Trehalose Alleviates Crystalline Silica-Induced Pulmonary Fibrosis via Activation of the TFEB-Mediated Autophagy-Lysosomal System in Alveolar Macrophages
Identifieur interne : 000E44 ( Ncbi/Merge ); précédent : 000E43; suivant : 000E45Trehalose Alleviates Crystalline Silica-Induced Pulmonary Fibrosis via Activation of the TFEB-Mediated Autophagy-Lysosomal System in Alveolar Macrophages
Auteurs : Xiu He ; Shi Chen ; Chao Li ; Jiaqi Ban ; Yungeng Wei ; Yangyang He ; Fangwei Liu ; Ying Chen ; Jie ChenSource :
- Cells [ 2073-4409 ] ; 2020.
Abstract
Silicosis is an occupational lung disease characterized by persistent inflammation and irreversible fibrosis. Crystalline silica (CS) particles are mainly phagocytized by alveolar macrophages (AMs), which trigger apoptosis, inflammation, and pulmonary fibrosis. Previously, we found that autophagy-lysosomal system dysfunction in AMs was involved in CS-induced inflammation and fibrosis. Induction of autophagy and lysosomal biogenesis by transcription factor EB (TFEB) nuclear translocation can rescue fibrotic diseases. However, the role of TFEB in silicosis is unknown. In this study, we found that CS induced TFEB nuclear localization and increased TFEB expression in macrophages both in vivo and in vitro. However, TFEB overexpression or treatment with the TFEB activator trehalose (Tre) alleviated lysosomal dysfunction and enhanced autophagic flux. It also reduced apoptosis, inflammatory cytokine levels, and fibrosis. Both pharmacologically inhibition of autophagy and TFEB knockdown in macrophages significantly abolished the antiapoptotic and anti-inflammatory effects elicited by either TFEB overexpression or Tre treatment. In conclusion, these results uncover a protective role of TFEB-mediated autophagy in silicosis. Our study suggests that restoration of autophagy-lysosomal function by Tre-induced TFEB activation may be a novel strategy for the treatment of silicosis.
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DOI: 10.3390/cells9010122
PubMed: 31947943
PubMed Central: 7016807
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Trehalose Alleviates Crystalline Silica-Induced Pulmonary Fibrosis via Activation of the TFEB-Mediated Autophagy-Lysosomal System in Alveolar Macrophages</title><author><name sortKey="He, Xiu" sort="He, Xiu" uniqKey="He X" first="Xiu" last="He">Xiu He</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Shi" sort="Chen, Shi" uniqKey="Chen S" first="Shi" last="Chen">Shi Chen</name><affiliation><nlm:aff id="af2-cells-09-00122">School of Medicine, Hunan Normal University, No.36 Lushan Road, Changsha 410013, China;<email>chenshonest@163.com</email></nlm:aff></affiliation></author><author><name sortKey="Li, Chao" sort="Li, Chao" uniqKey="Li C" first="Chao" last="Li">Chao Li</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Ban, Jiaqi" sort="Ban, Jiaqi" uniqKey="Ban J" first="Jiaqi" last="Ban">Jiaqi Ban</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Wei, Yungeng" sort="Wei, Yungeng" uniqKey="Wei Y" first="Yungeng" last="Wei">Yungeng Wei</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="He, Yangyang" sort="He, Yangyang" uniqKey="He Y" first="Yangyang" last="He">Yangyang He</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Liu, Fangwei" sort="Liu, Fangwei" uniqKey="Liu F" first="Fangwei" last="Liu">Fangwei Liu</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Ying" sort="Chen, Ying" uniqKey="Chen Y" first="Ying" last="Chen">Ying Chen</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Jie" sort="Chen, Jie" uniqKey="Chen J" first="Jie" last="Chen">Jie Chen</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">31947943</idno><idno type="pmc">7016807</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016807</idno><idno type="RBID">PMC:7016807</idno><idno type="doi">10.3390/cells9010122</idno><date when="2020">2020</date><idno type="wicri:Area/Pmc/Corpus">000479</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000479</idno><idno type="wicri:Area/Pmc/Curation">000479</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">000479</idno><idno type="wicri:Area/Pmc/Checkpoint">000008</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Checkpoint">000008</idno><idno type="wicri:Area/Ncbi/Merge">000E44</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Trehalose Alleviates Crystalline Silica-Induced Pulmonary Fibrosis via Activation of the TFEB-Mediated Autophagy-Lysosomal System in Alveolar Macrophages</title><author><name sortKey="He, Xiu" sort="He, Xiu" uniqKey="He X" first="Xiu" last="He">Xiu He</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Shi" sort="Chen, Shi" uniqKey="Chen S" first="Shi" last="Chen">Shi Chen</name><affiliation><nlm:aff id="af2-cells-09-00122">School of Medicine, Hunan Normal University, No.36 Lushan Road, Changsha 410013, China;<email>chenshonest@163.com</email></nlm:aff></affiliation></author><author><name sortKey="Li, Chao" sort="Li, Chao" uniqKey="Li C" first="Chao" last="Li">Chao Li</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Ban, Jiaqi" sort="Ban, Jiaqi" uniqKey="Ban J" first="Jiaqi" last="Ban">Jiaqi Ban</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Wei, Yungeng" sort="Wei, Yungeng" uniqKey="Wei Y" first="Yungeng" last="Wei">Yungeng Wei</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="He, Yangyang" sort="He, Yangyang" uniqKey="He Y" first="Yangyang" last="He">Yangyang He</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Liu, Fangwei" sort="Liu, Fangwei" uniqKey="Liu F" first="Fangwei" last="Liu">Fangwei Liu</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Ying" sort="Chen, Ying" uniqKey="Chen Y" first="Ying" last="Chen">Ying Chen</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author><author><name sortKey="Chen, Jie" sort="Chen, Jie" uniqKey="Chen J" first="Jie" last="Chen">Jie Chen</name><affiliation><nlm:aff id="af1-cells-09-00122">Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</nlm:aff></affiliation></author></analytic><series><title level="j">Cells</title><idno type="eISSN">2073-4409</idno><imprint><date when="2020">2020</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Silicosis is an occupational lung disease characterized by persistent inflammation and irreversible fibrosis. Crystalline silica (CS) particles are mainly phagocytized by alveolar macrophages (AMs), which trigger apoptosis, inflammation, and pulmonary fibrosis. Previously, we found that autophagy-lysosomal system dysfunction in AMs was involved in CS-induced inflammation and fibrosis. Induction of autophagy and lysosomal biogenesis by transcription factor EB (TFEB) nuclear translocation can rescue fibrotic diseases. However, the role of TFEB in silicosis is unknown. In this study, we found that CS induced TFEB nuclear localization and increased TFEB expression in macrophages both in vivo and in vitro. However, TFEB overexpression or treatment with the TFEB activator trehalose (Tre) alleviated lysosomal dysfunction and enhanced autophagic flux. It also reduced apoptosis, inflammatory cytokine levels, and fibrosis. Both pharmacologically inhibition of autophagy and TFEB knockdown in macrophages significantly abolished the antiapoptotic and anti-inflammatory effects elicited by either TFEB overexpression or Tre treatment. In conclusion, these results uncover a protective role of TFEB-mediated autophagy in silicosis. Our study suggests that restoration of autophagy-lysosomal function by Tre-induced TFEB activation may be a novel strategy for the treatment of silicosis.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Leung, C C" uniqKey="Leung C">C.C. Leung</name></author><author><name sortKey="Yu, I T S" uniqKey="Yu I">I.T.S. Yu</name></author><author><name sortKey="Chen, W" uniqKey="Chen W">W. Chen</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, C" uniqKey="Li C">C. Li</name></author><author><name sortKey="Du, S" uniqKey="Du S">S. Du</name></author><author><name sortKey="Lu, Y" uniqKey="Lu Y">Y. 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<front><journal-meta><journal-id journal-id-type="nlm-ta">Cells</journal-id><journal-id journal-id-type="iso-abbrev">Cells</journal-id><journal-id journal-id-type="publisher-id">cells</journal-id><journal-title-group><journal-title>Cells</journal-title></journal-title-group><issn pub-type="epub">2073-4409</issn><publisher><publisher-name>MDPI</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">31947943</article-id><article-id pub-id-type="pmc">7016807</article-id><article-id pub-id-type="doi">10.3390/cells9010122</article-id><article-id pub-id-type="publisher-id">cells-09-00122</article-id><article-categories><subj-group subj-group-type="heading"><subject>Article</subject></subj-group></article-categories><title-group><article-title>Trehalose Alleviates Crystalline Silica-Induced Pulmonary Fibrosis via Activation of the TFEB-Mediated Autophagy-Lysosomal System in Alveolar Macrophages</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>He</surname><given-names>Xiu</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref><xref ref-type="author-notes" rid="fn1-cells-09-00122">†</xref></contrib><contrib contrib-type="author"><name><surname>Chen</surname><given-names>Shi</given-names></name><xref ref-type="aff" rid="af2-cells-09-00122">2</xref><xref ref-type="author-notes" rid="fn1-cells-09-00122">†</xref></contrib><contrib contrib-type="author"><name><surname>Li</surname><given-names>Chao</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>Ban</surname><given-names>Jiaqi</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>Wei</surname><given-names>Yungeng</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>He</surname><given-names>Yangyang</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>Liu</surname><given-names>Fangwei</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>Chen</surname><given-names>Ying</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref></contrib><contrib contrib-type="author"><name><surname>Chen</surname><given-names>Jie</given-names></name><xref ref-type="aff" rid="af1-cells-09-00122">1</xref><xref rid="c1-cells-09-00122" ref-type="corresp">*</xref></contrib></contrib-group><aff id="af1-cells-09-00122"><label>1</label>Division of Pneumoconiosis, School of Public Health, China Medical University, No. 77 Puhe Road, Shenyang North New Area, Shenyang 110122, China;<email>xiuhe_cmu@163.com</email> (X.H.);<email>lichao@cmu.edu.cn</email> (C.L.);<email>17614027841@163.com</email> (J.B.);<email>weiyungeng1736@163.com</email> (Y.W.);<email>heyang129@sina.com</email> (Y.H.);<email>fwliu@cmu.edu.cn</email> (F.L.);<email>ychen25@cmu.edu.cn</email> (Y.C.)</aff><aff id="af2-cells-09-00122"><label>2</label>School of Medicine, Hunan Normal University, No.36 Lushan Road, Changsha 410013, China;<email>chenshonest@163.com</email></aff><author-notes><corresp id="c1-cells-09-00122"><label>*</label>Correspondence: <email>jchen@cmu.edu.cn</email>; Tel.: +86-24-31939079</corresp><fn id="fn1-cells-09-00122"><label>†</label><p>Equal contribution to this study.</p></fn></author-notes><pub-date pub-type="epub"><day>04</day><month>1</month><year>2020</year></pub-date><pub-date pub-type="collection"><month>1</month><year>2020</year></pub-date><volume>9</volume><issue>1</issue><elocation-id>122</elocation-id><history><date date-type="received"><day>19</day><month>11</month><year>2019</year></date><date date-type="accepted"><day>01</day><month>1</month><year>2020</year></date></history><permissions><copyright-statement>© 2020 by the authors.</copyright-statement><copyright-year>2020</copyright-year><license license-type="open-access"><license-p>Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (<ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">http://creativecommons.org/licenses/by/4.0/</ext-link>).</license-p></license></permissions><abstract><p>Silicosis is an occupational lung disease characterized by persistent inflammation and irreversible fibrosis. Crystalline silica (CS) particles are mainly phagocytized by alveolar macrophages (AMs), which trigger apoptosis, inflammation, and pulmonary fibrosis. Previously, we found that autophagy-lysosomal system dysfunction in AMs was involved in CS-induced inflammation and fibrosis. Induction of autophagy and lysosomal biogenesis by transcription factor EB (TFEB) nuclear translocation can rescue fibrotic diseases. However, the role of TFEB in silicosis is unknown. In this study, we found that CS induced TFEB nuclear localization and increased TFEB expression in macrophages both in vivo and in vitro. However, TFEB overexpression or treatment with the TFEB activator trehalose (Tre) alleviated lysosomal dysfunction and enhanced autophagic flux. It also reduced apoptosis, inflammatory cytokine levels, and fibrosis. Both pharmacologically inhibition of autophagy and TFEB knockdown in macrophages significantly abolished the antiapoptotic and anti-inflammatory effects elicited by either TFEB overexpression or Tre treatment. In conclusion, these results uncover a protective role of TFEB-mediated autophagy in silicosis. Our study suggests that restoration of autophagy-lysosomal function by Tre-induced TFEB activation may be a novel strategy for the treatment of silicosis.</p></abstract><kwd-group><kwd>TFEB</kwd><kwd>trehalose</kwd><kwd>silicosis</kwd><kwd>alveolar macrophages</kwd><kwd>autophagy</kwd><kwd>lysosome</kwd></kwd-group></article-meta></front><floats-group><fig id="cells-09-00122-f001" orientation="portrait" position="float"><label>Figure 1</label><caption><p>Crystalline silica (CS) induces transcription factor EB (TFEB) nuclear localization and increases TFEB expression in vivo. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of TFEB levels in the cytosol or nucleus of lung tissue on day 7 or day 56 post CS instillation; β-actin, loading control for cytosolic proteins; and lamin B, loading control for nuclear proteins (<italic>n</italic> = 4). (<bold>C</bold>) qPCR analysis of TFEB expression in lung tissue on day 7 or day 56 (<italic>n</italic> = 5 to 6). (<bold>D</bold>) Immunofluorescence analysis of the distribution of F4/80 and TFEB in lung tissue on day 7 or 56 post saline or CS treatment. Red arrows indicate alveolar macrophages (AMs) in the lungs containing silicotic lesions with predominantly nuclear TFEB staining (scale bar = 50 μm and <italic>n</italic> = 5). (<bold>E</bold>) Immunofluorescence analysis of TFEB nucleus translocation in primary AMs from bronchoalveolar lavage fluid (BALF) (scale bar = 10 μm and <italic>n</italic> = 3 to 4). (<bold>F</bold>) qPCR analysis of TFEB expression in primary AMs on day 7 or 56 post CS instillation (<italic>n</italic> = 4). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05; **, <italic>p</italic> < 0.01; and ##, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g001"></graphic></fig><fig id="cells-09-00122-f002" orientation="portrait" position="float"><label>Figure 2</label><caption><p>TFEB modulates CS-induced macrophage apoptosis and inflammatory response in vitro. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of TFEB levels in the cytosol and nucleus of MH-S cells 12 h post saline or CS treatment; β-actin, loading control for cytosolic proteins and lamin B, loading control for nuclear proteins (<italic>n</italic> = 4). (<bold>C</bold>) qPCR analysis of TFEB expression in MH-S cells 12 h post-saline or CS treatment (<italic>n</italic> = 4). (<bold>D</bold>,<bold>E</bold>) TFEB knockdown deteriorates CS-induced apoptosis and inflammation. Immunoblotting analysis of BAX and BCL-2 protein levels (<italic>n</italic> = 3 to 4). (<bold>F</bold>,<bold>G</bold>) TUNEL (green) and DAPI (blue) staining and ratios of TUNEL-positive apoptotic cells (scale bar = 50 μm and <italic>n</italic> = 4). (<bold>H</bold>–<bold>K</bold>) ELISA analysis of IL-6, MCP-1, TNF-α, and IL-1β levels in MH-S cell supernatant (<italic>n</italic> = 3 to 4). (<bold>L</bold>,<bold>M</bold>) TFEB overexpression relieves CS-induced apoptosis and inflammation. Immunoblotting analysis of BAX and BCL-2 protein levels (<italic>n</italic> = 4). (<bold>N</bold>,<bold>O</bold>) TUNEL (green) and DAPI (blue) staining and ratios of TUNEL-positive apoptotic cells (scale bar = 50 μm and <italic>n</italic> = 4). (<bold>P</bold>–<bold>S</bold>) ELISA analysis of IL-6, MCP-1, TNF-α, and IL-1β levels in MH-S cell supernatant (<italic>n</italic> = 3 to 4). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05; **, <italic>p</italic> < 0.01; and ##, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g002"></graphic></fig><fig id="cells-09-00122-f003" orientation="portrait" position="float"><label>Figure 3</label><caption><p>TFEB overexpression relieves CS-induced macrophage apoptosis and inflammation via autophagic substrate degradation in vitro. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of LC3II, Atg5, and Beclin 1 protein levels (<italic>n</italic> = 4). (<bold>C</bold>,<bold>D</bold>) Immunoblotting analysis of LAMP1 protein levels (<italic>n</italic> = 3). (<bold>E</bold>) Cells were grown on coverslips and treated with CS followed by staining with 50 nmol/L Lysotracker Red or 5 μg/mL acridine orange at 37 °C for 30 min (scale bar = 10 μm and <italic>n</italic> = 4). (<bold>F</bold>) Immunofluorescence analysis of the colocalization of LAMP1 and LC3 12 h post CS treatment (scale bar = 25 μm and <italic>n</italic> = 4). (<bold>G</bold>,<bold>H</bold>) Immunoblotting analysis of p62 protein levels (<italic>n</italic> = 4). (<bold>I</bold>) Immunofluorescence analysis of the colocalization of the p62 and ubiquitin 12 h post CS treatment (scale bar = 25 μm and <italic>n</italic> = 4). <bold>(J</bold>,<bold>K</bold>) TUNEL (green) and DAPI (blue) staining and ratios of TUNEL-positive apoptotic cells (scale bar = 50 μm and <italic>n</italic> = 4). (<bold>L</bold>–<bold>O</bold>) ELISA analysis of IL-6, MCP-1, TNF-α, and IL-1β levels in MH-S cell supernatant (<italic>n</italic> = 3 to 4). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05 and **, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g003"></graphic></fig><fig id="cells-09-00122-f004" orientation="portrait" position="float"><label>Figure 4</label><caption><p>Trehalose (Tre) affects autophagy-associated proteins in the lungs and AMs of CS-treated mice. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of cytosol and nuclear TFEB levels in lung tissue on day 7 or 56 post CS or Tre treatment; β-actin, loading control for cytosolic proteins; and LaminB, loading control for nuclear proteins (<italic>n</italic> = 3 to 4). (<bold>C</bold>) qPCR analysis of TFEB levels in lung tissue on day 7 or 56 post-CS or Tre treatment (<italic>n</italic> = 4). (<bold>D</bold>) Immunofluorescence analysis of TFEB nuclear translocation in primary AMs in BALF on day 7 or 56 (scale bar = 10 μm and <italic>n</italic> = 3 to 4). (<bold>E</bold>) qPCR analysis of TFEB expression in AMs on day 7 or 56 post CS or Tre treatment (<italic>n</italic> = 4). (<bold>F</bold>,<bold>G</bold>) Immunoblotting analysis of LC3II, Atg5, and Beclin 1 protein levels in lung tissue on day 7 or 56 post CS and Tre treatment (<italic>n</italic> = 5 to 6). (<bold>H</bold>) Immunofluorescence analysis of LC3 in AMs on day 7 or 56 post CS or Tre treatment (Scale bar = 25 μm and <italic>n</italic> = 3). (<bold>I</bold>,<bold>J</bold>) Immunoblotting analysis of LC3II and Atg5 protein levels in AMs on day 7 or 56 post CS or Tre treatment (<italic>n</italic> = 3 to 4). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05; **, <italic>p</italic> < 0.01; and ##, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g004"></graphic></fig><fig id="cells-09-00122-f005" orientation="portrait" position="float"><label>Figure 5</label><caption><p>Tre relieves CS-induced lysosome damage and disorder of autophagic substrate degradation in vivo. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of LAMP1 protein levels in lung tissue on day 7 or 56 post CS instillation (<italic>n</italic> = 5 to 6). (<bold>C</bold>,<bold>D</bold>) Immunoblotting analysis of LAMP1 protein levels in AMs on day 7 or 56 post CS instillation (<italic>n</italic> = 3 to 4). (<bold>E</bold>,<bold>F</bold>) Immunoblotting analysis of p62 and ubiquitin protein levels in lung tissue on day 7 or 56 post CS instillation (<italic>n</italic> = 5 to 6). (<bold>G</bold>,<bold>H</bold>) Immunoblotting analysis of p62 protein levels in AMs on day 7 or 56 post CS instillation (<italic>n</italic> = 3 to 4). (<bold>I</bold>) Immunofluorescence analysis of p62 in AMs on day 7 or 56 post CS instillation (scale bar = 25 μm and <italic>n</italic> = 3). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05 and **, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g005"></graphic></fig><fig id="cells-09-00122-f006" orientation="portrait" position="float"><label>Figure 6</label><caption><p>Tre relieves CS-induced lysosome damage and restores autophagic substrate degradation through TFEB activation in vitro. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of TFEB levels in the cytosol and nucleus of MH-S cells post CS and Tre treatment and β-actin, loading control for cytosolic proteins; LaminB, loading control for nuclear proteins (<italic>n</italic> = 3 to 4). (<bold>C</bold>) qPCR analysis of TFEB expression in MH-S cells 12 h post-CS and Tre treatment (<italic>n</italic> = 3 to 4). (<bold>D</bold>) MH-S cells were transfected with the adenovirus mRFP-GFP-LC3 plasmid. LC3 puncta were observed by confocal microscopy (scale bar = 10 μm and <italic>n</italic> = 3). (<bold>E</bold>,<bold>F</bold>) Immunoblotting analysis of LC3II, Atg5, and Beclin 1 protein levels (<italic>n</italic> = 3). (<bold>G</bold>,<bold>H</bold>) Immunoblotting analysis of LAMP1 protein levels in cell lysates (<italic>n</italic> = 4). (<bold>I</bold>) Cells were grown on coverslips and stained with Lysotracker Red or acridine orange 12 h post CS and Tre treatment (scale bar = 10 μm and <italic>n</italic> = 3). (<bold>J</bold>) Immunofluorescence analysis of the colocalization of LAMP1 and LC3 12 h post CS and Tre treatment (scale bar = 25 μ and <italic>n</italic> = 3). (<bold>K</bold>–<bold>M</bold>) Immunoblotting analysis of p62 and immunofluorescence analysis of the colocalization of p62 and ubiquitin 12 h post CS and Tre treatment (scale bar = 25 μm and <italic>n</italic> = 3). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05; **, <italic>p</italic> < 0.01; #, <italic>p</italic> < 0.05; and ##, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g006"></graphic></fig><fig id="cells-09-00122-f007" orientation="portrait" position="float"><label>Figure 7</label><caption><p>Tre may relieve CS-induced apoptosis and inflammation may through TFEB activation. (<bold>A</bold>,<bold>B</bold>) Immunoblotting analysis of BAX and BCL-2 protein levels in lungs on day 7 or 56 post CS instillation (<italic>n</italic> = 5). (<bold>C</bold>,<bold>D</bold>) Immunoblotting analysis of BAX and BCL-2 protein levels in AMs on day 7 or 56 post CS instillation (<italic>n</italic> = 3). (<bold>E–H</bold>) ELISA analysis of IL-6, MCP-1, TNF-α, and IL-1β levels in BALF on day 7 or 56 post CS instillation (<italic>n</italic> = 5 to 6). (<bold>I</bold>) H&E staining of mouse lungs on day 7 or 56 post-CS instillation (scale bar = 100 μm and <italic>n</italic> = 5). (<bold>J</bold>,<bold>K</bold>) Immunoblotting analysis of BAX and BCL-2 protein levels in MH-S cell lines (<italic>n</italic> = 3). (<bold>L</bold>,<bold>M</bold>) TUNEL (green) and DAPI (blue) staining and ratios of TUNEL-positive apoptotic MH-S cells (scale bar = 50 μm and <italic>n</italic> = 3). (<bold>N</bold>–<bold>Q</bold>) ELISA analysis of IL-6, MCP-1, TNF-α, and IL-1β levels in MH-S cell supernatant (<italic>n</italic> = 3). Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05 and **, <italic>p</italic> < 0.01.</p></caption><graphic xlink:href="cells-09-00122-g007"></graphic></fig><fig id="cells-09-00122-f008" orientation="portrait" position="float"><label>Figure 8</label><caption><p>Tre relieves CS-induced pulmonary fibrosis. (<bold>A</bold>) HYP content in lung tissue was measured on day 56 post CS instillation (<italic>n</italic> = 5). (<bold>B</bold>,<bold>C</bold>) Immunoblotting analysis of Col-1 and Fn protein levels in mouse lungs on day 56 post CS treatment (<italic>n</italic> = 5 to 6). (<bold>D</bold>,<bold>E</bold>) Immunohistochemical staining and analysis of Col-1 and Fn levels in paraffin sections of mouse lung on day 56 post CS treatment. (<bold>F</bold>) Masson’s trichrome staining of mouse lungs on day 56 (scale bar = 100 μm and <italic>n</italic> = 5). (<bold>G</bold>) Fibrotic score analysis of the lung sections on day 56 post CS instillation. The fibrotic area is presented as a percentage. Data are presented as the mean ± SD. *, <italic>p</italic> < 0.05 and **, <italic>p</italic> < 0.01. (<bold>H</bold>) Illustration of the protective effects of Tre on CS-induced lung inflammation and fibrosis through the activation of the TFEB-dependent autophagy-lysosome machinery. CS impairs lysosomal function, inhibits autophagosome-lysosome fusion, and impairs autophagic substrate degradation. Tre induces TFEB nuclear translocation and improves the function of the autophagy-lysosomal system, leading to decreased apoptosis and secretion of inflammatory factors to alleviate CS-induced lung tissue injury and fibrosis.</p></caption><graphic xlink:href="cells-09-00122-g008"></graphic></fig></floats-group></pmc><affiliations><list></list><tree><noCountry><name sortKey="Ban, Jiaqi" sort="Ban, Jiaqi" uniqKey="Ban J" first="Jiaqi" last="Ban">Jiaqi Ban</name><name sortKey="Chen, Jie" sort="Chen, Jie" uniqKey="Chen J" first="Jie" last="Chen">Jie Chen</name><name sortKey="Chen, Shi" sort="Chen, Shi" uniqKey="Chen S" first="Shi" last="Chen">Shi Chen</name><name sortKey="Chen, Ying" sort="Chen, Ying" uniqKey="Chen Y" first="Ying" last="Chen">Ying Chen</name><name sortKey="He, Xiu" sort="He, Xiu" uniqKey="He X" first="Xiu" last="He">Xiu He</name><name sortKey="He, Yangyang" sort="He, Yangyang" uniqKey="He Y" first="Yangyang" last="He">Yangyang He</name><name sortKey="Li, Chao" sort="Li, Chao" uniqKey="Li C" first="Chao" last="Li">Chao Li</name><name sortKey="Liu, Fangwei" sort="Liu, Fangwei" uniqKey="Liu F" first="Fangwei" last="Liu">Fangwei Liu</name><name sortKey="Wei, Yungeng" sort="Wei, Yungeng" uniqKey="Wei Y" first="Yungeng" last="Wei">Yungeng Wei</name></noCountry></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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