Roumudan formulation attenuated liver fibrosis by inhibiting TGF-β1/Smad4 pathway in mice

doi:10.12047/j.cjap.6010.2020.121

Abstract

Abstract: Objective: To investigate underlying mechanism involving Roumudan(RMD) formulation (Z20160012) suppressed liver fibrosis induced by CCl₄ injection in mice by inhibiting TGF-β1/Smad4 pathway. Methods: Male BALB/c mice were randomly divided into control group, liver fibrosis model group and RMD-treated group(n=11). Mice in liver fibrosis model and RMD-treaded groups were injected intraperitoneally with CCl₄ (20% in olive oil) at the dose of 2.5 mL/kg two times for one week and 5 mL/kg two times for 4 weeks. Mice in control group were treated intraperitoneally with the same volume of olive oil at the same time intervals. From sixth week, Mice in liver fibrosis model group were administrated with CCl₄ (20% in olive oil, 1.5 ml/kg once per week) intraperitoneally and given distilled water by intragastric gavage. Mice in the RMD-treated group were administrated with CCl₄ (20% in olive oil, 1.5 ml/kg/mouse once per week) intraperitoneally and given RMD(6.2 g/kg everyday) by intragastric gavage. Mice in the control group were administrated with olive oil (1.5 ml/kg/mouse once per week) intraperitoneally and given distilled water by intragastric gavage. The serum AST and ALT levels were estimated for assessment of liver function. The pathologic changes of mice' livers were examined by the HE, Masson, immunohistochemical staining, Western Blot, Q-PCR and so on. Results: After intraperitoneally injected with CCl₄ in mice, the pathological characteristics of liver fibrosis were observed compared with the control group at the sixth week. Compared with the liver fibrosis model group, RMD improved the liver function significantly through reducing liver index(P＜0.01) and the levels of ALT(P＜0.01), AST(P＜0.01) and HYP(P＜0.05). The expression of TGF-β1(P＜0.05), α-SMA(P＜0.05), COL1A1(P＜0.01) and COL3A1(P＜0.01) were decreased by RMD. The positive expression area of Smad4 mRNA in RMD treated group was lower than that in liver fibrosis model group. Conclusion: The RMD formulation could attenuate liver fibrosis by inhibiting TGF-β1/Smad4 pathway and extracellular matrix (ECM) production in mice.

Key words: fibrosis, liver, roumudan, carbon tetrachloride, mice

CLC Number:

R256.42

LI Qian, YAN Shu-guang, HUI Yi, LI Jing-tao, WEI Hai-liang, ZHANG Hong. Roumudan formulation attenuated liver fibrosis by inhibiting TGF-β1/Smad4 pathway in mice[J]. CJAP, 2020, 36(6): 576-581.

References

[1] Duran A, Hernandez ED, Reina-Campos M, et al. p62/SQSTM1 by binding to vitamin D receptor inhibits hepatic stellate cell activity, fibrosis, and liver cancer[J]. Cancer Cell, 2016, 30(4): 595-609.
[2] Higashi T, Friedman SL, Hoshida Y. Hepatic stellate cells as key target in liver fibrosis[J]. Advanced drug delivery reviews, 2017, 121: 27-42.
[3] Affo S, Yu LX, Schwabe RF. The role of cancer-associated fibroblasts and fibrosis in liver cancer[J]. Annu Rev Pathol, 2017, 12: 153-186.
[4] 王志旺, 付晓艳, 程小丽, 等. 育阴软肝颗粒剂对肝纤维化大鼠TGF-β1表达的影响[J].中国应用生理学杂志, 2018, 34(2): 169-172.
[5] Zhang X, Xu Y, Chen JM, et al. Huang Qi decoction prevents BDL-induced liver fibrosis through inhibition of notch signaling activation[J]. Am J Chin Med, 2017, 45(1): 85-104.
[6] 席奇, 刘宝咸, 刘亚珠, 等. 柔木丹颗粒治疗乙型肝炎肝纤维化随机对照研究[J]. 现代中西医结合杂志, 2016, 25(36): 3991-3993.
[7] 席奇, 刘亚珠, 宋春荣, 等. 柔木丹颗粒联合恩替卡韦治疗乙型肝炎肝纤维化45例疗效观察[J]. 中医药导报, 2015, 21(24): 46-48.
[8] 史晓燕, 许小凡, 宋亮, 等. 柔木丹颗粒对四氯化碳诱导小鼠肝纤维化的治疗作用[J]. 新乡医学院学报, 2019, 36(1): 1-6.
[9] 汪小玲, 杨红侠, 刘芸, 等. 柔木丹颗粒质量标准的研究[J]. 陕西中医学院学报, 2009, 32(1): 62-63.
[10] 黄继汉, 黄晓晖, 陈志扬, 等. 药理试验中动物间和动物与人体间的等效剂量换算[J]. 中国临床药理学与治疗学, 2004, 9(9): 1069-1072.
[11] 邓莉, 申宝德, 刘园, 等. 复方鳖甲软肝片减方对CCl₄诱导大鼠肝纤维化的影响及机制研究[J]. 中草药, 2018, 49(6): 1371-1378.
[12] 庄子锐, 王明亮, 张婷, 等. 肝肾纤维化动物模型的研究进展及评价[J/OL]. 中国实验动物学报, 2020: 1-8(2020-11-26).
[13] 肖敏, 屈小虎, 吕聚坪, 等. 吡非尼酮对四氯化碳诱导的小鼠肝纤维化的影响[J]. 中国应用生理学杂志, 2016, 32(4): 378-382.
[14] Huang Y, Deng X, Liang J. Modulation of hepatic stellate cells and reversibility of hepatic fibrosis[J]. Exp Cell Res, 2017, 352(2): 420-426.
[15] Watanabe T, Niioka M, Hozawa S, et al. Gene expression of interstitial collagenase in both progressive and recovery phase of rat liver fibrosis induced by carbon tetrachloride[J]. J Hepatol, 2000, 33(2): 224-235.
[16] Xu F, Liu C, Zhou D, et al. TGF-beta/SMAD pathway and its regulation in hepatic fibrosis[J]. J Histochem Cytochem, 2016, 64(3): 157-167.
[17] Yoshida K, Murata M, Yamaguchi T, et al. TGF-β/Smad signaling during hepatic fibro-carcinogenesis[J]. Int J Oncol, 2014, 45(4): 1363-1371.
[18] Zou Y, Cai Y, Lu D, et al. MicroRNA-146a-5p attenuates liver fibrosis by suppressing profibrogenic effects of TGF-β1 and lipopolysaccharide[J]. Cell Signal, 2017, 39: 1-8.
[19] Wu X, Wu X, Ma Y, et al. CUG-binding protein 1 regulates HSC activation and liver fibrogenesis[J]. Nature Communications, 2016, 7(13498): 1-14.
[20] Bruschi FV, Claudel T, Tardelli M, et al. The PNPLA3 I148M variant modulates the fibrogenic phenotype of human hepatic stellate cells[J]. Hepatology, 2017, 65(6): 1875-1890.