大鼠SV40LT抗原基因转染的肝细胞脾内移植后早期组织形态学和超微结构改变

背景:基础研究表明,肝细胞移植可以改善急性肝功能衰竭动物的生化参数及提高生存率.但其广泛应用于临床之前,还有细胞来源和免疫排斥、移植肝细胞在受体中的分布、形态结构变化等方面的问题需要解决.目的:观察大鼠脾内移植SV40LT抗原基因转染肝细胞的早期组织形态学和超微结构特点.设计、时间及地点:随机对照动物实验,细胞病理学观察,于2001-03/12在卫生部肝胆肠外科研究中心实验室完成.材料:选用Wistar大鼠60只,制作脾内肝细胞移植模型.方法:将60只大鼠按随机数字表法分4组,每组15只.原代组、原代 环孢素A组脾内注射原代肝细胞;SV40LT抗原基因组、SV40LT抗原基因 环孢素A组脾内注射SV40LT抗原基因转染的肝细胞.移植前24 h至术后14 d,原代组、SV40LT抗原基因组每天经尾静脉注入0.5 mL生理盐水;原代 环孢素A组、SV40LT抗原基因 环孢素A组每天经尾静脉注入环孢素A10mg/(kg·d).主要观察指标:术后每天每组取1只大鼠,取脾脏行光镜及电镜检查,观察移植肝细胞的存活率、组织形态学和超微结构特点,共观察14d.结果:1]与原代组、SV40LT抗原基因组比较,原代 环孢素A组和SV40LT抗原基因 环孢素A组移植肝细胞的组织形态学及超微结构改变较小,移植肝细胞的存活数增高(P<0.05).2]SV40LT抗原基因 环孢素A组、原代 环孢素A组移植肝细胞的存活率在术后1～7 d差异无显著性意义(P>0.05):术后8～14d,SV40LT抗原基因 环孢素A组肝细胞存活率高于原代 环孢素A组(P<0.01).结论:SV40LT抗原基因转染的肝细胞脾内移植后,在使用免疫抑制剂的情况下能长时间保持较高的存活细胞数,维持完整的组织形态学结构.

Early-stage histomorphology and ultrastructure of SV40LT antigen gene transfected hepatocytes transplanted into rat spleen

BACKGROUND:Hepatocyte transplantation can improve biochemical parameters and survival of animals with acute liver failure.However,cell sources.immunological rejection,distribution and histomorphological alternation of transplanted hepatocytes are important issues for its wide application in clinic.OBJECTIVE:To observe early-stage histomorphological and ultrastructural changes of SV40LT antigen gene transfected hepatocytes after transplanted into rat spleen.DESIGN,TIME AND SETTING:Randomized controlled animal trial was performed at Laboratory of National Hepatobiliary and Enteric Surgery Research Center.Ministry of Health from March to December 2001.MATERIALS:Sixty male Wistar rats were selected for hepatocyte transplantation.METHODS:Sixty rats were randomly divided into 4 groups(n=15).Primary cell group and primary cell plus cyclosporine A group were intrasplenically transplanted with primarily cultured hepatocytes;SV40LT antigen gene group and SV40LT antigen gene plus cyclosporine A group were intrasplenic transplanted with SV40LT antigen gene transfected hepatocytes.Twenty-four hours before and 14 days after transplantation,the rats in primary cell group and SV40LT antigen gene group were injected with 0.5 mL normal saline through tail vein daily,while the other groups were injected with cyclosporine A (10mg/kg per day).MAIN OUTCOME MEASURES:The spleen of one rat was harvested every day postoperatively for light microscopic and electron microscopic examinations to observe survival rate,histomorphological and ultrastructural features of transplanted hepatocytes for 14 days.RESULTS:Compared with primary cell group and SV40LT antigen gene group,the survival rate of transplanted hepatocytes in the other groups was significantly increased(P<0.05),but the histomorphological and ultrastructural changes were minor.There were no significant differences in the survival rate between SV40LT zntigen gene transfected hepatocytes and primarily cultured hepatocytes during the first week after transplantation(P>0.05),while the survival rate in SV40LT antigen gene transfected hepatocytes was significantly higher than primarily cultured hepatocytes 8-14 days postoperatively(P<0.01).CONCLUSION:Under treatment of cyclosporine A,the SV40LT antigen gene transfected hepatocytes can maintain a stable ultrastructure and a long survival rate.