TGF-β1-Smads信号传导通路在感染日本血吸虫小鼠肝纤维化中的表达  

The expression of TGF-β1-Smads pathway in mice liver fibrosis induced by Schistosomajaponicum

作　　者：张彬彬[1] 蔡卫民[2] 陶君[3] 

机构地区：[1]黑龙江省医院消化病院消化一科,哈尔滨150001 [2]浙江大学医学院附属第一医院传染病研究所,杭州310003 [3]解放军杭州疗养院,杭州310003

出　　处：《国际医学寄生虫病杂志》2013年第3期121-129,共9页International JOurnal of Medical Parasitic Diseases

摘　　要：目的探讨在BALB／c小鼠感染日本血吸虫后形成肝纤维化的过程中，转化生长因子-β（transforming growth factorsβ，TGF—β）l及其两类受体：TGF—β受体I（TβRI）、TGF-β受体Ⅱ（TBRⅡ）以及Smad2、Smad3、Smad4和Smad7在转录水平的表达。方法50只BALB／c小鼠感染日本血吸虫尾蚴，（20-4-1）条／只，用于构建肝纤维化模型，10只未感染的BALB／c小鼠作为健康对照组，分别在感染后8、12、16和24周处死小鼠取肝组织．同时取对照组小鼠肝组织。所获肝组织一部分立即液氮冷冻后，保存于-80℃，通过RT—PCR方法测定TGF-β1、TBRI、TBRⅡ和Smad2、Smad3、Smad4以及Smad7的mRNA水平，结果为相对值，以待测mRNA密度扫描计数与内参照B—actinmRNA密度扫描计数的比值表示。另一部分肝组织置入常规10％甲醛固定液中．用于伊红-苏木素（HE）染色和天狼猩红染色．其中HE染色用于测定血吸虫虫卵肉芽肿面积（每一个虫卵肉芽肿的最大长度与最大宽度的乘积，以mm^2表示），每份标本随机测量5个虫卵肉芽肿面积．求平均值．天狼猩红染色用于判断肝纤维化程度．并以下述方法计分：正常肝组织为0级，以20=1计分；胶原纤维包绕肉芽肿周围并插入其中为I级，以21=2计分；汇管区有大量纤维化，小叶间仅有少量纤维为Ⅱ级，以2z=4计分：纤维组织大量延伸至小叶间为Ⅲ级。以23=8计分。结果小鼠感染日本血吸虫8周后其肝脏中形成的虫卵肉芽肿周围出现胶原纤维．并随着感染时间的延长，胶原纤维逐渐增加。感染后16周，胶原纤维在肝小叶中分布明显，其肝纤维化程度计分为4．27±1．03分；至感染24周时，胶原沉积量达到顶峰，为6．90±1．57分；而正常小鼠肝组织的肝纤维化计分为1分。正常小鼠肝脏中TGF—BltuRNA的表达水平为0．30±0．18，其表达量在感染后8周达到高峰（0．87±0．76），而�Objective To investigate the transcription level expression of TGF-β1, its two transmembrane receptors TGF-β receptor I （TβRI） and TGF-β receptor II （TβR II） and Smad2, Smad3, Smad4 and Smad7 during the development of liver fibrosis in the BALB/c mice infected with Schistosomajaponcum. Methods Fifty BALB/c mice infected with cercariae of Schistosomajaponicum （20 ± 1 ） were used as the liver fibrosis model and 10 uninfected mice belonged to the normal control group. Liver specimens were got at the 8th, 12th, 16th and 24th week post infection respectively and the normal controls were sacrificed at the same period mentioned above. Some liver tissues were frozen in the fluid nitrogen immediately and then conserved in the -80℃ refrigerator for reverse transcription polymerase chain reaction （RT-PCR） to detect the mRNA level of TGF-β1, TβRI , TβR II, Smad2, Smad3, Smad4 and Smad7. The final value was expressed by the ratio of the scan density of the amplified fragment with the internal control of β-actin, which is a relative value. Other liver pieces were fixed in 10% buffered formalin for histology assay to detect the size of egg granuloma measured by the product of maximum width and the maximum length and expressed in square micrometers and the final results were the average of the five readings randomly under the microscope and thus to determine the liver fibrosis degree, by the criteria： grade Ⅰ ： 2^0=1 no significant collagen was observed in liver tissue; grade Ⅱ： 2^1=2 collagen distributed around and in the granuloma; grade Ⅲ： 2^2=4 more collagen appeared in the portal tracts while few collagen a- mong liver lobules; grade Ⅳ： 2^3=8 fibrous tissues penetrated into liver lobules. Results Collagen fibers appeared around egg granulomas after 8 weeks of Schistosomajaponcum infection and increased gradually. At the 16th week after infection, fibrous tissue distributed evidently in liver lobules and the score of liver fibrosis was 4.27± 1.03 and fibrosis degree peaked at th

关 键 词：日本血吸虫 SMAD 纤维化 肝脏 TGF一81 受体 

分 类 号：R57[医药卫生—消化系统]