转化生长因子β1和结缔组织生长因子在小鼠血吸虫病肝纤维化中的表达

中国寄生虫学与寄生虫病杂志 ›› 2012, Vol. 30 ›› Issue (1): 4-20-26.

转化生长因子β1和结缔组织生长因子在小鼠血吸虫病肝纤维化中的表达

黄加权1，朱俊2，李兰1，焦云桃1，徐蕾1，陶然1，范翔雪1，马科1，郭威1，宁琴1 *

1 华中科技大学同济医学院附属同济医院感染科，武汉 430030；2 同济咸宁医院感染性疾病科，咸宁 437100

出版日期:2012-02-29 发布日期:2012-09-28

Expression of Transforming Growth Factor-β1 and Connective Tissue Growing Factor in Schistosomal Hepatic Fibrosis of Mice

HUANG Jia-quan1，ZHU Jun2，LI Lan1，JIAO Yun-tao1，XU Lei1，TAO Ran1，FAN Xiang-xue1，MA Ke1，GUO Wei1，NING Qin1*

1 Department of Infectious Diseases，Tongji Hospital，Tongji Medical College，Huazhong University of Science and Technology，Wuhan 430030，China；2 Department of Infectious Diseases，Tongji Xianning Hospital，Xianning 437000，China

Online:2012-02-29 Published:2012-09-28

摘要/Abstract

摘要： 目的建立血吸虫病肝纤维化小鼠模型，观察转化生长因子β1 （TGF-β1）和结缔组织生长因子（CTGF）在小鼠血吸虫肝纤维化中的表达状况。方法 30只BALB/c小鼠随机分为模型组和对照组(每组15只)，模型组小鼠以腹部贴片法经皮肤攻击感染日本血吸虫尾蚴（30±1）条，分别于感染后第4、6、8、10和12周两组各取3只小鼠摘眼球取血，测定血清中丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）水平；采血后处死小鼠取肝组织，常规切片，经HE染色后观察各组小鼠肝组织的病理学改变，Masson染色后观察肝组织胶原显微增生情况；免疫组织化学和荧光定量PCR方法检测小鼠肝组织中TGF-β1、CTGF的蛋白和mRNA表达水平。结果感染后6~12周，模型组的ALT和AST水平与同期对照组比较差异统计学意义（P＜0.05）。感染12周时，模型组的ALT和AST水平分别为（173.53±31.12）U/L和（301.00±34.87）U/L均高于对照组的[（42.00±3.53） U/L和（96.58±11.26） U/L]。肝组织切片经HE染色后发现，模型组的肝组织有虫卵肉芽肿沉积，汇管区的纤维化特别明显，呈干线性纤维化改变，切面上见门静脉分支周围纤维组织增生呈树枝状分布；Masson染色显示，模型组的肝组织胶原纤维增生面积于感染后8周明显增加，感染后12周为（23.83±1.68）%，与对照组［（1.23±0.14）%］的比较，差异有统计学意义（P<0.05）。感染后10和12周，模型组的TGF-β1 [（22.34±2.58）%和（25.82±3.01）%]和CTGF[（11.32±2.44）%和（14.51±2.05）%]的蛋白阳性区域面积与对照组[（2.56±0.87）%和（1.09±0.73）%]的比较，差异有统计学意义（P＜0.05）。感染后6、8、10和12周，模型组的TGF-β1和CTGF mRNA相对转录水平呈上升趋势，与同时段对照组的比较差异均有统计学意义（P＜0.05）；其中在感染后10周时，TGF-β1 mRNA相对转录水平最高，为0.0721±0.0187，对照组的则为0.0089±0.0037；在感染后12周时，CTGF mRNA相对转录水平最高，为0.1136±0.0365，对照组的则为0.0293±0.0184；CTGF mRNA相对转录水平与日本血吸虫感染时间有明显的相关性（r=0.927，P＜0.05）。结论 BALB/c小鼠感染日本血吸虫后，肝组织中TGF-β1和CTGF蛋白阳性表达类型和表达分布区域基本一致，CTGF mRNA的相对转录水平与日本血吸虫感染时间有明显相关性。

关键词: 转化生长因子-&beta, 1；结缔组织生长因子；肝纤维化；血吸虫病；BALB/c小鼠

Abstract: Objective To establish the model of hepatic fibrosis in mice infected with Schistosoma japonicum and observe the expression of transforming growth factor-β1 （TGF-β1） and connective tissue growing factor （CTGF） in mice model. Methods Thirty BALB/c mice were randomly assigned to control group and model group. Each mouse of model group was infected with (30±1) S. japonicum cercariae through the abdominal skin. Serum samples were collected at 4，6，8，10, and 12 weeks after infection, and were analyzed for the levels of ALT and AST. Pathological changes and proliferation of hepatic collagen fibers in liver tissue were observed after HE staining and Masson staining. Immunohis-tochemistry and real-time PCR were used to detect the protein and mRNA expression of CTGF and TGF-β1. Results 6-12 weeks after infection, there was significant difference in ALT and AST between model group and control group（P＜0.05）. At 12th week, ALT [（173.53±31.12）U/L ] and AST [（301.00±34.87）U/L] in model group were higher than those in control group [(42.00±3.53) and (96.58±11.26)U/L]. In model group, egg granulomas formed in the liver, and the formation of hepatic fibrosis was significant in portal areas, and there was tree-like hepatic fibrosis around the portal vein branch. 8 weeks after infection, hepatic fibrosis area in mice of model group increased considerably，and there was significant difference in percentage of positive area of collagen between 12th week [（23.83±1.68）%] and control group［（1.23±0.14）%］（P<0.05）. 10 and 12 weeks after infection, the percentage of positive area of TGF-β1 [（22.34±2.58）% and （25.82±3.01）%] and CTGF[（11.32±2.44）% and （14.51±2.05）%]was higher respectively than that of the control ［（2.56±0.87）%，and （1.09±0.73）% （P<0.05）. 6，8，10，and 12 weeks after infection，both TGF-β1 and CTGF mRNA increased gradually, higher than that in control group（P<0.05）. 10 weeks after infection, TGF-β1 mRNA relative transcription level was the highest ( 0.0721±0.0187), and it was 0.0089±0.0037 in control group. CTGF mRNA relative transcription level reached the highest value（0.1136±0.0365）in 12 weeks after infection, while it was 0.0293±0.0184 in control group. CTGF mRNA expression was positively correlated with the duration of infection （r=0.927，P＜0.05）. Conclusion The area and cell types of TGF-β1 positive expression is the same as that of CTGF in liver tissue of schistosome-infected mice (BALB/c). CTGF mRNA expression is significantly related to the duration of infection, but it is not the case for TGF-β1.

Key words: Transforming growing factor-&beta, 1；Connective tissue growing factor；Hepatic fibrosis； Schistosomasis；BALB/c mouse

黄加权1，朱俊2，李兰1，焦云桃1，徐蕾1，陶然1，范翔雪1，马科1，郭威1，宁琴1 *. 转化生长因子β1和结缔组织生长因子在小鼠血吸虫病肝纤维化中的表达[J]. 中国寄生虫学与寄生虫病杂志, 2012, 30(1): 4-20-26.

HUANG Jia-quan1，ZHU Jun2，LI Lan1，JIAO Yun-tao1，XU Lei1，TAO Ran1，FAN Xiang-xue1，MA Ke1，GUO Wei1，NING Qin1*. Expression of Transforming Growth Factor-β1 and Connective Tissue Growing Factor in Schistosomal Hepatic Fibrosis of Mice[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2012, 30(1): 4-20-26.