Huaxin Chen1, Jianye Cai2, Jiancheng Wang3, Yuan Qiu4, Chenhao Jiang4, Yi Wang4, Yiqin Wang4, Chenju Yi5, Lijie Pan1, Yuanjun Guan6, Jun Zheng7, Dongbo Qiu8, Cong Du8, Qiuli Liu9, Guihua Chen7, Yang Yang10, Yan Xu11, Andy Peng Xiang12, Qi Zhang13. 1. Biotherapy Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China; Cell-gene Therapy Translational Medicine Research Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. 2. Centre for Stem Cell Biology and Tissue Engineering, Key Laboratory for Stem Cells and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, China; Department of Hepatic Surgery and Liver Transplantation Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. 3. Centre for Stem Cell Biology and Tissue Engineering, Key Laboratory for Stem Cells and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, China; Scientific Research Centre, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, China. 4. Centre for Stem Cell Biology and Tissue Engineering, Key Laboratory for Stem Cells and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, China. 5. Scientific Research Centre, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, China. 6. Core Facility Centre, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China. 7. Department of Hepatic Surgery and Liver Transplantation Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. 8. Biotherapy Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China; Cell-gene Therapy Translational Medicine Research Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China; Guangdong Provincial Key Laboratory of Liver Disease Research, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. 9. Biotherapy Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. 10. Department of Hepatic Surgery and Liver Transplantation Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. Electronic address: yysysu@163.com. 11. Biotherapy Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. Electronic address: xuyan55@mail.sysu.edu.cn. 12. Centre for Stem Cell Biology and Tissue Engineering, Key Laboratory for Stem Cells and Tissue Engineering, Ministry of Education, Sun Yat-sen University, Guangzhou, China. Electronic address: xiangp@mail.sysu.edu.cn. 13. Biotherapy Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China; Cell-gene Therapy Translational Medicine Research Centre, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China; Guangdong Provincial Key Laboratory of Liver Disease Research, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China. Electronic address: zhangq27@mail.sysu.edu.cn.
Abstract
BACKGROUND & AIMS: Liver fibrosis is a wound healing response that arises from various aetiologies. The intermediate filament protein Nestin has been reported to participate in maintaining tissue homeostasis during wound healing responses. However, little is known about the role Nestin plays in liver fibrosis. This study investigated the function and precise regulatory network of Nestin during liver fibrosis. METHODS: Nestin expression was assessed via immunostaining and quantitative real-time PCR (qPCR) in fibrotic/cirrhotic samples. The induction of Nestin expression by transforming growth factor beta (TGFβ)-Smad2/3 signalling was investigated through luciferase reporter assays. The functional role of Nestin in hepatic stellate cells (HSCs) was investigated by examining the pathway activity of profibrogenic TGFβ-Smad2/3 signalling and degradation of TGFβ receptor I (TβRI) after interfering with Nestin. The in vivo effects of knocking down Nestin were examined with an adeno-associated virus vector (serotype 6, AAV6) carrying short-hairpin RNA targeting Nestin in fibrotic mouse models. RESULTS: Nestin was mainly expressed in activated HSCs and increased with the progression of liver fibrosis. The profibrogenic pathway TGFβ-Smad2/3 induced Nestin expression directly. Knocking down Nestin promoted caveolin 1-mediated TβRI degradation, resulting in TGFβ-Smad2/3 pathway impairment and reduced fibrosis marker expression in HSCs. In AAV6-treated murine fibrotic models, knocking down Nestin resulted in decreased levels of inflammatory infiltration, hepatocellular damage, and a reduced degree of fibrosis. CONCLUSION: The expression of Nestin in HSCs was induced by TGFβ and positively correlated with the degree of liver fibrosis. Knockdown of Nestin decreased activation of the TGFβ pathway and alleviated liver fibrosis both in vitro and in vivo. Our data demonstrate a novel role of Nestin in controlling HSC activation in liver fibrosis. LAY SUMMARY: Liver fibrosis has various aetiologies but represents a common process in chronic liver diseases that is associated with high morbidity and mortality. Herein, we demonstrate that the intermediate filament protein Nestin plays an essential profibrogenic role in liver fibrosis by forming a positive feedback loop with the TGFβ-Smad2/3 pathway, providing a potential therapeutic target for the treatment of liver fibrosis.
BACKGROUND & AIMS: Liver fibrosis is a wound healing response that arises from various aetiologies. The intermediate filament protein Nestin has been reported to participate in maintaining tissue homeostasis during wound healing responses. However, little is known about the role Nestin plays in liver fibrosis. This study investigated the function and precise regulatory network of Nestin during liver fibrosis. METHODS: Nestin expression was assessed via immunostaining and quantitative real-time PCR (qPCR) in fibrotic/cirrhotic samples. The induction of Nestin expression by transforming growth factor beta (TGFβ)-Smad2/3 signalling was investigated through luciferase reporter assays. The functional role of Nestin in hepatic stellate cells (HSCs) was investigated by examining the pathway activity of profibrogenic TGFβ-Smad2/3 signalling and degradation of TGFβ receptor I (TβRI) after interfering with Nestin. The in vivo effects of knocking down Nestin were examined with an adeno-associated virus vector (serotype 6, AAV6) carrying short-hairpin RNA targeting Nestin in fibrotic mouse models. RESULTS: Nestin was mainly expressed in activated HSCs and increased with the progression of liver fibrosis. The profibrogenic pathway TGFβ-Smad2/3 induced Nestin expression directly. Knocking down Nestin promoted caveolin 1-mediated TβRI degradation, resulting in TGFβ-Smad2/3 pathway impairment and reduced fibrosis marker expression in HSCs. In AAV6-treated murine fibrotic models, knocking down Nestin resulted in decreased levels of inflammatory infiltration, hepatocellular damage, and a reduced degree of fibrosis. CONCLUSION: The expression of Nestin in HSCs was induced by TGFβ and positively correlated with the degree of liver fibrosis. Knockdown of Nestin decreased activation of the TGFβ pathway and alleviated liver fibrosis both in vitro and in vivo. Our data demonstrate a novel role of Nestin in controlling HSC activation in liver fibrosis. LAY SUMMARY: Liver fibrosis has various aetiologies but represents a common process in chronic liver diseases that is associated with high morbidity and mortality. Herein, we demonstrate that the intermediate filament protein Nestin plays an essential profibrogenic role in liver fibrosis by forming a positive feedback loop with the TGFβ-Smad2/3 pathway, providing a potential therapeutic target for the treatment of liver fibrosis.
Authors: Patricia P Bloom; Trisha S Pasricha; Karin L Andersson; Daniel S Pratt; Nikroo Hashemi; Irun Bhan; Kathleen Viveiros Journal: Dig Dis Sci Date: 2020-07-29 Impact factor: 3.199