Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion

Xiaojie Yu; Natalia Elfimova; Marion Müller; Daniel Bachurski; Ulrike Koitzsch; Uta Drebber; Esther Mahabir; Hinrich P. Hansen; Scott L. Friedman; Sabine Klein; Hans Peter Dienes; Marianna Hösel; Reinhard Buettner; Jonel Trebicka; Vangelis Kondylis; Inge Mannaerts; Margarete Odenthal

doi:10.1016/j.jcmgh.2022.02.013

Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion

Xiaojie Yu, Natalia Elfimova, Marion Müller, Daniel Bachurski, Ulrike Koitzsch, Uta Drebber, Esther Mahabir, Hinrich P. Hansen, Scott L. Friedman, Sabine Klein, Hans Peter Dienes, Marianna Hösel, Reinhard Buettner, Jonel Trebicka, Vangelis Kondylis, Inge Mannaerts, Margarete Odenthal

Research output: Contribution to journal › Article › peer-review

2 Scopus citations

Abstract

Background & Aims: Liver fibrosis arises from long-term chronic liver injury, accompanied by an accelerated wound healing response with interstitial accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HSC) are the main source for ECM production. MicroRNA29a (miR-29a) is a crucial antifibrotic miRNA that is repressed during fibrosis, resulting in up-regulation of collagen synthesis. Methods: Intracellular and extracellular miRNA levels of primary and immortalized myofibroblastic HSC in response to profibrogenic stimulation by transforming growth factor β (TGFβ) or platelet-derived growth factor-BB (PDGF-BB) or upon inhibition of vesicular transport and autophagy processes were determined by quantitative polymerase chain reaction. Autophagy flux was studied by electron microscopy, flow cytometry, immunoblotting, and immunocytochemistry. Hepatic and serum miR-29a levels were quantified by using both liver tissue and serum samples from a cohort of chronic hepatitis C virus patients and a murine CCl₄ induced liver fibrosis model. Results: In our study, we show that TGFβ and PDGF-BB resulted in decrease of intracellular miR-29a and a pronounced increase of vesicular miR-29a release into the supernatant. Strikingly, miR-29a vesicular release was accompanied by enhanced autophagic activity and up-regulation of the autophagy marker protein LC3. Moreover, autophagy inhibition strongly prevented miR-29a secretion and repressed its targets’ expression such as Col1A1. Consistently, hepatic miR-29a loss and increased LC3 expression in myofibroblastic HSC were associated with increased serum miR-29a levels in CCl₄-treated murine liver fibrosis and specimens of hepatitis C virus patients with chronic liver disease. Conclusions: We provide evidence that activation-associated autophagy in HSC induces release of miR-29a, whereas inhibition of autophagy represses fibrogenic gene expression in part through attenuated miR-29a secretion.

Original language	English
Pages (from-to)	1701-1716
Number of pages	16
Journal	Cellular and Molecular Gastroenterology and Hepatology
Volume	13
Issue number	6
DOIs	https://doi.org/10.1016/j.jcmgh.2022.02.013
State	Published - Jan 2022

Keywords

Circulating MiRNA
Liver Fibrosis
MiR-29

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10.1016/j.jcmgh.2022.02.013

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Yu, X., Elfimova, N., Müller, M., Bachurski, D., Koitzsch, U., Drebber, U., Mahabir, E., Hansen, H. P., Friedman, S. L., Klein, S., Dienes, H. P., Hösel, M., Buettner, R., Trebicka, J., Kondylis, V., Mannaerts, I., & Odenthal, M. (2022). Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion. Cellular and Molecular Gastroenterology and Hepatology, 13(6), 1701-1716. https://doi.org/10.1016/j.jcmgh.2022.02.013

@article{152ba960f17e48eaa29559c38b965ed2,

title = "Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion",

abstract = "Background & Aims: Liver fibrosis arises from long-term chronic liver injury, accompanied by an accelerated wound healing response with interstitial accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HSC) are the main source for ECM production. MicroRNA29a (miR-29a) is a crucial antifibrotic miRNA that is repressed during fibrosis, resulting in up-regulation of collagen synthesis. Methods: Intracellular and extracellular miRNA levels of primary and immortalized myofibroblastic HSC in response to profibrogenic stimulation by transforming growth factor β (TGFβ) or platelet-derived growth factor-BB (PDGF-BB) or upon inhibition of vesicular transport and autophagy processes were determined by quantitative polymerase chain reaction. Autophagy flux was studied by electron microscopy, flow cytometry, immunoblotting, and immunocytochemistry. Hepatic and serum miR-29a levels were quantified by using both liver tissue and serum samples from a cohort of chronic hepatitis C virus patients and a murine CCl4 induced liver fibrosis model. Results: In our study, we show that TGFβ and PDGF-BB resulted in decrease of intracellular miR-29a and a pronounced increase of vesicular miR-29a release into the supernatant. Strikingly, miR-29a vesicular release was accompanied by enhanced autophagic activity and up-regulation of the autophagy marker protein LC3. Moreover, autophagy inhibition strongly prevented miR-29a secretion and repressed its targets{\textquoteright} expression such as Col1A1. Consistently, hepatic miR-29a loss and increased LC3 expression in myofibroblastic HSC were associated with increased serum miR-29a levels in CCl4-treated murine liver fibrosis and specimens of hepatitis C virus patients with chronic liver disease. Conclusions: We provide evidence that activation-associated autophagy in HSC induces release of miR-29a, whereas inhibition of autophagy represses fibrogenic gene expression in part through attenuated miR-29a secretion.",

keywords = "Circulating MiRNA, Liver Fibrosis, MiR-29",

author = "Xiaojie Yu and Natalia Elfimova and Marion M{\"u}ller and Daniel Bachurski and Ulrike Koitzsch and Uta Drebber and Esther Mahabir and Hansen, {Hinrich P.} and Friedman, {Scott L.} and Sabine Klein and Dienes, {Hans Peter} and Marianna H{\"o}sel and Reinhard Buettner and Jonel Trebicka and Vangelis Kondylis and Inge Mannaerts and Margarete Odenthal",

note = "Funding Information: Funding Supported by German Liver Foundation (MO), the Heinrich Hertz Foundation (MO), the Center for Molecular Medicine Cologne (CMMC) to RB, MO, and the German Research Foundation (DFG OD/6-1) to MO, (SFB TRR57/ P18) to JT, (DFG KO 5827/2-1) to VK, and (DFG HA2432/3-2) to HH. In addition, financial support was received from the Research and Education programme of the Medical Faculty of the University of Cologne and the German Competence Network for Viral Hepatitis (HepNet), funded by the German Federal Ministry of Education and Research (BMBF), Grant No 01KI0601 (HPD/MO). Jonel Trebicka is supported by the European Union{\textquoteright}s Horizon 2020 Research and Innovation Program (Galaxy, No. 668031, MICROB-PREDICT, No. 825694 and DECISION No.84794), and Societal Challenges - Health, Demographic Change and Wellbeing (No. 731875), and Cellex Foundation (PREDICT). S.L.F. is supported by NIH grant R01 DK128289 and I.M. by a FWO–V post-doctoral fellowship (12N5419 N). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Funding Information: Funding Supported by German Liver Foundation (MO), the Heinrich Hertz Foundation (MO), the Center for Molecular Medicine Cologne (CMMC) to RB, MO, and the German Research Foundation (DFG OD/6-1) to MO, (SFB TRR57/ P18) to JT, (DFG KO 5827/2-1) to VK, and (DFG HA2432/3-2) to HH. In addition, financial support was received from the Research and Education programme of the Medical Faculty of the University of Cologne and the German Competence Network for Viral Hepatitis (HepNet), funded by the German Federal Ministry of Education and Research (BMBF), Grant No 01KI0601 (HPD/MO). Jonel Trebicka is supported by the European Union's Horizon 2020 Research and Innovation Program (Galaxy, No. 668031, MICROB-PREDICT, No. 825694 and DECISION No.84794), and Societal Challenges - Health, Demographic Change and Wellbeing (No. 731875), and Cellex Foundation (PREDICT). S.L.F. is supported by NIH grant R01 DK128289 and I.M. by a FWO?V post-doctoral fellowship (12N5419 N). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Publisher Copyright: {\textcopyright} 2022 The Authors",

year = "2022",

month = jan,

doi = "10.1016/j.jcmgh.2022.02.013",

language = "English",

volume = "13",

pages = "1701--1716",

journal = "Cellular and Molecular Gastroenterology and Hepatology",

issn = "2352-345X",

publisher = "Elsevier Inc.",

number = "6",

}

Yu, X, Elfimova, N, Müller, M, Bachurski, D, Koitzsch, U, Drebber, U, Mahabir, E, Hansen, HP, Friedman, SL, Klein, S, Dienes, HP, Hösel, M, Buettner, R, Trebicka, J, Kondylis, V, Mannaerts, I & Odenthal, M 2022, 'Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion', Cellular and Molecular Gastroenterology and Hepatology, vol. 13, no. 6, pp. 1701-1716. https://doi.org/10.1016/j.jcmgh.2022.02.013

TY - JOUR

T1 - Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion

AU - Yu, Xiaojie

AU - Elfimova, Natalia

AU - Müller, Marion

AU - Bachurski, Daniel

AU - Koitzsch, Ulrike

AU - Drebber, Uta

AU - Mahabir, Esther

AU - Hansen, Hinrich P.

AU - Friedman, Scott L.

AU - Klein, Sabine

AU - Dienes, Hans Peter

AU - Hösel, Marianna

AU - Buettner, Reinhard

AU - Trebicka, Jonel

AU - Kondylis, Vangelis

AU - Mannaerts, Inge

AU - Odenthal, Margarete

N1 - Funding Information: Funding Supported by German Liver Foundation (MO), the Heinrich Hertz Foundation (MO), the Center for Molecular Medicine Cologne (CMMC) to RB, MO, and the German Research Foundation (DFG OD/6-1) to MO, (SFB TRR57/ P18) to JT, (DFG KO 5827/2-1) to VK, and (DFG HA2432/3-2) to HH. In addition, financial support was received from the Research and Education programme of the Medical Faculty of the University of Cologne and the German Competence Network for Viral Hepatitis (HepNet), funded by the German Federal Ministry of Education and Research (BMBF), Grant No 01KI0601 (HPD/MO). Jonel Trebicka is supported by the European Union’s Horizon 2020 Research and Innovation Program (Galaxy, No. 668031, MICROB-PREDICT, No. 825694 and DECISION No.84794), and Societal Challenges - Health, Demographic Change and Wellbeing (No. 731875), and Cellex Foundation (PREDICT). S.L.F. is supported by NIH grant R01 DK128289 and I.M. by a FWO–V post-doctoral fellowship (12N5419 N). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Funding Information: Funding Supported by German Liver Foundation (MO), the Heinrich Hertz Foundation (MO), the Center for Molecular Medicine Cologne (CMMC) to RB, MO, and the German Research Foundation (DFG OD/6-1) to MO, (SFB TRR57/ P18) to JT, (DFG KO 5827/2-1) to VK, and (DFG HA2432/3-2) to HH. In addition, financial support was received from the Research and Education programme of the Medical Faculty of the University of Cologne and the German Competence Network for Viral Hepatitis (HepNet), funded by the German Federal Ministry of Education and Research (BMBF), Grant No 01KI0601 (HPD/MO). Jonel Trebicka is supported by the European Union's Horizon 2020 Research and Innovation Program (Galaxy, No. 668031, MICROB-PREDICT, No. 825694 and DECISION No.84794), and Societal Challenges - Health, Demographic Change and Wellbeing (No. 731875), and Cellex Foundation (PREDICT). S.L.F. is supported by NIH grant R01 DK128289 and I.M. by a FWO?V post-doctoral fellowship (12N5419 N). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Publisher Copyright: © 2022 The Authors

PY - 2022/1

Y1 - 2022/1

N2 - Background & Aims: Liver fibrosis arises from long-term chronic liver injury, accompanied by an accelerated wound healing response with interstitial accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HSC) are the main source for ECM production. MicroRNA29a (miR-29a) is a crucial antifibrotic miRNA that is repressed during fibrosis, resulting in up-regulation of collagen synthesis. Methods: Intracellular and extracellular miRNA levels of primary and immortalized myofibroblastic HSC in response to profibrogenic stimulation by transforming growth factor β (TGFβ) or platelet-derived growth factor-BB (PDGF-BB) or upon inhibition of vesicular transport and autophagy processes were determined by quantitative polymerase chain reaction. Autophagy flux was studied by electron microscopy, flow cytometry, immunoblotting, and immunocytochemistry. Hepatic and serum miR-29a levels were quantified by using both liver tissue and serum samples from a cohort of chronic hepatitis C virus patients and a murine CCl4 induced liver fibrosis model. Results: In our study, we show that TGFβ and PDGF-BB resulted in decrease of intracellular miR-29a and a pronounced increase of vesicular miR-29a release into the supernatant. Strikingly, miR-29a vesicular release was accompanied by enhanced autophagic activity and up-regulation of the autophagy marker protein LC3. Moreover, autophagy inhibition strongly prevented miR-29a secretion and repressed its targets’ expression such as Col1A1. Consistently, hepatic miR-29a loss and increased LC3 expression in myofibroblastic HSC were associated with increased serum miR-29a levels in CCl4-treated murine liver fibrosis and specimens of hepatitis C virus patients with chronic liver disease. Conclusions: We provide evidence that activation-associated autophagy in HSC induces release of miR-29a, whereas inhibition of autophagy represses fibrogenic gene expression in part through attenuated miR-29a secretion.

AB - Background & Aims: Liver fibrosis arises from long-term chronic liver injury, accompanied by an accelerated wound healing response with interstitial accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HSC) are the main source for ECM production. MicroRNA29a (miR-29a) is a crucial antifibrotic miRNA that is repressed during fibrosis, resulting in up-regulation of collagen synthesis. Methods: Intracellular and extracellular miRNA levels of primary and immortalized myofibroblastic HSC in response to profibrogenic stimulation by transforming growth factor β (TGFβ) or platelet-derived growth factor-BB (PDGF-BB) or upon inhibition of vesicular transport and autophagy processes were determined by quantitative polymerase chain reaction. Autophagy flux was studied by electron microscopy, flow cytometry, immunoblotting, and immunocytochemistry. Hepatic and serum miR-29a levels were quantified by using both liver tissue and serum samples from a cohort of chronic hepatitis C virus patients and a murine CCl4 induced liver fibrosis model. Results: In our study, we show that TGFβ and PDGF-BB resulted in decrease of intracellular miR-29a and a pronounced increase of vesicular miR-29a release into the supernatant. Strikingly, miR-29a vesicular release was accompanied by enhanced autophagic activity and up-regulation of the autophagy marker protein LC3. Moreover, autophagy inhibition strongly prevented miR-29a secretion and repressed its targets’ expression such as Col1A1. Consistently, hepatic miR-29a loss and increased LC3 expression in myofibroblastic HSC were associated with increased serum miR-29a levels in CCl4-treated murine liver fibrosis and specimens of hepatitis C virus patients with chronic liver disease. Conclusions: We provide evidence that activation-associated autophagy in HSC induces release of miR-29a, whereas inhibition of autophagy represses fibrogenic gene expression in part through attenuated miR-29a secretion.

KW - Circulating MiRNA

KW - Liver Fibrosis

KW - MiR-29

UR - http://www.scopus.com/inward/record.url?scp=85128387181&partnerID=8YFLogxK

U2 - 10.1016/j.jcmgh.2022.02.013

DO - 10.1016/j.jcmgh.2022.02.013

M3 - Article

C2 - 35219894

AN - SCOPUS:85128387181

VL - 13

SP - 1701

EP - 1716

JO - Cellular and Molecular Gastroenterology and Hepatology

JF - Cellular and Molecular Gastroenterology and Hepatology

SN - 2352-345X

IS - 6

ER -

Autophagy-Related Activation of Hepatic Stellate Cells Reduces Cellular miR-29a by Promoting Its Vesicular Secretion

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