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  • 邵佳亮,胡国信,郑洁,万赞燕,姜瑞娇.羟基喜树碱对肝纤维化大鼠肝组织BaxBcl-2基因和α-SMA蛋白表达及肝纤维化的影响[J].第二军医大学学报,2014,35(4):399-405    [点击复制]
  • SHAO Jia-liang,HU Guo-xin,ZHENG Jie,WAN Zan-yan,JIANG Rui-jiao.Effects of hydroxycamptothecin on hepatic expression of Bax and Bcl-2 genes and α-SMA protein and hepatic fibrosis in rats[J].Acad J Sec Mil Med Univ,2014,35(4):399-405   [点击复制]
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羟基喜树碱对肝纤维化大鼠肝组织BaxBcl-2基因和α-SMA蛋白表达及肝纤维化的影响
邵佳亮1,胡国信2*,郑洁3,万赞燕4,姜瑞娇5
0
(1. 江苏省高邮市人民医院消化科, 高邮 225600;
2. 北京大学深圳医院感染科, 深圳 518036;
3. 北京大学深圳医院中医科, 深圳 518036;
4. 江西省胸科医院感染科, 南昌 330006;
5. 河北省赵县人民医院病理科, 石家庄 051530
*通信作者)
摘要:
目的 考察羟基喜树碱(hydroxycamptothecin,HCPT)对肝纤维化大鼠肝组织中BaxBcl-2基因和α-平滑肌肌动蛋白(α-SMA)表达及肝纤维化的影响。方法 64只SD大鼠随机分为5组:正常组、模型组、低剂量 HCPT治疗组、中剂量HCPT治疗组、高剂量HCPT治疗组。采用40%四氯化碳(CCl4)诱导大鼠肝纤维化模型。正常组给予生理盐水腹腔注射;3个治疗组在造模同时分别给予0.25、0.5、1.0 mg/kg HCPT腹腔注射,3次/周,共8周。各组分别在第8周末取肝脏组织,行H-E、Masson染色观察肝脏组织病理学改变;RT-PCR检测肝脏组织中BaxBcl-2 mRNA表达并计算Bax/Bcl-2 mRNA比值;免疫组化染色检测肝脏组织中α-SMA蛋白表达;TUNEL染色观察细胞凋亡情况。结果 模型组大鼠出现明显的肝纤维化(Ⅲ期2例,Ⅳ期8例),各HCPT治疗组的肝纤维化程度较模型组减轻(低剂量组Ⅱ期1例,Ⅲ期8例,Ⅳ期1例;中剂量组Ⅱ期7例,Ⅲ期3例;高剂量组Ⅰ期1例,Ⅱ期7例,Ⅲ期2例),差异有统计学意义(P均<0.05)。RT-PCR检测显示模型组BaxBcl-2 mRNA较正常组升高,而各HCPT治疗组较模型组降低(P均<0.05);模型组Bax/Bcl-2 mRNA比值低于各HCPT治疗组(P均<0.05)。免疫组化染色检测显示模型组α-SMA蛋白表达水平高于中、高剂量HCPT组(P<0.05)。TUNEL染色结果显示正常组、模型组无明显阳性染色,各HCPT治疗组均有阳性染色。结论 HCPT对CCl4诱导的肝纤维化大鼠模型具有防治作用,抑制肝星状细胞活化增殖,上调Bax/Bcl-2 mRNA比值可能是HCPT抗肝纤维化的部分机制。
关键词:  肝纤维化  肝星状细胞  细胞凋亡  羟基喜树碱
DOI:10.3724/SP.J.1008.2014.00399
投稿时间:2013-11-17修订日期:2014-01-19
基金项目:国家自然科学基金(30760230).
Effects of hydroxycamptothecin on hepatic expression of Bax and Bcl-2 genes and α-SMA protein and hepatic fibrosis in rats
SHAO Jia-liang1,HU Guo-xin2*,ZHENG Jie3,WAN Zan-yan4,JIANG Rui-jiao5
(1. Department of Gastroenterology, People's Hospital of Gaoyou, Gaoyou 225600, Jiangsu, China;
2. Department of Infectious Diseases, Peking University Shenzhen Hospital, Shenzhen 518036, Guangdong, China;
3. Department of Traditional Chinese Medicine, Peking University Shenzhen Hospital, Shenzhen 518036, Guangdong, China;
4. Department of Infectious Diseases, Jiangxi Provincial Chest Hospital, Nanchang 330006, Jiangxi, China;
5. Department of Pathology, People's Hospital of Zhaoxian, Shijiazhuang 051530, Hebei, China
*Corresponding author.)
Abstract:
Objective To observe the effects of hydroxycamptothecin (HCPT) on hepatic expression of Bax, Bcl-2 genes, and α-smooth muscle actin (α-SMA) and hepatic fibrosis in rats. Methods Sixty-four SD rats were randomly divided into 5 groups: normal control group, model group, low-dose HCPT group, intermediate-dose HCPT group and high-dose HCPT group. Hepatic fibrosis was induced in rats by abdominal injection of CCl4. The normal control group was injected with normal saline only; the low-dose, intermediate-dose, and high-dose groups were injected with HCPT (3 times a week for 8 weeks) at 0.25, 0.5 and 1.0 mg/kg, respectively. At the end of the 8th week, liver tissues were obtained from each group for H-E staining and Masson staining to observe the degree of hepatic fibrosis. The expression of Bax, Bcl-2 mRNA and the Bax/Bcl-2 mRNA ratio in liver tissues were examined by RT-PCR. Immunohistochemical staining was used to observe α-SMA protein expression and TUNEL staining was used to observe cell apoptosis. Results Notable hepatic fibrosis was found in model group (Ⅲ stage in 2 rats,Ⅳ stage in 8 rats). Compared with the model group, each HCPT group had significantly improved fibrosis (P<0.05; low-dose HCPT group: Ⅱ stage in 1,Ⅲ stage in 8, Ⅳ stage in 1; intermediate-dose HCPT group:Ⅱ stage in 7, Ⅲ stage in 3; and high-dose HCPT group: Ⅱ stage in 1,Ⅲ stage in 7,Ⅳ stage in 2). RT-PCR results showed that the expression of Bax and Bcl-2 mRNA in the model group was significantly higher than that in the normal control group (P<0.05), and the expression in the three HCPT groups were significantly lower than that in the model group (P<0.05), with the ratio of Bax/Bcl-2 mRNA in the model group being significantly lower than those in the three HCPT groups (all P<0.05).The immunohistochemistry result showed that the hepatic α-SMA level in the model group was significantly higher than those in the intermediate-and high-dose HCPT groups (P<0.05). The TUNEL staining showed no significantly positive staining in the normal control group or model group, and positive staining in all the three HCPT groups. Conclusion HCPT has protective effect on CCl4-induced hepatic fibrosis in rats; inhibiting proliferation of hepatic stellate cells, up-regulating Bax/Bcl-2 mRNA ratio might be part of the related mechanism.
Key words:  liver fibrosis  hepatic stellate cells  apoptosis  hydroxycamptothecine