Autophagy mediated by endoplasmic reticulum stress enhances the caffeine-induced apoptosis of hepatic stellate cells

Li,Yongjian; Chen,Yunyang; Huang,Haiyan; Shi,Minmin; Yang,Weiping; Kuang,Jie; Yan,Jiqi

doi:10.3892/ijmm.2017.3145

Autophagy mediated by endoplasmic reticulum stress enhances the caffeine-induced apoptosis of hepatic stellate cells

Authors:
- Yongjian Li
- Yunyang Chen
- Haiyan Huang
- Minmin Shi
- Weiping Yang
- Jie Kuang
- Jiqi Yan
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Affiliations: Department of General Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, P.R. China, Shanghai Institute of Digestive Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, P.R. China
Published online on: September 20, 2017 https://doi.org/10.3892/ijmm.2017.3145
Pages: 1405-1414
Copyright: © Li et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Caffeine has been identified to have beneficial effects against chronic liver diseases, particularly liver fibrosis. Many studies have reported that caffeine ameliorates liver fibrosis by directly inducing hepatic stellate cell (HSC) apoptosis; however, the molecular mechanisms involved in this process remain unclear. The presents study aimed to detect the underlying mechanisms by which caffeine mediates HSC apoptosis and eliminates activated HSCs. For this purpose, the LX-2 cell line was applied in this study and the cells were exposed to various concentrations of caffeine for the indicated times. The effects of caffeine on cell viability and apoptosis were assessed by Cell Counting Kit-8 assay and flow cytometry, respectively. Autophagy and endoplasmic reticulum (ER) stress were explored by morphological assessment and western blotting. In the present study, caffeine was found to inhibit the viability and increase the apoptosis of the LX-2 cells in dose- and time-dependent manners. ER stress was stimulated by caffeine as demonstrated by increased levels of GRP78/Bip, CHOP and inositol‑requiring enzyme 1 (IRE1)-α as well as many enlarged ERs detected by electron microscopy. Caffeine induced autophagy as shown by increased p62 and LC3Ⅱ accumulation and the number of GFP/RFP-LC3 puncta and autophagosomes/autolysosomes. Moreover, IRE1-α knockdown decreased the level of autophagic flux, and inhibition of autophagy protected LX-2 cells from apoptotic death. In conclusion, our study showed that the caffeine-enhanced autophagic flux in HSCs was stimulated by ER stress via the IRE1-α signaling pathway, which further weakened HSC viability via the induction of apoptosis. These findings provide new insight into the mechanism of caffeine's anti-fibrotic effects.

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1	Tsochatzis EA, Bosch J and Burroughs AK: Liver cirrhosis. Lancet. 383:1749–1761. 2014. View Article : Google Scholar : PubMed/NCBI
2	Friedman SL: Hepatic stellate cells: protean, multifunctional, and enigmatic cells of the liver. Physiol Rev. 88:125–172. 2008. View Article : Google Scholar : PubMed/NCBI
3	Mederacke I, Hsu CC, Troeger JS, Huebener P, Mu X, Dapito DH, Pradere JP and Schwabe RF: Fate tracing reveals hepatic stellate cells as dominant contributors to liver fibrosis independent of its aetiology. Nat Commun. 4:28232013. View Article : Google Scholar : PubMed/NCBI
4	Thoen LF, Guimarães EL, Dollé L, Mannaerts I, Najimi M, Sokal E and van Grunsven LA: A role for autophagy during hepatic stellate cell activation. J Hepatol. 55:1353–1360. 2011. View Article : Google Scholar : PubMed/NCBI
5	Parola M, Marra F and Pinzani M: Myofibroblast-like cells and liver fibrogenesis: emerging concepts in a rapidly moving scenario. Mol Aspects Med. 29:58–66. 2008. View Article : Google Scholar
6	Glick D, Barth S and Macleod KF: Autophagy: cellular and molecular mechanisms. J Pathol. 221:3–12. 2010. View Article : Google Scholar : PubMed/NCBI
7	Hernández-Gea V, Hilscher M, Rozenfeld R, Lim MP, Nieto N, Werner S, Devi LA and Friedman SL: Endoplasmic reticulum stress induces fibrogenic activity in hepatic stellate cells through autophagy. J Hepatol. 59:98–104. 2013. View Article : Google Scholar : PubMed/NCBI
8	Ding WX and Yin XM: Sorting, recognition and activation of the misfolded protein degradation pathways through macroautophagy and the proteasome. Autophagy. 4:141–150. 2008. View Article : Google Scholar
9	Bernales S, Papa FR and Walter P: Intracellular signaling by the unfolded protein response. Annu Rev Cell Dev Biol. 22:487–508. 2006. View Article : Google Scholar : PubMed/NCBI
10	Momoi T: Conformational diseases and ER stress-mediated cell death: apoptotic cell death and autophagic cell death. Curr Mol Med. 6:111–118. 2006. View Article : Google Scholar : PubMed/NCBI
11	Koo JH, Lee HJ, Kim W and Kim SG: Endoplasmic reticulum stress in hepatic stellate cells promotes liver fibrosis via PERK-mediated degradation of HNRNPA1 and up-regulation of SMAD2. Gastroenterology. 150:181–193.e8. 2016. View Article : Google Scholar
12	Ron D and Walter P: Signal integration in the endoplasmic reticulum unfolded protein response. Nat Rev Mol Cell Biol. 8:519–529. 2007. View Article : Google Scholar : PubMed/NCBI
13	Xu C, Bailly-Maitre B and Reed JC: Endoplasmic reticulum stress: cell life and death decisions. J Clin Invest. 115:2656–2664. 2005. View Article : Google Scholar : PubMed/NCBI
14	Meares GP, Mines MA, Beurel E, Eom TY, Song L, Zmijewska AA and Jope RS: Glycogen synthase kinase-3 regulates endoplasmic reticulum (ER) stress-induced CHOP expression in neuronal cells. Exp Cell Res. 317:1621–1628. 2011. View Article : Google Scholar : PubMed/NCBI
15	Choi AY, Choi JH, Yoon H, Hwang KY, Noh MH, Choe W, Yoon KS, Ha J, Yeo EJ and Kang I: Luteolin induces apoptosis through endoplasmic reticulum stress and mitochondrial dysfunction in Neuro-2a mouse neuroblastoma cells. Eur J Pharmacol. 668:115–126. 2011. View Article : Google Scholar : PubMed/NCBI
16	Hsu SJ, Lee FY, Wang SS, Hsin IF, Lin TY, Huang HC, Chang CC, Chuang CL, Ho HL, Lin HC, et al: Caffeine ameliorates hemodynamic derangements and portosystemic collaterals in cirrhotic rats. Hepatology. 61:1672–1684. 2015. View Article : Google Scholar : PubMed/NCBI
17	Gressner OA, Lahme B, Rehbein K, Siluschek M, Weiskirchen R and Gressner AM: Pharmacological application of caffeine inhibits TGF-β-stimulated connective tissue growth factor expression in hepatocytes via PPARgamma and SMAD2/3-dependent pathways. J Hepatol. 49:758–767. 2008. View Article : Google Scholar : PubMed/NCBI
18	Shim SG, Jun DW, Kim EK, Saeed WK, Lee KN, Lee HL, Lee OY, Choi HS and Yoon BC: Caffeine attenuates liver fibrosis via defective adhesion of hepatic stellate cells in cirrhotic model. J Gastroenterol Hepatol. 28:1877–1884. 2013. View Article : Google Scholar : PubMed/NCBI
19	Saiki S, Sasazawa Y, Imamichi Y, Kawajiri S, Fujimaki T, Tanida I, Kobayashi H, Sato F, Sato S, Ishikawa K, et al: Caffeine induces apoptosis by enhancement of autophagy via PI3K/Akt/mTOR/p70S6K inhibition. Autophagy. 7:176–187. 2011. View Article : Google Scholar :
20	Yan X, Shan Z, Yan L, Zhu Q, Liu L, Xu B, Liu S, Jin Z and Gao Y: High expression of zinc-finger protein X-linked promotes tumor growth and predicts a poor outcome for stage II/III colorectal cancer patients. Oncotarget. 7:19680–19692. 2016. View Article : Google Scholar : PubMed/NCBI
21	Ou B, Zhao J, Guan S, Wangpu X, Zhu C, Zong Y, Ma J, Sun J, Zheng M, Feng H, et al: Plk2 promotes tumor growth and inhibits apoptosis by targeting Fbxw7/Cyclin E in colorectal cancer. Cancer Lett. 380:457–466. 2016. View Article : Google Scholar : PubMed/NCBI
22	Shi YH, Ding ZB, Zhou J, Hui B, Shi GM, Ke AW, Wang XY, Dai Z, Peng YF, Gu CY, et al: Targeting autophagy enhances sorafenib lethality for hepatocellular carcinoma via ER stress-related apoptosis. Autophagy. 7:1159–1172. 2011. View Article : Google Scholar : PubMed/NCBI
23	Ding WX, Ni HM, Gao W, Hou YF, Melan MA, Chen X, Stolz DB, Shao ZM and Yin XM: Differential effects of endoplasmic reticulum stress-induced autophagy on cell survival. J Biol Chem. 282:4702–4710. 2007. View Article : Google Scholar
24	Xu WH, Liu ZB, Hou YF, Hong Q, Hu DL and Shao ZM: Inhibition of autophagy enhances the cytotoxic effect of PA-MSHA in breast cancer. BMC Cancer. 14:2732014. View Article : Google Scholar : PubMed/NCBI
25	Friedrich K, Smit M, Wannhoff A, Rupp C, Scholl SG, Antoni C, Dollinger M, Neumann-Haefelin C, Stremmel W, Weiss KH, et al: Coffee consumption protects against progression in liver cirrhosis and increases long-term survival after liver transplantation. J Gastroenterol Hepatol. 31:1470–1475. 2016. View Article : Google Scholar : PubMed/NCBI
26	Wadhawan M and Anand AC: Coffee and liver disease. J Clin Exp Hepatol. 6:40–46. 2016. View Article : Google Scholar : PubMed/NCBI
27	Sinha RA, Farah BL, Singh BK, Siddique MM, Li Y, Wu Y, Ilkayeva OR, Gooding J, Ching J, Zhou J, et al: Caffeine stimulates hepatic lipid metabolism by the autophagy-lysosomal pathway in mice. Hepatology. 59:1366–1380. 2014. View Article : Google Scholar
28	Elsharkawy AM, Oakley F and Mann DA: The role and regulation of hepatic stellate cell apoptosis in reversal of liver fibrosis. Apoptosis. 10:927–939. 2005. View Article : Google Scholar : PubMed/NCBI
29	Jeschke MG, Gauglitz GG, Song J, Kulp GA, Finnerty CC, Cox RA, Barral JM, Herndon DN and Boehning D: Calcium and ER stress mediate hepatic apoptosis after burn injury. J Cell Mol Med. 13(8B): 1857–1865. 2009. View Article : Google Scholar
30	Ji C: Dissection of endoplasmic reticulum stress signaling in alcoholic and non-alcoholic liver injury. J Gastroenterol Hepatol. 23(Suppl 1): S16–S24. 2008. View Article : Google Scholar : PubMed/NCBI
31	Wang Y, Gao J, Zhang D, Zhang J, Ma J and Jiang H: New insights into the antifibrotic effects of sorafenib on hepatic stellate cells and liver fibrosis. J Hepatol. 53:132–144. 2010. View Article : Google Scholar : PubMed/NCBI
32	Shimada Y, Kobayashi H, Kawagoe S, Aoki K, Kaneshiro E, Shimizu H, Eto Y, Ida H and Ohashi T: Endoplasmic reticulum stress induces autophagy through activation of p38 MAPK in fibroblasts from Pompe disease patients carrying c.546G>T mutation. Mol Genet Metab. 104:566–573. 2011. View Article : Google Scholar : PubMed/NCBI
33	Malhi H and Kaufman RJ: Endoplasmic reticulum stress in liver disease. J Hepatol. 54:795–809. 2011. View Article : Google Scholar
34	Bertolotti A, Zhang Y, Hendershot LM, Harding HP and Ron D: Dynamic interaction of BiP and ER stress transducers in the unfolded-protein response. Nat Cell Biol. 2:326–332. 2000. View Article : Google Scholar : PubMed/NCBI
35	Oikawa D, Kimata Y, Kohno K and Iwawaki T: Activation of mammalian IRE1alpha upon ER stress depends on dissociation of BiP rather than on direct interaction with unfolded proteins. Exp Cell Res. 315:2496–2504. 2009. View Article : Google Scholar : PubMed/NCBI
36	Szegezdi E, Logue SE, Gorman AM and Samali A: Mediators of endoplasmic reticulum stress-induced apoptosis. EMBO Rep. 7:880–885. 2006. View Article : Google Scholar : PubMed/NCBI
37	Hwang MS and Baek WK: Glucosamine induces autophagic cell death through the stimulation of ER stress in human glioma cancer cells. Biochem Biophys Res Commun. 399:111–116. 2010. View Article : Google Scholar : PubMed/NCBI
38	Deniaud A, Sharaf el dein O, Maillier E, Poncet D, Kroemer G, Lemaire C and Brenner C: Endoplasmic reticulum stress induces calcium-dependent permeability transition, mitochondrial outer membrane permeabilization and apoptosis. Oncogene. 27:285–299. 2008. View Article : Google Scholar
39	Lawless MW, Mankan AK, White M, O'Dwyer MJ and Norris S: Expression of hereditary hemochromatosis C282Y HFE protein in HEK293 cells activates specific endoplasmic reticulum stress responses. BMC Cell Biol. 8:302007. View Article : Google Scholar : PubMed/NCBI
40	Kondo Y, Kanzawa T, Sawaya R and Kondo S: The role of autophagy in cancer development and response to therapy. Nat Rev Cancer. 5:726–734. 2005. View Article : Google Scholar : PubMed/NCBI
41	Levine B and Kroemer G: Autophagy in the pathogenesis of disease. Cell. 132:27–42. 2008. View Article : Google Scholar : PubMed/NCBI
42	Criollo A, Maiuri MC, Tasdemir E, Vitale I, Fiebig AA, Andrews D, Molgó J, Díaz J, Lavandero S, Harper F, et al: Regulation of autophagy by the inositol trisphosphate receptor. Cell Death Differ. 14:1029–1039. 2007.PubMed/NCBI
43	Levine B and Klionsky DJ: Development by self-digestion: molecular mechanisms and biological functions of autophagy. Dev Cell. 6:463–477. 2004. View Article : Google Scholar : PubMed/NCBI
44	Hu G, Tuomilehto J, Pukkala E, Hakulinen T, Antikainen R, Vartiainen E and Jousilahti P: Joint effects of coffee consumption and serum gamma-glutamyltransferase on the risk of liver cancer. Hepatology. 48:129–136. 2008. View Article : Google Scholar : PubMed/NCBI