肝纤维化大鼠肝脏中组蛋白修饰水平的变化
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摘要
目的:观察肝纤维化大鼠肝脏中组蛋白修饰的变化,并探讨其在肝纤维化发生发展过程中可能的作用。方法:雄性Wistar大鼠20只,随机分为正常对照组和肝纤维化组,其中肝纤维化组采用CCl_4皮下注射以制备大鼠肝纤维化模型,正常组注射等量植物油溶液。实验第8周末,股动脉放血处死大鼠,取2组血清,采用生化和放射免疫法测定血清肝功能指标丙氨酸氨基转移酶(ALT)和天门冬氨酸氨基转移酶(AST),以及肝纤维化标志物血清透明质酸(HA)、层粘连蛋白(LN)、Ⅳ型胶原(Col Ⅳ)和Ⅲ型前胶原(PCⅢ)的水平;取2组大鼠肝脏,测定肝脏指数;取肝组织常规固定,HE染色和Masson染色观察组织病理改变及胶原纤维沉积情况;Western blot检测2组大鼠肝脏组织中α-平滑肌肌动蛋白(α-SMA)和I型胶原(ColⅠ)表达情况,以及acH4K12、acH3K9、H3K4me2和H3K9me2修饰水平的变化。结果:与对照组相比,模型组大鼠肝脏指数及ALT、AST、HA、LN、ColⅣ和PCⅢ水平明显增高(P<0.05);Western blot检测发现,与对照组比较,肝纤维化组大鼠肝组织的acH4K12修饰水平减少(P<0.05),acH3K9和H3K9me2修饰水平及α-SMA和ColⅠ表达明显增加(P<0.05),H3K4me2修饰水平的差异无统计学显著性。结论:肝纤维化大鼠肝脏中acH4K12、acH3K9和H3K9me2修饰水平改变可能与某些细胞外基质代谢相关基因转录调控有关,从而参与了大鼠肝纤维化发生。
AIM: To observe the changes of histone modifications in the liver of rats with hepatic fibrosis and its possible role in the development of hepatic fibrosis. METHODS: Male Wistar rats(n=20) were randomly divided into liver fibrosis group and normal control group. The liver fibrosis model was established by hypodermic injection of CCl_4, and the rats in normal control group were injected with vegetable oils. At the end of the 8 th week, all rats were killed. Liver function indexes including alanine aminotransferase(ALT) and aspartate aminotransferase(AST), and liver fibrosis indexes including haluronic acid(HA), laminin(LN), collagen type IV(Col Ⅳ) and procollagen type III(PCⅢ) were determined by biochemical and RIA methods. The liver index was analyzed, and the liver fibrosis degree and the morphological change of the liver were detected by HE and Masson staining. The levels of α-smooth muscle actin(α-SMA), collagen type Ⅰ(ColⅠ), H3 K4 me2, H3 K9 me2, acH3 K9 and acH4 K12 were detected by Western blot. RESULTS: After hypodermic injection of CCl_4 for 8 weeks, the liver index, ALT, AST, HA, LN, Col Ⅳ and PCⅢ of the rats in liver fibrosis group were higher than those in control group(P<0.05). Compared with control group, the level of acH4 K12 was decreased(P<0.05), while acH3 K9, H3 K9 me2, α-SMA and ColⅠ were increased(P<0.05), but H3 K4 me2 had no significant change.CONCLUSION: The levels of acH4 K12, acH3 K9 and H3 K9 me2 may play essential roles in the pathogenesis of liver fibrosis, and these histone modifications may regulate gene expression associated with extracellular matrix metabolism.
AIM: To observe the changes of histone modifications in the liver of rats with hepatic fibrosis and its possible role in the development of hepatic fibrosis. METHODS: Male Wistar rats(n=20) were randomly divided into liver fibrosis group and normal control group. The liver fibrosis model was established by hypodermic injection of CCl_4, and the rats in normal control group were injected with vegetable oils. At the end of the 8 th week, all rats were killed. Liver function indexes including alanine aminotransferase(ALT) and aspartate aminotransferase(AST), and liver fibrosis indexes including haluronic acid(HA), laminin(LN), collagen type IV(Col Ⅳ) and procollagen type III(PCⅢ) were determined by biochemical and RIA methods. The liver index was analyzed, and the liver fibrosis degree and the morphological change of the liver were detected by HE and Masson staining. The levels of α-smooth muscle actin(α-SMA), collagen type Ⅰ(ColⅠ), H3 K4 me2, H3 K9 me2, acH3 K9 and acH4 K12 were detected by Western blot. RESULTS: After hypodermic injection of CCl_4 for 8 weeks, the liver index, ALT, AST, HA, LN, Col Ⅳ and PCⅢ of the rats in liver fibrosis group were higher than those in control group(P<0.05). Compared with control group, the level of acH4 K12 was decreased(P<0.05), while acH3 K9, H3 K9 me2, α-SMA and ColⅠ were increased(P<0.05), but H3 K4 me2 had no significant change.CONCLUSION: The levels of acH4 K12, acH3 K9 and H3 K9 me2 may play essential roles in the pathogenesis of liver fibrosis, and these histone modifications may regulate gene expression associated with extracellular matrix metabolism.
引文
[1] Roeb E. Matrix metalloproteinases and liver fibrosis (translational aspects)[J]. Matrix Biol, 2018, 68-69:463-473.
[2] Lo RC, Kim H. Histopathological evaluation of liver fibrosis and cirrhosis regression[J]. Clin Mol Hepatol, 2017, 23(4):302-307.
[3] Naim A, Pan Q, Baig MS. Matrix metalloproteinases (MMPs) in liver diseases[J]. J Clin Exp Hepatol, 2017, 7(4):367-372.
[4] 田甜, 张金娟, 罗新华, 等. 大鼠原代肝星状细胞活化后组蛋白修饰水平观察[J]. 中国病理生理杂志, 2015, 31(5):871-876.
[5] 刘幸, 田甜, 余蕾, 等. 辛二酰苯胺异羟肟酸诱导大鼠原代肝星状细胞凋亡[J]. 中国病理生理杂志, 2017, 33(5):913-918.
[6] Jun HJ, Kim JY, Lee SJ, et al. Hepatic lipid accumulation alters global histone H3 lysine 9 and 4 trimethylation in the peroxisome proliferator-activated receptor alpha network[J]. PLoS One, 2012, 7(9):e44345.
[7] Zhao ZK, Yu HF, Wang DR. Overexpression of lysine specific demethylase 1 predicts worse prognosis in primary hepatocellular carcinoma patients[J].World J Gastroenterol, 2012, 18(45):6651-6656.
[8] Park PH, Miller R, Shukla SD. Acetylation of histone H3 at lysine 9 by ethanol in rat hepatocytes[J]. Biochem Biophys Res Commun, 2003, 306(2):501-504.
[9] Kim JS, Shukla AD. Histone H3 modifications in rat hepatic stellate cells by ethanol[J]. Alcohol Alcohol, 2005, 40(5):367-372.
[10] Sheen-Chen SM, Lin CR, Chen KH, et al. Epigenetic histone methylation regulates transforming growth factor β-1 expression following bile duct ligation in rats[J]. J Gastroenterol, 2014, 49(8):1285-1297.
[11] Wang Q, Wang LX, Zeng JP, et al. Histone demethylase retinoblastoma binding protein 2 regulates the expression of α-smooth muscle actin and vimentin in cirrhotic livers[J]. Braz J Med Biol Res, 2013, 46(9):739-745.
[2] Lo RC, Kim H. Histopathological evaluation of liver fibrosis and cirrhosis regression[J]. Clin Mol Hepatol, 2017, 23(4):302-307.
[3] Naim A, Pan Q, Baig MS. Matrix metalloproteinases (MMPs) in liver diseases[J]. J Clin Exp Hepatol, 2017, 7(4):367-372.
[4] 田甜, 张金娟, 罗新华, 等. 大鼠原代肝星状细胞活化后组蛋白修饰水平观察[J]. 中国病理生理杂志, 2015, 31(5):871-876.
[5] 刘幸, 田甜, 余蕾, 等. 辛二酰苯胺异羟肟酸诱导大鼠原代肝星状细胞凋亡[J]. 中国病理生理杂志, 2017, 33(5):913-918.
[6] Jun HJ, Kim JY, Lee SJ, et al. Hepatic lipid accumulation alters global histone H3 lysine 9 and 4 trimethylation in the peroxisome proliferator-activated receptor alpha network[J]. PLoS One, 2012, 7(9):e44345.
[7] Zhao ZK, Yu HF, Wang DR. Overexpression of lysine specific demethylase 1 predicts worse prognosis in primary hepatocellular carcinoma patients[J].World J Gastroenterol, 2012, 18(45):6651-6656.
[8] Park PH, Miller R, Shukla SD. Acetylation of histone H3 at lysine 9 by ethanol in rat hepatocytes[J]. Biochem Biophys Res Commun, 2003, 306(2):501-504.
[9] Kim JS, Shukla AD. Histone H3 modifications in rat hepatic stellate cells by ethanol[J]. Alcohol Alcohol, 2005, 40(5):367-372.
[10] Sheen-Chen SM, Lin CR, Chen KH, et al. Epigenetic histone methylation regulates transforming growth factor β-1 expression following bile duct ligation in rats[J]. J Gastroenterol, 2014, 49(8):1285-1297.
[11] Wang Q, Wang LX, Zeng JP, et al. Histone demethylase retinoblastoma binding protein 2 regulates the expression of α-smooth muscle actin and vimentin in cirrhotic livers[J]. Braz J Med Biol Res, 2013, 46(9):739-745.