Optimizing Western Blots for the Detection of Endogenous α-Synuclein in the Enteric Nervous System.
Identifieur interne : 000423 ( PubMed/Curation ); précédent : 000422; suivant : 000424Optimizing Western Blots for the Detection of Endogenous α-Synuclein in the Enteric Nervous System.
Auteurs : Cécile Preterre [France] ; Anne-Gaëlle Corbillé [France] ; Gaëlle Balloy [France] ; Franck Letournel [France] ; Michel Neunlist [France] ; Pascal Derkinderen [France]Source :
- Journal of Parkinson's disease [ 1877-718X ] ; 2015.
English descriptors
- KwdEn :
- MESH :
- chemical , analysis : alpha-Synuclein.
- chemical , metabolism : Biomarkers.
- diagnosis : Parkinson Disease.
- innervation : Intestine, Small.
- metabolism : Enteric Nervous System, Intestine, Small.
- methods : Blotting, Western.
- standards : Blotting, Western.
- Animals, Humans, Rats.
Abstract
Alpha-synuclein containing inclusions in neurons, the characteristic pathological lesions of Parkinson's disease (PD), are not limited to the central nervous system, but also affect the enteric nervous system (ENS). This suggests that the ENS offer some potential as a surrogate of central nervous system pathology and that it may represent an original source of biomarkers for PD. However, the usefulness of α-synuclein detection in gastrointestinal biopsies as a biomarker for PD is still unclear, as the different immunohistochemical methods employed to date have led to conflicting results.
DOI: 10.3233/JPD-150670
PubMed: 26599299
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uniqKey="Balloy G" first="Gaëlle" last="Balloy">Gaëlle Balloy</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author><author><name sortKey="Letournel, Franck" sort="Letournel, Franck" uniqKey="Letournel F" first="Franck" last="Letournel">Franck Letournel</name><affiliation wicri:level="1"><nlm:affiliation>CHU Angers, Neurobiology and Neuropathology Laboratory, Angers F-49033, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>CHU Angers, Neurobiology and Neuropathology Laboratory, Angers F-49033</wicri:regionArea></affiliation></author><author><name sortKey="Neunlist, Michel" sort="Neunlist, Michel" uniqKey="Neunlist M" first="Michel" last="Neunlist">Michel Neunlist</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, 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wicri:stream="PubMed" wicri:step="Curation">000423</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Optimizing Western Blots for the Detection of Endogenous α-Synuclein in the Enteric Nervous System.</title><author><name sortKey="Preterre, Cecile" sort="Preterre, Cecile" uniqKey="Preterre C" first="Cécile" last="Preterre">Cécile Preterre</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author><author><name sortKey="Corbille, Anne Gaelle" sort="Corbille, Anne Gaelle" uniqKey="Corbille A" first="Anne-Gaëlle" last="Corbillé">Anne-Gaëlle Corbillé</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author><author><name sortKey="Balloy, Gaelle" sort="Balloy, Gaelle" uniqKey="Balloy G" first="Gaëlle" last="Balloy">Gaëlle Balloy</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author><author><name sortKey="Letournel, Franck" sort="Letournel, Franck" uniqKey="Letournel F" first="Franck" last="Letournel">Franck Letournel</name><affiliation wicri:level="1"><nlm:affiliation>CHU Angers, Neurobiology and Neuropathology Laboratory, Angers F-49033, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>CHU Angers, Neurobiology and Neuropathology Laboratory, Angers F-49033</wicri:regionArea></affiliation></author><author><name sortKey="Neunlist, Michel" sort="Neunlist, Michel" uniqKey="Neunlist M" first="Michel" last="Neunlist">Michel Neunlist</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author><author><name sortKey="Derkinderen, Pascal" sort="Derkinderen, Pascal" uniqKey="Derkinderen P" first="Pascal" last="Derkinderen">Pascal Derkinderen</name><affiliation wicri:level="1"><nlm:affiliation>Inserm, U913, Nantes, F-44093, France.</nlm:affiliation><country xml:lang="fr">France</country><wicri:regionArea>Inserm, U913, Nantes, F-44093</wicri:regionArea></affiliation></author></analytic><series><title level="j">Journal of Parkinson's disease</title><idno type="eISSN">1877-718X</idno><imprint><date when="2015" type="published">2015</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>Biomarkers (metabolism)</term><term>Blotting, Western (methods)</term><term>Blotting, Western (standards)</term><term>Enteric Nervous System (metabolism)</term><term>Humans</term><term>Intestine, Small (innervation)</term><term>Intestine, Small (metabolism)</term><term>Parkinson Disease (diagnosis)</term><term>Rats</term><term>alpha-Synuclein (analysis)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="analysis" xml:lang="en"><term>alpha-Synuclein</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Biomarkers</term></keywords><keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Parkinson Disease</term></keywords><keywords scheme="MESH" qualifier="innervation" xml:lang="en"><term>Intestine, Small</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Enteric Nervous System</term><term>Intestine, Small</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Blotting, Western</term></keywords><keywords scheme="MESH" qualifier="standards" xml:lang="en"><term>Blotting, Western</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Humans</term><term>Rats</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Alpha-synuclein containing inclusions in neurons, the characteristic pathological lesions of Parkinson's disease (PD), are not limited to the central nervous system, but also affect the enteric nervous system (ENS). This suggests that the ENS offer some potential as a surrogate of central nervous system pathology and that it may represent an original source of biomarkers for PD. However, the usefulness of α-synuclein detection in gastrointestinal biopsies as a biomarker for PD is still unclear, as the different immunohistochemical methods employed to date have led to conflicting results.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">26599299</PMID><DateCreated><Year>2015</Year><Month>12</Month><Day>19</Day></DateCreated><DateCompleted><Year>2016</Year><Month>09</Month><Day>29</Day></DateCompleted><DateRevised><Year>2015</Year><Month>12</Month><Day>19</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Electronic">1877-718X</ISSN><JournalIssue CitedMedium="Internet"><Volume>5</Volume><Issue>4</Issue><PubDate><Year>2015</Year></PubDate></JournalIssue><Title>Journal of Parkinson's disease</Title><ISOAbbreviation>J Parkinsons Dis</ISOAbbreviation></Journal><ArticleTitle>Optimizing Western Blots for the Detection of Endogenous α-Synuclein in the Enteric Nervous System.</ArticleTitle><Pagination><MedlinePgn>765-72</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.3233/JPD-150670</ELocationID><Abstract><AbstractText Label="BACKGROUND" NlmCategory="BACKGROUND">Alpha-synuclein containing inclusions in neurons, the characteristic pathological lesions of Parkinson's disease (PD), are not limited to the central nervous system, but also affect the enteric nervous system (ENS). This suggests that the ENS offer some potential as a surrogate of central nervous system pathology and that it may represent an original source of biomarkers for PD. However, the usefulness of α-synuclein detection in gastrointestinal biopsies as a biomarker for PD is still unclear, as the different immunohistochemical methods employed to date have led to conflicting results.</AbstractText><AbstractText Label="OBJECTIVE" NlmCategory="OBJECTIVE">Our aim is to propose an optimized immunoblotting method for the detection of endogenous α-synuclein in the healthy ENS that may be used to supplement the immunohistochemical analysis.</AbstractText><AbstractText Label="METHODS" NlmCategory="METHODS">Primary culture of rat ENS and homogenates of human small intestine were analyzed by Western Blot using seven different α-synuclein and phospho-α-synuclein antibodies along with two methods that increase α-synuclein retention on blot membranes, namely incubation of the membranes with paraformaldehyde (PFA) or treatment of samples with the crosslinker dithiobis[succinimidylpropionate] (DSP).</AbstractText><AbstractText Label="RESULTS" NlmCategory="RESULTS">A moderate improvement in the detection of endogenous enteric α-synuclein was observed following membrane fixation with PFA for only two of the seven antibodies we tested. Immunodetection of total and phosphorylated α-synuclein in the ENS was markedly improved when samples were treated with DSP, regardless of the antibody used.</AbstractText><AbstractText Label="CONCLUSIONS" NlmCategory="CONCLUSIONS">Our results demonstrate that the detection of α-synuclein in the gut by Western Blot can be optimized by using methods for enhanced membrane retention of the protein along with the appropriate antibody. Such an optimized protocol opens the way to the development of novel biomarkers for PD that will enable a quantification of α-synuclein in gastrointestinal biopsies.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Preterre</LastName><ForeName>Cécile</ForeName><Initials>C</Initials><AffiliationInfo><Affiliation>Inserm, U913, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Nantes University, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Department of Neurology, Nantes, F-44093, France.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Corbillé</LastName><ForeName>Anne-Gaëlle</ForeName><Initials>AG</Initials><AffiliationInfo><Affiliation>Inserm, U913, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Nantes University, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Department of Neurology, Nantes, F-44093, France.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Balloy</LastName><ForeName>Gaëlle</ForeName><Initials>G</Initials><AffiliationInfo><Affiliation>Inserm, U913, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Nantes University, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Department of Neurology, Nantes, F-44093, France.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Letournel</LastName><ForeName>Franck</ForeName><Initials>F</Initials><AffiliationInfo><Affiliation>CHU Angers, Neurobiology and Neuropathology Laboratory, Angers F-49033, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Université of Angers, UPRES EA3143, F-49033, France.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Neunlist</LastName><ForeName>Michel</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Inserm, U913, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Nantes University, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Department of Neurology, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Institut des Maladies de l'Appareil Digestif, Nantes, France.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Derkinderen</LastName><ForeName>Pascal</ForeName><Initials>P</Initials><AffiliationInfo><Affiliation>Inserm, U913, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Nantes University, Nantes, F-44093, France.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>CHU Nantes, Department of Neurology, Nantes, F-44093, France.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>Netherlands</Country><MedlineTA>J Parkinsons Dis</MedlineTA><NlmUniqueID>101567362</NlmUniqueID><ISSNLinking>1877-7171</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D015415">Biomarkers</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="C497604">SNCA protein, human</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="C497606">Snca protein, rat</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D051844">alpha-Synuclein</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D000818" MajorTopicYN="N">Animals</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D015415" MajorTopicYN="N">Biomarkers</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D015153" MajorTopicYN="N">Blotting, Western</DescriptorName><QualifierName UI="Q000379" MajorTopicYN="Y">methods</QualifierName><QualifierName UI="Q000592" MajorTopicYN="N">standards</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D017615" MajorTopicYN="N">Enteric Nervous System</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D006801" MajorTopicYN="N">Humans</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D007421" MajorTopicYN="N">Intestine, Small</DescriptorName><QualifierName UI="Q000294" MajorTopicYN="N">innervation</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010300" MajorTopicYN="N">Parkinson Disease</DescriptorName><QualifierName UI="Q000175" MajorTopicYN="N">diagnosis</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D051381" MajorTopicYN="N">Rats</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D051844" MajorTopicYN="N">alpha-Synuclein</DescriptorName><QualifierName UI="Q000032" MajorTopicYN="Y">analysis</QualifierName></MeshHeading></MeshHeadingList><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Alpha-synuclein</Keyword><Keyword MajorTopicYN="N">Parkinson’s disease</Keyword><Keyword MajorTopicYN="N">Western Blot</Keyword><Keyword MajorTopicYN="N">enteric nervous system</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate 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