Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats
Background/Aims: The apoptosis of activated hepatic stellate cells (HSCs) is the central event in the reversal of liver fibrosis. K63 de-ubiquitinated receptor-interacting protein (RIP)1 promotes apoptosis in tumor necrosis factor (TNF)-α signaling pathway. In the previous study, we have proved that...
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| Format: | Article |
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Cell Physiol Biochem Press GmbH & Co KG
2017-03-01
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| Series: | Cellular Physiology and Biochemistry |
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| Online Access: | http://www.karger.com/Article/FullText/470650 |
| _version_ | 1811267162705231872 |
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| author | Bin Li Meng Cong Yanan Zhu Ying Xiong Wenyi Jin Yang Wan Yunjiao Zhou Ying Ao Hui Wang |
| author_facet | Bin Li Meng Cong Yanan Zhu Ying Xiong Wenyi Jin Yang Wan Yunjiao Zhou Ying Ao Hui Wang |
| author_sort | Bin Li |
| collection | DOAJ |
| description | Background/Aims: The apoptosis of activated hepatic stellate cells (HSCs) is the central event in the reversal of liver fibrosis. K63 de-ubiquitinated receptor-interacting protein (RIP)1 promotes apoptosis in tumor necrosis factor (TNF)-α signaling pathway. In the previous study, we have proved that indole-3-carbinol (I3C) could reverse different models of liver fibrosis in rats, but the mechanism is still unclear. Thus, the present research aimed to demonstrate the induction of I3C on apoptosis of HSCs and the underlying molecular mechanism. Methods: HSC-T6, an immortalized rat liver stellate cell line, was treated for 24 hours with 25, 50 and 100 µM of I3C. The apoptosis of HSC-T6 was analyzed by flow cytometric analysis, acridine orange staining and RT-PCR, respectively. K63 de-ubiquitination of RIP1 and the expression of ubiquitin ligases and deubiquitinases were analyzed by Co-IP assay and western blot. Knockdown of deubiquitinases was undertaken by small interfering RNA (siRNA). Results: The results of flow cytometric analysis indicated that the apoptotic rate of HSC-T6 was induced by I3C in a dose-dependent manner. Observation by acridine orange staining exhibited nuclear condensation and apoptotic bodies in I3C treated cells. Consistently, the expression ratio of Bax/bcl-2 was markedly increased by I3C. These results indicated that I3C could significantly induce apoptosis of HSC-T6 cells. Furthermore, Co-IP assay revealed K63 de-ubiquitination of RIP1 by I3C, associated with the induction of caspase 8. Although I3C had no effect on the expression of ubiquitin ligases cellular inhibitor of apoptosis 1/2 (cIAP1/2), the protein level of deubiquitinase cylindromatosis (CYLD) was significantly induced by I3C. Moreover, CYLD silencing reversed the pro-apoptosis induction effect of I3C and reduced the expression ratio of Bax/bcl-2 in HSC-T6 cells. Conclusion: These results demonstrated that I3C could induce apoptosis of HSC through RIP1 K63 de-ubiquitination by upregulating deubiquitinase CYLD. |
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| id | doaj.art-e885fdf4c4d748efab1a6840cecc7172 |
| institution | Directory Open Access Journal |
| issn | 1015-8987 1421-9778 |
| language | English |
| last_indexed | 2024-04-12T20:56:23Z |
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| series | Cellular Physiology and Biochemistry |
| spelling | doaj.art-e885fdf4c4d748efab1a6840cecc71722022-12-22T03:16:58ZengCell Physiol Biochem Press GmbH & Co KGCellular Physiology and Biochemistry1015-89871421-97782017-03-014141481149010.1159/000470650470650Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in RatsBin LiMeng CongYanan ZhuYing XiongWenyi JinYang WanYunjiao ZhouYing AoHui WangBackground/Aims: The apoptosis of activated hepatic stellate cells (HSCs) is the central event in the reversal of liver fibrosis. K63 de-ubiquitinated receptor-interacting protein (RIP)1 promotes apoptosis in tumor necrosis factor (TNF)-α signaling pathway. In the previous study, we have proved that indole-3-carbinol (I3C) could reverse different models of liver fibrosis in rats, but the mechanism is still unclear. Thus, the present research aimed to demonstrate the induction of I3C on apoptosis of HSCs and the underlying molecular mechanism. Methods: HSC-T6, an immortalized rat liver stellate cell line, was treated for 24 hours with 25, 50 and 100 µM of I3C. The apoptosis of HSC-T6 was analyzed by flow cytometric analysis, acridine orange staining and RT-PCR, respectively. K63 de-ubiquitination of RIP1 and the expression of ubiquitin ligases and deubiquitinases were analyzed by Co-IP assay and western blot. Knockdown of deubiquitinases was undertaken by small interfering RNA (siRNA). Results: The results of flow cytometric analysis indicated that the apoptotic rate of HSC-T6 was induced by I3C in a dose-dependent manner. Observation by acridine orange staining exhibited nuclear condensation and apoptotic bodies in I3C treated cells. Consistently, the expression ratio of Bax/bcl-2 was markedly increased by I3C. These results indicated that I3C could significantly induce apoptosis of HSC-T6 cells. Furthermore, Co-IP assay revealed K63 de-ubiquitination of RIP1 by I3C, associated with the induction of caspase 8. Although I3C had no effect on the expression of ubiquitin ligases cellular inhibitor of apoptosis 1/2 (cIAP1/2), the protein level of deubiquitinase cylindromatosis (CYLD) was significantly induced by I3C. Moreover, CYLD silencing reversed the pro-apoptosis induction effect of I3C and reduced the expression ratio of Bax/bcl-2 in HSC-T6 cells. Conclusion: These results demonstrated that I3C could induce apoptosis of HSC through RIP1 K63 de-ubiquitination by upregulating deubiquitinase CYLD.http://www.karger.com/Article/FullText/470650Indole-3-carbinolHepatic stellate cellsReceptor-interacting protein 1K63 de-ubiquitinationCylindromatosis |
| spellingShingle | Bin Li Meng Cong Yanan Zhu Ying Xiong Wenyi Jin Yang Wan Yunjiao Zhou Ying Ao Hui Wang Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats Cellular Physiology and Biochemistry Indole-3-carbinol Hepatic stellate cells Receptor-interacting protein 1 K63 de-ubiquitination Cylindromatosis |
| title | Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats |
| title_full | Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats |
| title_fullStr | Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats |
| title_full_unstemmed | Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats |
| title_short | Indole-3-Carbinol Induces Apoptosis of Hepatic Stellate Cells through K63 De-Ubiquitination of RIP1 in Rats |
| title_sort | indole 3 carbinol induces apoptosis of hepatic stellate cells through k63 de ubiquitination of rip1 in rats |
| topic | Indole-3-carbinol Hepatic stellate cells Receptor-interacting protein 1 K63 de-ubiquitination Cylindromatosis |
| url | http://www.karger.com/Article/FullText/470650 |
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