname>Larsen</surname><given-names>PD</given-names></name>, <name><surname>Harding</surname><given-names>SA</given-names></name>. <article-title>Platelet Toll-like receptor (TLR) expression and TLR-mediated platelet activation in acute myocardial infarction</article-title>. <source><italic toggle="yes">Thromb Res</italic></source>. <year>2017</year>; <volume>158</volume>: <fpage>8</fpage>–<lpage>15</lpage>. <pub-id pub-id-type="doi">10.1016/j.thromres.2017.07.031</pub-id><pub-id pub-id-type="pmid">28783513</pub-id></mixed-citation></ref><ref id="pone.0223444.ref007"><label>7</label><mixed-citation publication-type="journal"><name><surname>Blair</surname><given-names>P</given-names></name>, <name><surname>Rex</surname><given-names>S</given-names></name>, <name><surname>Vitseva</surname><given-names>O</given-names></name>, <name><surname>Beaulieu</surname><given-names>L</given-names></name>, <name><surname>Tanriverdi</surname><given-names>K</given-names></name>, <name><surname>Chakrabarti</surname><given-names>S</given-names></name>, <etal>et al</etal><article-title>Stimulation of Toll-like receptor 2 in human platelets induces a thromboinflammatory response through activation of phosphoinositide 3-kinase</article-title>. <source><italic toggle="yes">Circ Res</italic></source>. <year>2009</year>; <volume>104</volume>: <fpage>346</fpage>–<lpage>54</lpage>. <pub-id pub-id-type="doi">10.1161/CIRCRESAHA.108.185785</pub-id><pub-id pub-id-type="pmid">19106411</pub-id></mixed-citation></ref><ref id="pone.0223444.ref008"><label>8</label><mixed-citation publication-type="journal"><name><surname>Rivadeneyra</surname><given-names>L</given-names></name>, <name><surname>Carestia</surname><given-names>A</given-names></name>, <name><surname>Etulain</surname><given-names>J</given-names></name>, <name><surname>Pozner</surname><given-names>RG</given-names></name>, <name><surname>Fondevila</surname><given-names>C</given-names></name>, <name><surname>Negrotto</surname><given-names>S</given-names></name>, <etal>et al</etal><article-title>Regulation of platelet responses triggered by Toll-like receptor 2 and 4 ligands is another non-genomic role of nuclear factor-kappaB</article-title>. <source><italic toggle="yes">Thromb Res</italic></source>. <year>2014</year>; <volume>133</volume>: <fpage>235</fpage>–<lpage>43</lpage>. <pub-id pub-id-type="doi">10.1016/j.thromres.2013.11.028</pub-id><pub-id pub-id-type="pmid">24331207</pub-id></mixed-citation></ref><ref id="pone.0223444.ref009"><label>9</label><mixed-citation publication-type="journal"><name><surname>Zhang</surname><given-names>G</given-names></name>, <name><surname>Han</surname><given-names>J</given-names></name>, <name><surname>Welch</surname><given-names>EJ</given-names></name>, <name><surname>Ye</surname><given-names>RD</given-names></name>, <name><surname>Voyno-Yasenetskaya</surname><given-names>TA</given-names></name>, <name><surname>Malik</surname><given-names>AB</given-names></name>, <etal>et al</etal><article-title>Lipopolysaccharide stimulates platelet secretion and potentiates platelet aggregation via TLR4/MyD88 and the cGMP-dependent protein kinase pathway</article-title>. <source><italic toggle="yes">J Immunol</italic></source>. <year>2009</year>; <volume>182</volume>: <fpage>7997</fpage>–<lpage>8004</lpage>. <pub-id pub-id-type="doi">10.4049/jimmunol.0802884</pub-id><pub-id pub-id-type="pmid">19494325</pub-id></mixed-citation></ref><ref id="pone.0223444.ref010"><label>10</label><mixed-citation publication-type="journal"><name><surname>Hally</surname><given-names>KE</given-names></name>, <name><surname>La Flamme</surname><given-names>AC</given-names></name>, <name><surname>Harding</surname><given-names>SA</given-names></name>, <name><surname>Larsen</surname><given-names>PD</given-names></name>. <article-title>The effects of aspirin and ticagrelor on Toll-like receptor (TLR)-mediated platelet activation: results of a randomized, cross-over trial.</article-title><source><italic toggle="yes">Platelets</italic></source>. <year>2018</year>: <fpage>1</fpage>–<lpage>9</lpage>.</mixed-citation></ref><ref id="pone.0223444.ref011"><label>11</label><mixed-citation publication-type="journal"><name><surname>Stocker</surname><given-names>TJ</given-names></name>, <name><surname>Ishikawa-Ankerhold</surname><given-names>H</given-names></name>, <name><surname>Massberg</surname><given-names>S</given-names></name>, <name><surname>Schulz</surname><given-names>C</given-names></name>. <article-title>Small but mighty: Platelets as central effectors of host defense</article-title>. <source><italic toggle="yes">Thromb Haemost</italic></source>. <year>2017</year>; <volume>117</volume>: <fpage>651</fpage>–<lpage>61</lpage>. <pub-id pub-id-type="doi">10.1160/TH16-12-0921</pub-id><pub-id pub-id-type="pmid">28203681</pub-id></mixed-citation></ref><ref id="pone.0223444.ref012"><label>12</label><mixed-citation publication-type="journal"><name><surname>Ståhl</surname><given-names>A-l</given-names></name>, <name><surname>Svensson</surname><given-names>M</given-names></name>, <name><surname>Mörgelin</surname><given-names>M</given-names></name>, <name><surname>Svanborg</surname><given-names>C</given-names></name>, <name><surname>Tarr</surname><given-names>PI</given-names></name>, <name><surname>Mooney</surname><given-names>JC</given-names></name>, <etal>et al</etal><article-title>Lipopolysaccharide from enterohemorrhagic Escherichia coli binds to platelets through TLR4 and CD62 and is detected on circulating platelets in patients with hemolytic uremic syndrome</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2006</year>; <volume>108</volume>: <fpage>167</fpage>–<lpage>76</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2005-08-3219</pub-id><pub-id pub-id-type="pmid">16514062</pub-id></mixed-citation></ref><ref id="pone.0223444.ref013"><label>13</label><mixed-citation publication-type="journal"><name><surname>Linke</surname><given-names>B</given-names></name>, <name><surname>Schreiber</surname><given-names>Y</given-names></name>, <name><surname>Picard-Willems</surname><given-names>B</given-names></name>, <name><surname>Slattery</surname><given-names>P</given-names></name>, <name><surname>Nusing</surname><given-names>RM</given-names></name>, <name><surname>Harder</surname><given-names>S</given-names></name>, <etal>et al</etal><article-title>Activated Platelets Induce an Anti-Inflammatory Response of Monocytes/Macrophages through Cross-Regulation of PGE2 and Cytokines</article-title>. <source><italic toggle="yes">Mediators Inflamm</italic></source>. <year>2017</year>; <volume>2017</volume>: <fpage>1463216</fpage><pub-id pub-id-type="doi">10.1155/2017/1463216</pub-id><pub-id pub-id-type="pmid">28592915</pub-id></mixed-citation></ref><ref id="pone.0223444.ref014"><label>14</label><mixed-citation publication-type="journal"><name><surname>Jancinova</surname><given-names>V</given-names></name>, <name><surname>Drabikova</surname><given-names>K</given-names></name>, <name><surname>Petrikova</surname><given-names>M</given-names></name>, <name><surname>Nosal</surname><given-names>R</given-names></name>. <article-title>Blood platelets decrease concentration of reactive oxygen species produced by polymorphonuclear leukocytes</article-title>. <source><italic toggle="yes">Bratisl Lek Listy</italic></source>. <year>2004</year>; <volume>105</volume>: <fpage>250</fpage>–<lpage>5</lpage>. <pub-id pub-id-type="pmid">15543845</pub-id></mixed-citation></ref><ref id="pone.0223444.ref015"><label>15</label><mixed-citation publication-type="journal"><name><surname>Reinisch</surname><given-names>CM</given-names></name>, <name><surname>Dunzendorfer</surname><given-names>S</given-names></name>, <name><surname>Pechlaner</surname><given-names>C</given-names></name>, <name><surname>Ricevuti</surname><given-names>G</given-names></name>, <name><surname>Wiedermann</surname><given-names>CJ</given-names></name>. <article-title>The inhibition of oxygen radical release from human neutrophils by resting platelets is reversed by administration of acetylsalicylic acid or clopidogrel</article-title>. <source><italic toggle="yes">Free Radic Res</italic></source>. <year>2001</year>; <volume>34</volume>: <fpage>461</fpage>–<lpage>6</lpage>. <pub-id pub-id-type="doi">10.1080/10715760100300401</pub-id><pub-id pub-id-type="pmid">11378529</pub-id></mixed-citation></ref><ref id="pone.0223444.ref016"><label>16</label><mixed-citation publication-type="journal"><name><surname>Hally</surname><given-names>KE</given-names></name>, <name><surname>La Flamme</surname><given-names>AC</given-names></name>, <name><surname>Harding</surname><given-names>SA</given-names></name>, <name><surname>Larsen</surname><given-names>PD</given-names></name>. <article-title>Platelets regulate leucocyte responses to Toll-like receptor stimulation.</article-title><source>Clinical & Translational <italic toggle="yes">Immunology</italic></source>. <year>2018</year>; <volume>7</volume>: <fpage>e1036</fpage>.<pub-id pub-id-type="pmid">30065836</pub-id></mixed-citation></ref><ref id="pone.0223444.ref017"><label>17</label><mixed-citation publication-type="journal"><name><surname>Ziegler</surname><given-names>M</given-names></name>, <name><surname>Wang</surname><given-names>X</given-names></name>, <name><surname>Peter</surname><given-names>K</given-names></name>. <article-title>Platelets in cardiac ischaemia/reperfusion injury: a promising therapeutic target</article-title>. <source><italic toggle="yes">Cardiovasc Res</italic></source>. <year>2019</year>; <volume>115</volume>: <fpage>1178</fpage>–<lpage>88</lpage>. <pub-id pub-id-type="doi">10.1093/cvr/cvz070</pub-id><pub-id pub-id-type="pmid">30906948</pub-id></mixed-citation></ref><ref id="pone.0223444.ref018"><label>18</label><mixed-citation publication-type="journal"><name><surname>Xiang</surname><given-names>B</given-names></name>, <name><surname>Zhang</surname><given-names>G</given-names></name>, <name><surname>Guo</surname><given-names>L</given-names></name>, <name><surname>Li</surname><given-names>XA</given-names></name>, <name><surname>Morris</surname><given-names>AJ</given-names></name>, <name><surname>Daugherty</surname><given-names>A</given-names></name>, <etal>et al</etal><article-title>Platelets protect from septic shock by inhibiting macrophage-dependent inflammation via the cyclooxygenase 1 signalling pathway</article-title>. <source><italic toggle="yes">Nature Communications</italic></source>. <year>2013</year>; <volume>4</volume>: <fpage>2657</fpage><pub-id pub-id-type="doi">10.1038/ncomms3657</pub-id><pub-id pub-id-type="pmid">24150174</pub-id></mixed-citation></ref><ref id="pone.0223444.ref019"><label>19</label><mixed-citation publication-type="journal"><name><surname>Bain</surname><given-names>W</given-names></name>, <name><surname>Olonisakin</surname><given-names>T</given-names></name>, <name><surname>Yu</surname><given-names>M</given-names></name>, <name><surname>Qu</surname><given-names>Y</given-names></name>, <name><surname>Hulver</surname><given-names>M</given-names></name>, <name><surname>Xiong</surname><given-names>Z</given-names></name>, <etal>et al</etal><article-title>Platelets inhibit apoptotic lung epithelial cell death and protect mice against infection-induced lung injury</article-title>. <source><italic toggle="yes">Blood Advances</italic></source>. <year>2019</year>; <volume>3</volume>: <fpage>432</fpage>–<lpage>45</lpage>. <pub-id pub-id-type="doi">10.1182/bloodadvances.2018026286</pub-id><pub-id pub-id-type="pmid">30733303</pub-id></mixed-citation></ref><ref id="pone.0223444.ref020"><label>20</label><mixed-citation publication-type="journal"><name><surname>Walsh</surname><given-names>TG</given-names></name>, <name><surname>Poole</surname><given-names>AW</given-names></name>. <article-title>Do platelets promote cardiac recovery after myocardial infarction: roles beyond occlusive ischemic damage</article-title>. <source><italic toggle="yes">Am J Physiol Heart Circ Physiol</italic></source>. <year>2018</year>; <volume>314</volume>: <fpage>H1043</fpage>–<lpage>h8</lpage>. <pub-id pub-id-type="doi">10.1152/ajpheart.00134.2018</pub-id><pub-id pub-id-type="pmid">29547023</pub-id></mixed-citation></ref><ref id="pone.0223444.ref021"><label>21</label><mixed-citation publication-type="journal"><name><surname>Gros</surname><given-names>A</given-names></name>, <name><surname>Ollivier</surname><given-names>V</given-names></name>, <name><surname>Ho-Tin-Noe</surname><given-names>B</given-names></name>. <article-title>Platelets in inflammation: regulation of leukocyte activities and vascular repair</article-title>. <source><italic toggle="yes">Front Immunol</italic></source>. <year>2014</year>; <volume>5</volume>: <fpage>678</fpage><pub-id pub-id-type="doi">10.3389/fimmu.2014.00678</pub-id><pub-id pub-id-type="pmid">25610439</pub-id></mixed-citation></ref><ref id="pone.0223444.ref022"><label>22</label><mixed-citation publication-type="journal"><name><surname>Assinger</surname><given-names>A</given-names></name>, <name><surname>Laky</surname><given-names>M</given-names></name>, <name><surname>Badrnya</surname><given-names>S</given-names></name>, <name><surname>Esfandeyari</surname><given-names>A</given-names></name>, <name><surname>Volf</surname><given-names>I</given-names></name>. <article-title>Periodontopathogens induce expression of CD40L on human platelets via TLR2 and TLR4</article-title>. <source><italic toggle="yes">Thromb Res</italic></source>. <year>2012</year>; <volume>130</volume>: <fpage>e73</fpage>–<lpage>e8</lpage>. <pub-id pub-id-type="doi">10.1016/j.thromres.2012.04.017</pub-id><pub-id pub-id-type="pmid">22608210</pub-id></mixed-citation></ref><ref id="pone.0223444.ref023"><label>23</label><mixed-citation publication-type="journal"><name><surname>Zarbock</surname><given-names>A</given-names></name>, <name><surname>Singbartl</surname><given-names>K</given-names></name>, <name><surname>Ley</surname><given-names>K</given-names></name>. <article-title>Complete reversal of acid-induced acute lung injury by blocking of platelet-neutrophil aggregation</article-title>. <source><italic toggle="yes">J Clin Invest</italic></source>. <year>2006</year>; <volume>116</volume>: <fpage>3211</fpage>–<lpage>9</lpage>. <pub-id pub-id-type="doi">10.1172/JCI29499</pub-id><pub-id pub-id-type="pmid">17143330</pub-id></mixed-citation></ref><ref id="pone.0223444.ref024"><label>24</label><mixed-citation publication-type="journal"><name><surname>McDonald</surname><given-names>B</given-names></name>, <name><surname>Davis</surname><given-names>RP</given-names></name>, <name><surname>Kim</surname><given-names>SJ</given-names></name>, <name><surname>Tse</surname><given-names>M</given-names></name>, <name><surname>Esmon</surname><given-names>CT</given-names></name>, <name><surname>Kolaczkowska</surname><given-names>E</given-names></name>, <etal>et al</etal><article-title>Platelets and neutrophil extracellular traps collaborate to promote intravascular coagulation during sepsis in mice</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2017</year>; <volume>129</volume>: <fpage>1357</fpage>–<lpage>67</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2016-09-741298</pub-id><pub-id pub-id-type="pmid">28073784</pub-id></mixed-citation></ref><ref id="pone.0223444.ref025"><label>25</label><mixed-citation publication-type="journal"><name><surname>Herbertsson</surname><given-names>H</given-names></name>, <name><surname>Bengtsson</surname><given-names>T</given-names></name>. <article-title>Role of platelets and the arachidonic acid pathway in the regulation of neutrophil oxidase activity</article-title>. <source><italic toggle="yes">Scand J Clin Lab Invest</italic></source>. <year>2001</year>; <volume>61</volume>: <fpage>641</fpage>–<lpage>9</lpage>. <pub-id pub-id-type="doi">10.1080/003655101753268008</pub-id><pub-id pub-id-type="pmid">11768324</pub-id></mixed-citation></ref><ref id="pone.0223444.ref026"><label>26</label><mixed-citation publication-type="journal"><name><surname>Losche</surname><given-names>W</given-names></name>, <name><surname>Dressel</surname><given-names>M</given-names></name>, <name><surname>Krause</surname><given-names>S</given-names></name>, <name><surname>Redlich</surname><given-names>H</given-names></name>, <name><surname>Spangenberg</surname><given-names>P</given-names></name>, <name><surname>Heptinstall</surname><given-names>S</given-names></name>. <article-title>Contact-induced modulation of neutrophil elastase secretion and phagocytic activity by platelets</article-title>. <source><italic toggle="yes">Blood Coagul Fibrinolysis</italic></source>. <year>1996</year>; <volume>7</volume>: <fpage>210</fpage>–<lpage>3</lpage>. <pub-id pub-id-type="doi">10.1097/00001721-199603000-00025</pub-id><pub-id pub-id-type="pmid">8735821</pub-id></mixed-citation></ref><ref id="pone.0223444.ref027"><label>27</label><mixed-citation publication-type="journal"><name><surname>Del Principe</surname><given-names>D</given-names></name>, <name><surname>Menichelli</surname><given-names>A</given-names></name>, <name><surname>Di Giulio</surname><given-names>S</given-names></name>, <name><surname>De Matteis</surname><given-names>W</given-names></name>, <name><surname>Giordani</surname><given-names>M</given-names></name>, <name><surname>Pentassuglio</surname><given-names>AM</given-names></name>, <etal>et al</etal><article-title>Stimulated platelets release factor(s) affecting the in vitro response of human polymorphonuclear cells</article-title>. <source><italic toggle="yes">J Leukoc Biol</italic></source>. <year>1990</year>; <volume>48</volume>: <fpage>7</fpage>–<lpage>14</lpage>. <pub-id pub-id-type="doi">10.1002/jlb.48.1.7</pub-id><pub-id pub-id-type="pmid">2113564</pub-id></mixed-citation></ref><ref id="pone.0223444.ref028"><label>28</label><mixed-citation publication-type="journal"><name><surname>Fortunati</surname><given-names>E</given-names></name>, <name><surname>Kazemier</surname><given-names>KM</given-names></name>, <name><surname>Grutters</surname><given-names>JC</given-names></name>, <name><surname>Koenderman</surname><given-names>L</given-names></name>, <article-title>Van den Bosch vJMM. Human neutrophils switch to an activated phenotype after homing to the lung irrespective of inflammatory disease</article-title>. <source><italic toggle="yes">Clin Exp Immunol</italic></source>. <year>2009</year>; <volume>155</volume>: <fpage>559</fpage>–<lpage>66</lpage>. <pub-id pub-id-type="doi">10.1111/j.1365-2249.2008.03791.x</pub-id><pub-id pub-id-type="pmid">19077082</pub-id></mixed-citation></ref><ref id="pone.0223444.ref029"><label>29</label><mixed-citation publication-type="journal"><name><surname>van Oostrom</surname><given-names>AJ</given-names></name>, <name><surname>van Wijk</surname><given-names>JP</given-names></name>, <name><surname>Sijmonsma</surname><given-names>TP</given-names></name>, <name><surname>Rabelink</surname><given-names>TJ</given-names></name>, <name><surname>Castro Cabezas</surname><given-names>M</given-names></name>. <article-title>Increased expression of activation markers on monocytes and neutrophils in type 2 diabetes</article-title>. <source><italic toggle="yes">The Netherlands Journal of Medicine</italic></source>. <year>2004</year>; <volume>62</volume>: <fpage>320</fpage>–<lpage>5</lpage>. <pub-id pub-id-type="pmid">15635816</pub-id></mixed-citation></ref><ref id="pone.0223444.ref030"><label>30</label><mixed-citation publication-type="journal"><name><surname>Stokes</surname><given-names>KY</given-names></name>, <name><surname>Granger</surname><given-names>DN</given-names></name>. <article-title>Platelets: a critical link between inflammation and microvascular dysfunction</article-title>. <source><italic toggle="yes">The Journal of Physiology</italic></source>. <year>2012</year>; <volume>590</volume>: <fpage>1023</fpage>–<lpage>34</lpage>. <pub-id pub-id-type="doi">10.1113/jphysiol.2011.225417</pub-id><pub-id pub-id-type="pmid">22183721</pub-id></mixed-citation></ref><ref id="pone.0223444.ref031"><label>31</label><mixed-citation publication-type="journal"><name><surname>Gudbrandsdottir</surname><given-names>S</given-names></name>, <name><surname>Hasselbalch</surname><given-names>HC</given-names></name>, <name><surname>Nielsen</surname><given-names>CH</given-names></name>. <article-title>Activated platelets enhance IL-10 secretion and reduce TNF-alpha secretion by monocytes</article-title>. <source><italic toggle="yes">J Immunol</italic></source>. <year>2013</year>; <volume>191</volume>: <fpage>4059</fpage>–<lpage>67</lpage>. <pub-id pub-id-type="doi">10.4049/jimmunol.1201103</pub-id><pub-id pub-id-type="pmid">24048901</pub-id></mixed-citation></ref><ref id="pone.0223444.ref032"><label>32</label><mixed-citation publication-type="journal"><name><surname>Nami</surname><given-names>N</given-names></name>, <name><surname>Feci</surname><given-names>L</given-names></name>, <name><surname>Napoliello</surname><given-names>L</given-names></name>, <name><surname>Giordano</surname><given-names>A</given-names></name>, <name><surname>Lorenzini</surname><given-names>S</given-names></name>, <name><surname>Galeazzi</surname><given-names>M</given-names></name>, <etal>et al</etal><article-title>Crosstalk between platelets and PBMC: New evidence in wound healing</article-title>. <source><italic toggle="yes">Platelets</italic></source>. <year>2016</year>; <volume>27</volume>: <fpage>143</fpage>–<lpage>8</lpage>. <pub-id pub-id-type="doi">10.3109/09537104.2015.1048216</pub-id><pub-id pub-id-type="pmid">26030799</pub-id></mixed-citation></ref><ref id="pone.0223444.ref033"><label>33</label><mixed-citation publication-type="journal"><name><surname>Sadallah</surname><given-names>S</given-names></name>, <name><surname>Eken</surname><given-names>C</given-names></name>, <name><surname>Martin</surname><given-names>PJ</given-names></name>, <name><surname>Schifferli</surname><given-names>JA</given-names></name>. <article-title>Microparticles (ectosomes) shed by stored human platelets downregulate macrophages and modify the development of dendritic cells</article-title>. <source><italic toggle="yes">J Immunol</italic></source>. <year>2011</year>; <volume>186</volume>: <fpage>6543</fpage>–<lpage>52</lpage>. <pub-id pub-id-type="doi">10.4049/jimmunol.1002788</pub-id><pub-id pub-id-type="pmid">21525379</pub-id></mixed-citation></ref><ref id="pone.0223444.ref034"><label>34</label><mixed-citation publication-type="journal"><name><surname>Jancinova</surname><given-names>V</given-names></name>, <name><surname>Drabikova</surname><given-names>K</given-names></name>, <name><surname>Nosal</surname><given-names>R</given-names></name>, <name><surname>Petrikova</surname><given-names>M</given-names></name>, <name><surname>Ciz</surname><given-names>M</given-names></name>, <name><surname>Lojek</surname><given-names>A</given-names></name>, <etal>et al</etal><article-title>Inhibition of FMLP-stimulated neutrophil chemiluminescence by blood platelets increased in the presence of the serotonin-liberating drug chloroquine</article-title>. <source><italic toggle="yes">Thromb Res</italic></source>. <year>2003</year>; <volume>109</volume>: <fpage>293</fpage>–<lpage>8</lpage>. <pub-id pub-id-type="doi">10.1016/s0049-3848(03)00239-1</pub-id><pub-id pub-id-type="pmid">12818253</pub-id></mixed-citation></ref><ref id="pone.0223444.ref035"><label>35</label><mixed-citation publication-type="journal"><name><surname>Deree</surname><given-names>J</given-names></name>, <name><surname>Lall</surname><given-names>R</given-names></name>, <name><surname>Melbostad</surname><given-names>H</given-names></name>, <name><surname>Grant</surname><given-names>M</given-names></name>, <name><surname>Hoyt</surname><given-names>DB</given-names></name>, <name><surname>Coimbra</surname><given-names>R</given-names></name>. <article-title>Neutrophil degranulation and the effects of phosphodiesterase inhibition</article-title>. <source><italic toggle="yes">J Surg Res</italic></source>. <year>2006</year>; <volume>133</volume>: <fpage>22</fpage>–<lpage>8</lpage>. <pub-id pub-id-type="doi">10.1016/j.jss.2006.02.031</pub-id><pub-id pub-id-type="pmid">16690368</pub-id></mixed-citation></ref><ref id="pone.0223444.ref036"><label>36</label><mixed-citation publication-type="journal"><name><surname>Lacy</surname><given-names>P</given-names></name>. <article-title>Mechanisms of Degranulation in Neutrophils.</article-title><source>Allergy, Asthma, and Clinical Immunology: Official Journal of the Canadian Society of Allergy and Clinical <italic toggle="yes">Immunology</italic></source>. <year>2006</year>; <volume>2</volume>: <fpage>98</fpage>–<lpage>108</lpage>.</mixed-citation></ref><ref id="pone.0223444.ref037"><label>37</label><mixed-citation publication-type="journal"><name><surname>Le Cabec</surname><given-names>V</given-names></name>, <name><surname>Carreno</surname><given-names>S</given-names></name>, <name><surname>Moisand</surname><given-names>A</given-names></name>, <name><surname>Bordier</surname><given-names>C</given-names></name>, <name><surname>Maridonneau-Parini</surname><given-names>I</given-names></name>. <article-title>Complement receptor 3 (CD11b/CD18) mediates type I and type II phagocytosis during nonopsonic and opsonic phagocytosis, respectively</article-title>. <source><italic toggle="yes">J Immunol</italic></source>. <year>2002</year>; <volume>169</volume>: <fpage>2003</fpage>–<lpage>9</lpage>. <pub-id pub-id-type="doi">10.4049/jimmunol.169.4.2003</pub-id><pub-id pub-id-type="pmid">12165526</pub-id></mixed-citation></ref><ref id="pone.0223444.ref038"><label>38</label><mixed-citation publication-type="journal"><name><surname>Diacovo</surname><given-names>TG</given-names></name>, <name><surname>Roth</surname><given-names>SJ</given-names></name>, <name><surname>Buccola</surname><given-names>JM</given-names></name>, <name><surname>Bainton</surname><given-names>DF</given-names></name>, <name><surname>Springer</surname><given-names>TA</given-names></name>. <article-title>Neutrophil rolling, arrest, and transmigration across activated, surface-adherent platelets via sequential action of P-selectin and the beta 2-integrin CD11b/CD18</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>1996</year>; <volume>88</volume>: <fpage>146</fpage>–<lpage>57</lpage>. <pub-id pub-id-type="pmid">8704169</pub-id></mixed-citation></ref><ref id="pone.0223444.ref039"><label>39</label><mixed-citation publication-type="journal"><name><surname>Corken</surname><given-names>A</given-names></name>, <name><surname>Russell</surname><given-names>S</given-names></name>, <name><surname>Dent</surname><given-names>J</given-names></name>, <name><surname>Post</surname><given-names>SR</given-names></name>, <name><surname>Ware</surname><given-names>J</given-names></name>. <article-title>Platelet glycoprotein Ib-IX as a regulator of systemic inflammation</article-title>. <source><italic toggle="yes">Arterioscler Thromb Vasc Biol</italic></source>. <year>2014</year>; <volume>34</volume>: <fpage>996</fpage>–<lpage>1001</lpage>. <pub-id pub-id-type="doi">10.1161/ATVBAHA.113.303113</pub-id><pub-id pub-id-type="pmid">24504734</pub-id></mixed-citation></ref><ref id="pone.0223444.ref040"><label>40</label><mixed-citation publication-type="journal"><name><surname>Klarstrom Engstrom</surname><given-names>K</given-names></name>, <name><surname>Brommesson</surname><given-names>C</given-names></name>, <name><surname>Kalvegren</surname><given-names>H</given-names></name>, <name><surname>Bengtsson</surname><given-names>T</given-names></name>. <article-title>Toll like receptor 2/1 mediated platelet adhesion and activation on bacterial mimetic surfaces is dependent on src/Syk-signaling and purinergic receptor P2X1 and P2Y12 activation</article-title>. <source><italic toggle="yes">Biointerphases</italic></source>. <year>2014</year>; <volume>9</volume>: <fpage>041003</fpage><pub-id pub-id-type="doi">10.1116/1.4901135</pub-id><pub-id pub-id-type="pmid">25553878</pub-id></mixed-citation></ref><ref id="pone.0223444.ref041"><label>41</label><mixed-citation publication-type="journal"><name><surname>Falker</surname><given-names>K</given-names></name>, <name><surname>Klarstrom-Engstrom</surname><given-names>K</given-names></name>, <name><surname>Bengtsson</surname><given-names>T</given-names></name>, <name><surname>Lindahl</surname><given-names>TL</given-names></name>, <name><surname>Grenegard</surname><given-names>M</given-names></name>. <article-title>The toll-like receptor 2/1 (TLR2/1) complex initiates human platelet activation via the src/Syk/LAT/PLCgamma2 signalling cascade</article-title>. <source><italic toggle="yes">Cell Signal</italic></source>. <year>2014</year>; <volume>26</volume>: <fpage>279</fpage>–<lpage>86</lpage>. <pub-id pub-id-type="doi">10.1016/j.cellsig.2013.11.011</pub-id><pub-id pub-id-type="pmid">24240055</pub-id></mixed-citation></ref><ref id="pone.0223444.ref042"><label>42</label><mixed-citation publication-type="journal"><name><surname>Senis</surname><given-names>YA</given-names></name>, <name><surname>Mazharian</surname><given-names>A</given-names></name>, <name><surname>Mori</surname><given-names>J</given-names></name>. <article-title>Src family kinases: at the forefront of platelet activation</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2014</year>; <volume>124</volume>: <fpage>2013</fpage>–<lpage>24</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2014-01-453134</pub-id><pub-id pub-id-type="pmid">25115887</pub-id></mixed-citation></ref><ref id="pone.0223444.ref043"><label>43</label><mixed-citation publication-type="journal"><name><surname>Assinger</surname><given-names>A</given-names></name>, <name><surname>Laky</surname><given-names>M</given-names></name>, <name><surname>Schabbauer</surname><given-names>G</given-names></name>, <name><surname>Hirschl</surname><given-names>AM</given-names></name>, <name><surname>Buchberger</surname><given-names>E</given-names></name>, <name><surname>Binder</surname><given-names>BR</given-names></name>, <etal>et al</etal><article-title>Efficient phagocytosis of periodontopathogens by neutrophils requires plasma factors, platelets and TLR2</article-title>. <source><italic toggle="yes">J Thromb Haemost</italic></source>. <year>2011</year>; <volume>9</volume>: <fpage>799</fpage>–<lpage>809</lpage>. <pub-id pub-id-type="doi">10.1111/j.1538-7836.2011.04193.x</pub-id><pub-id pub-id-type="pmid">21251195</pub-id></mixed-citation></ref><ref id="pone.0223444.ref044"><label>44</label><mixed-citation publication-type="journal"><name><surname>Belaaouaj</surname><given-names>A</given-names></name>, <name><surname>Kim</surname><given-names>KS</given-names></name>, <name><surname>Shapiro</surname><given-names>SD</given-names></name>. <article-title>Degradation of outer membrane protein A in Escherichia coli killing by neutrophil elastase</article-title>. <source><italic toggle="yes">Science</italic></source>. <year>2000</year>; <volume>289</volume>: <fpage>1185</fpage>–<lpage>8</lpage>. <pub-id pub-id-type="doi">10.1126/science.289.5482.1185</pub-id><pub-id pub-id-type="pmid">10947984</pub-id></mixed-citation></ref><ref id="pone.0223444.ref045"><label>45</label><mixed-citation publication-type="journal"><name><surname>Kawabata</surname><given-names>K</given-names></name>, <name><surname>Hagio</surname><given-names>T</given-names></name>, <name><surname>Matsuoka</surname><given-names>S</given-names></name>. <article-title>The role of neutrophil elastase in acute lung injury</article-title>. <source><italic toggle="yes">Eur J Pharmacol</italic></source>. <year>2002</year>; <volume>451</volume>: <fpage>1</fpage>–<lpage>10</lpage>. <pub-id pub-id-type="doi">10.1016/s0014-2999(02)02182-9</pub-id><pub-id pub-id-type="pmid">12223222</pub-id></mixed-citation></ref><ref id="pone.0223444.ref046"><label>46</label><mixed-citation publication-type="journal"><name><surname>Doring</surname><given-names>G</given-names></name>. <article-title>The role of neutrophil elastase in chronic inflammation</article-title>. <source><italic toggle="yes">Am J Respir Crit Care Med</italic></source>. <year>1994</year>; <volume>150</volume>: <fpage>S114</fpage>–<lpage>7</lpage>. <pub-id pub-id-type="doi">10.1164/ajrccm/150.6_Pt_2.S114</pub-id><pub-id pub-id-type="pmid">7952645</pub-id></mixed-citation></ref><ref id="pone.0223444.ref047"><label>47</label><mixed-citation publication-type="journal"><name><surname>Hammond</surname><given-names>ME</given-names></name>, <name><surname>Lapointe</surname><given-names>GR</given-names></name>, <name><surname>Feucht</surname><given-names>PH</given-names></name>, <name><surname>Hilt</surname><given-names>S</given-names></name>, <name><surname>Gallegos</surname><given-names>CA</given-names></name>, <name><surname>Gordon</surname><given-names>CA</given-names></name>, <etal>et al</etal><article-title>IL-8 induces neutrophil chemotaxis predominantly via type I IL-8 receptors</article-title>. <source><italic toggle="yes">J Immunol</italic></source>. <year>1995</year>; <volume>155</volume>: <fpage>1428</fpage>–<lpage>33</lpage>. <pub-id pub-id-type="pmid">7636208</pub-id></mixed-citation></ref><ref id="pone.0223444.ref048"><label>48</label><mixed-citation publication-type="journal"><name><surname>Sahoo</surname><given-names>M</given-names></name>, <name><surname>del Barrio</surname><given-names>L</given-names></name>, <name><surname>Miller</surname><given-names>MA</given-names></name>, <name><surname>Re</surname><given-names>F</given-names></name>. <article-title>Neutrophil Elastase Causes Tissue Damage That Decreases Host Tolerance to Lung Infection with Burkholderia Species.</article-title><source><italic toggle="yes">PLoS Pathog</italic></source>. <year>2014</year>; <volume>10</volume>: <fpage>e1004327</fpage><pub-id pub-id-type="doi">10.1371/journal.ppat.1004327</pub-id><pub-id pub-id-type="pmid">25166912</pub-id></mixed-citation></ref><ref id="pone.0223444.ref049"><label>49</label><mixed-citation publication-type="journal"><name><surname>Bardoel Bart</surname><given-names>W</given-names></name>, <name><surname>Kenny Elaine</surname><given-names>F</given-names></name>, <name><surname>Sollberger</surname><given-names>G</given-names></name>, <name><surname>Zychlinsky</surname><given-names>A</given-names></name>. <article-title>The Balancing Act of Neutrophils</article-title>. <source><italic toggle="yes">Cell Host Microbe</italic></source>. <year>2014</year>; <volume>15</volume>: <fpage>526</fpage>–<lpage>36</lpage>. <pub-id pub-id-type="doi">10.1016/j.chom.2014.04.011</pub-id><pub-id pub-id-type="pmid">24832448</pub-id></mixed-citation></ref><ref id="pone.0223444.ref050"><label>50</label><mixed-citation publication-type="journal"><name><surname>Middleton</surname><given-names>EA</given-names></name>, <name><surname>Rondina</surname><given-names>MT</given-names></name>, <name><surname>Schwertz</surname><given-names>H</given-names></name>, <name><surname>Zimmerman</surname><given-names>GA</given-names></name>. <article-title>Amicus or Adversary Revisited: Platelets in Acute Lung Injury and Acute Respiratory Distress Syndrome</article-title>. <source><italic toggle="yes">Am J Respir Cell Mol Biol</italic></source>. <year>2018</year>; <volume>59</volume>: <fpage>18</fpage>–<lpage>35</lpage>. <pub-id pub-id-type="doi">10.1165/rcmb.2017-0420TR</pub-id><pub-id pub-id-type="pmid">29553813</pub-id></mixed-citation></ref><ref id="pone.0223444.ref051"><label>51</label><mixed-citation publication-type="journal"><name><surname>Lee</surname><given-names>KH</given-names></name>, <name><surname>Hui</surname><given-names>KP</given-names></name>, <name><surname>Tan</surname><given-names>WC</given-names></name>. <article-title>Thrombocytopenia in sepsis: a predictor of mortality in the intensive care unit.</article-title><source><italic toggle="yes">Singapore Med J</italic></source>. <year>1993</year>; <volume>34</volume>: <fpage>245</fpage>–<lpage>6</lpage>. <pub-id pub-id-type="pmid">8266183</pub-id></mixed-citation></ref><ref id="pone.0223444.ref052"><label>52</label><mixed-citation publication-type="journal"><name><surname>Wuescher</surname><given-names>LM</given-names></name>, <name><surname>Takashima</surname><given-names>A</given-names></name>, <name><surname>Worth</surname><given-names>RG</given-names></name>. <article-title>A novel conditional platelet depletion mouse model reveals the importance of platelets in protection against Staphylococcus aureus bacteremia</article-title>. <source><italic toggle="yes">J Thromb Haemost</italic></source>. <year>2015</year>; <volume>13</volume>: <fpage>303</fpage>–<lpage>13</lpage>. <pub-id pub-id-type="doi">10.1111/jth.12795</pub-id><pub-id pub-id-type="pmid">25418277</pub-id></mixed-citation></ref><ref id="pone.0223444.ref053"><label>53</label><mixed-citation publication-type="journal"><name><surname>de Stoppelaar</surname><given-names>SF</given-names></name>, <name><surname>van 't Veer</surname><given-names>C</given-names></name>, <name><surname>Claushuis</surname><given-names>TA</given-names></name>, <name><surname>Albersen</surname><given-names>BJ</given-names></name>, <name><surname>Roelofs</surname><given-names>JJ</given-names></name>, <name><surname>van der Poll</surname><given-names>T</given-names></name>. <article-title>Thrombocytopenia impairs host defense in gram-negative pneumonia-derived sepsis in mice</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2014</year>; <volume>124</volume>: <fpage>3781</fpage>–<lpage>90</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2014-05-573915</pub-id><pub-id pub-id-type="pmid">25301709</pub-id></mixed-citation></ref><ref id="pone.0223444.ref054"><label>54</label><mixed-citation publication-type="journal"><name><surname>Hechler</surname><given-names>B</given-names></name>, <name><surname>Zimmermann</surname><given-names>C</given-names></name>, <name><surname>Rabouel</surname><given-names>Y</given-names></name>, <name><surname>Magnenat</surname><given-names>S</given-names></name>, <name><surname>Burban</surname><given-names>M</given-names></name>, <name><surname>Boisramé-Helms</surname><given-names>J</given-names></name>, <etal>et al</etal><article-title>A Potential Protective Role of Platelets during Septic Shock Does Not Depend on Their Purinergic Receptors</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2016</year>; <volume>128</volume>: <fpage>2537</fpage>–.</mixed-citation></ref><ref id="pone.0223444.ref055"><label>55</label><mixed-citation publication-type="journal"><name><surname>Martinod</surname><given-names>K</given-names></name>, <name><surname>Wagner</surname><given-names>DD</given-names></name>. <article-title>Thrombosis: tangled up in NETs.</article-title><source><italic toggle="yes">Blood</italic></source>. <year>2014</year>; <volume>123</volume>: <fpage>2768</fpage>–<lpage>76</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2013-10-463646</pub-id><pub-id pub-id-type="pmid">24366358</pub-id></mixed-citation></ref><ref id="pone.0223444.ref056"><label>56</label><mixed-citation publication-type="journal"><name><surname>Schauer</surname><given-names>C</given-names></name>, <name><surname>Janko</surname><given-names>C</given-names></name>, <name><surname>Munoz</surname><given-names>LE</given-names></name>, <name><surname>Zhao</surname><given-names>Y</given-names></name>, <name><surname>Kienhofer</surname><given-names>D</given-names></name>, <name><surname>Frey</surname><given-names>B</given-names></name>, <etal>et al</etal><article-title>Aggregated neutrophil extracellular traps limit inflammation by degrading cytokines and chemokines</article-title>. <source><italic toggle="yes">Nat Med</italic></source>. <year>2014</year>; <volume>20</volume>: <fpage>511</fpage>–<lpage>7</lpage>. <pub-id pub-id-type="doi">10.1038/nm.3547</pub-id><pub-id pub-id-type="pmid">24784231</pub-id></mixed-citation></ref><ref id="pone.0223444.ref057"><label>57</label><mixed-citation publication-type="journal"><name><surname>Looney</surname><given-names>MR</given-names></name>, <name><surname>Nguyen</surname><given-names>JX</given-names></name>, <name><surname>Hu</surname><given-names>Y</given-names></name>, <name><surname>Van Ziffle</surname><given-names>JA</given-names></name>, <name><surname>Lowell</surname><given-names>CA</given-names></name>, <name><surname>Matthay</surname><given-names>MA</given-names></name>. <article-title>Platelet depletion and aspirin treatment protect mice in a two-event model of transfusion-related acute lung injury</article-title>. <source><italic toggle="yes">The Journal of Clinical Investigation</italic></source>. <year>2009</year>; <volume>119</volume>: <fpage>3450</fpage>–<lpage>61</lpage>. <pub-id pub-id-type="doi">10.1172/JCI38432</pub-id><pub-id pub-id-type="pmid">19809160</pub-id></mixed-citation></ref><ref id="pone.0223444.ref058"><label>58</label><mixed-citation publication-type="journal"><name><surname>Luo</surname><given-names>S</given-names></name>, <name><surname>Wang</surname><given-names>Y</given-names></name>, <name><surname>An</surname><given-names>Q</given-names></name>, <name><surname>Chen</surname><given-names>H</given-names></name>, <name><surname>Zhao</surname><given-names>J</given-names></name>, <name><surname>Zhang</surname><given-names>J</given-names></name>, <etal>et al</etal><article-title>Platelets protect lung from injury induced by systemic inflammatory response</article-title>. <source><italic toggle="yes">Sci Rep</italic></source>. <year>2017</year>; <volume>7</volume>: <fpage>42080</fpage><pub-id pub-id-type="doi">10.1038/srep42080</pub-id><pub-id pub-id-type="pmid">28155889</pub-id></mixed-citation></ref><ref id="pone.0223444.ref059"><label>59</label><mixed-citation publication-type="journal"><name><surname>Abdulnour</surname><given-names>RE</given-names></name>, <name><surname>Dalli</surname><given-names>J</given-names></name>, <name><surname>Colby</surname><given-names>JK</given-names></name>, <name><surname>Krishnamoorthy</surname><given-names>N</given-names></name>, <name><surname>Timmons</surname><given-names>JY</given-names></name>, <name><surname>Tan</surname><given-names>SH</given-names></name>, <etal>et al</etal><article-title>Maresin 1 biosynthesis during platelet-neutrophil interactions is organ-protective</article-title>. <source><italic toggle="yes">Proc Natl Acad Sci U S A</italic></source>. <year>2014</year>; <volume>111</volume>: <fpage>16526</fpage>–<lpage>31</lpage>. <pub-id pub-id-type="doi">10.1073/pnas.1407123111</pub-id><pub-id pub-id-type="pmid">25369934</pub-id></mixed-citation></ref><ref id="pone.0223444.ref060"><label>60</label><mixed-citation publication-type="journal"><name><surname>Frangogiannis</surname><given-names>NG</given-names></name>. <article-title>The inflammatory response in myocardial injury, repair, and remodelling</article-title>. <source><italic toggle="yes">Nat Rev Cardiol</italic></source>. <year>2014</year>; <volume>11</volume>: <fpage>255</fpage>–<lpage>65</lpage>. <pub-id pub-id-type="doi">10.1038/nrcardio.2014.28</pub-id><pub-id pub-id-type="pmid">24663091</pub-id></mixed-citation></ref><ref id="pone.0223444.ref061"><label>61</label><mixed-citation publication-type="journal"><name><surname>Lefer</surname><given-names>AM</given-names></name>, <name><surname>Campbell</surname><given-names>B</given-names></name>, <name><surname>Scalia</surname><given-names>R</given-names></name>, <name><surname>Lefer</surname><given-names>DJ</given-names></name>. <article-title>Synergism Between Platelets and Neutrophils in Provoking Cardiac Dysfunction After Ischemia and Reperfusion.</article-title><source><italic toggle="yes">Role of Selectins</italic></source>. <year>1998</year>; <volume>98</volume>: <fpage>1322</fpage>–<lpage>8</lpage>.</mixed-citation></ref><ref id="pone.0223444.ref062"><label>62</label><mixed-citation publication-type="journal"><name><surname>Hargrave</surname><given-names>B</given-names></name>, <name><surname>Li</surname><given-names>F</given-names></name>. <article-title>Nanosecond pulse electric field activation of platelet-rich plasma reduces myocardial infarct size and improves left ventricular mechanical function in the rabbit heart</article-title>. <source><italic toggle="yes">The journal of extra-corporeal technology</italic></source>. <year>2012</year>; <volume>44</volume>: <fpage>198</fpage>–<lpage>204</lpage>. <pub-id pub-id-type="pmid">23441560</pub-id></mixed-citation></ref><ref id="pone.0223444.ref063"><label>63</label><mixed-citation publication-type="journal"><name><surname>Milioli</surname><given-names>M</given-names></name>, <name><surname>Ibanez-Vea</surname><given-names>M</given-names></name>, <name><surname>Sidoli</surname><given-names>S</given-names></name>, <name><surname>Palmisano</surname><given-names>G</given-names></name>, <name><surname>Careri</surname><given-names>M</given-names></name>, <name><surname>Larsen</surname><given-names>MR</given-names></name>. <article-title>Quantitative proteomics analysis of platelet-derived microparticles reveals distinct protein signatures when stimulated by different physiological agonists</article-title>. <source><italic toggle="yes">J Proteomics</italic></source>. <year>2015</year>; <volume>121</volume>: <fpage>56</fpage>–<lpage>66</lpage>. <pub-id pub-id-type="doi">10.1016/j.jprot.2015.03.013</pub-id><pub-id pub-id-type="pmid">25835965</pub-id></mixed-citation></ref><ref id="pone.0223444.ref064"><label>64</label><mixed-citation publication-type="journal"><name><surname>Vélez</surname><given-names>P</given-names></name>, <name><surname>Izquierdo</surname><given-names>I</given-names></name>, <name><surname>Rosa</surname><given-names>I</given-names></name>, <name><surname>García</surname><given-names>Á</given-names></name>. <article-title>A 2D-DIGE-based proteomic analysis reveals differences in the platelet releasate composition when comparing thrombin and collagen stimulations</article-title>. <source><italic toggle="yes">Sci Rep</italic></source>. <year>2015</year>; <volume>5</volume>: <fpage>8198</fpage><pub-id pub-id-type="doi">10.1038/srep08198</pub-id><pub-id pub-id-type="pmid">25645904</pub-id></mixed-citation></ref><ref id="pone.0223444.ref065"><label>65</label><mixed-citation publication-type="journal"><name><surname>Chatterjee</surname><given-names>M</given-names></name>, <name><surname>Huang</surname><given-names>Z</given-names></name>, <name><surname>Zhang</surname><given-names>W</given-names></name>, <name><surname>Jiang</surname><given-names>L</given-names></name>, <name><surname>Hultenby</surname><given-names>K</given-names></name>, <name><surname>Zhu</surname><given-names>L</given-names></name>, <etal>et al</etal><article-title>Distinct platelet packaging, release, and surface expression of proangiogenic and antiangiogenic factors on different platelet stimuli</article-title>. <source><italic toggle="yes">Blood</italic></source>. <year>2011</year>; <volume>117</volume>: <fpage>3907</fpage>–<lpage>11</lpage>. <pub-id pub-id-type="doi">10.1182/blood-2010-12-327007</pub-id><pub-id pub-id-type="pmid">21330475</pub-id></mixed-citation></ref><ref id="pone.0223444.ref066"><label>66</label><mixed-citation publication-type="journal"><name><surname>Pokrovskaya</surname><given-names>ID</given-names></name>, <name><surname>Aronova</surname><given-names>MA</given-names></name>, <name><surname>Kamykowski</surname><given-names>JA</given-names></name>, <name><surname>Prince</surname><given-names>AA</given-names></name>, <name><surname>Hoyne</surname><given-names>JD</given-names></name>, <name><surname>Calco</surname><given-names>GN</given-names></name>, <etal>et al</etal><article-title>STEM tomography reveals that the canalicular system and alpha-granules remain separate compartments during early secretion stages in blood platelets</article-title>. <source><italic toggle="yes">J Thromb Haemost</italic></source>. <year>2016</year>; <volume>14</volume>: <fpage>572</fpage>–<lpage>84</lpage>. <pub-id pub-id-type="doi">10.1111/jth.13225</pub-id><pub-id pub-id-type="pmid">26663480</pub-id></mixed-citation></ref></ref-list></back></pmc></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Sante/explor/ChloroquineV1/Data/Pmc/Corpus
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 0007760 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Pmc/Corpus/biblio.hfd -nk 0007760 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Sante
|area= ChloroquineV1
|flux= Pmc
|étape= Corpus
|type= RBID
|clé=
|texte=
}}
| This area was generated with Dilib version V0.6.33. |  |