低剂量三氯乙烯诱导人正常肝细胞损伤耐受差异表达基因的研究

目的寻找低剂量三氯乙烯(TCE)诱导人正常肝细胞(L02)损伤耐受差异表达的基因。方法参照本实验室用MTT法所得的TCE对L02毒性剂量-效应关系,选择5μmolL的三氯乙烯为低剂量,40μmolL为高剂量,按以下方式对细胞进行染毒:空白对照、低剂量、高剂量、损伤耐受组(先用低剂量刺激24小时,再用高剂量刺激6小时),然后用荧光差异显示PCR(FluoroDDPCR)寻找不同方式刺激差异表达的基因,并对差异表达基因进行克隆、鉴定。结果按方法所示的方式处理细胞,处理完毕后提取各组细胞总RNA,进行荧光差异显示PCR,找到51个差异条带。对其中的11个差异条带进行克隆鉴定同源性比较,发现其中9个已知基因,2个新基因。结论用荧光差异显示PCR方法寻找TCE诱导L02损伤耐受过程差异表达的基因,通过鉴定基因,为进一步研究TCE诱导L02的损伤耐受的机理提供科学依据。

Research on differential display genes of tolerant-damage induced by trichloroethylene

OBJECTIVE: To observe the differentially expressed genes of the human normal liver cell (L-02) induced by low concentration of trichloroethylene (TCE). METHODS: The dose-effect relation of TCE toxicity was analyzed by means of MTT. 5 micromol/L was chosen as low concentration while 40 micromol/L as high concentration. Then L-02 were treated with low concentration, high concentration, pretreated with low concentration then attacked with high concentration of TCE respectively. Fluorescence differential display polymerase chain reaction (Fluoro DD-PCR) was used to search differentially expressed genes of the different treatment groups of TCE. Results 51 differential expressed strap were found, 11 differential straps have been cloned and sequenced. 9 of them were known gene, while 2 of them were new genes. CONCLUSION: The differential straps would be identified to provide scientific base for research deeply the mechanism of low concentration TCE inducing adaptive response of L-02.