Down-regulation of αGal epitopes by co-transfection of α1,3-galactosidase gene and α1,2-fucosyltransferase gene  

作　　者：GONG Feng ZHANG Yangpei JIA Yanjun WANG Yingli TAN Yingxia TIAN shuguang 

机构地区：[1]Beijing Institute of Transfusion Medicine, Beijing 100850

出　　处：《Chinese Science Bulletin》2005年第23期2723-2727,共5页

基　　金：This work was supported by grants from the National Basic Research Program of China(Grant No.9732002CB713804);Hi-Tech Research and Development Foundation of China(Grant No.8632003AA205101)

摘　　要：The polycarbohydrate structure of Galα1- 3Galβ1-4GluNAc-R (known as αGal epitopes of xenoantigen), produced by α1-3-galactosyltransferase (α1,3-GT) in the course of animal development, is the major xenoantigen on the cell surface of porcine which causes hyperacute rejection in pig-to-human xenotransplantation. Alpha-1,3-galactosi- dase (AGL), a hydrolytic enzyme, can remove the terminal α-1,3-galactosyl from the Galα1-3Galβ1-4GluNAc-R struc-ture resulting in cleaning αGal epitopes from the porcine cells. Alpha-1,2-fucosyltransferase (HT) can modify the sur-face carbohydrate phenotype of porcine cells, bringing about reduction of αGal epitopes expression. In this study, human AGL and HT gene were co-transfected to porcine fetal fibro-blast (PFFb) in equimolar concentration to reduce the xeno-antigen. Gene and protein of hAGL and HT were both de-tected to express at high level by RT-PCR and Western blot, respectively. There was an 84% reduction in αGal xenoanti-gen and an 82% increase in H antigen as assayed by flow cytometry in the AGL and HT gene co-transfected PFFb. The number and morphology of transgenic PFFb chromosome were normal. Findings indicate that Galα1-3Gal epitopes of PFFb could be down regulated by AGL and HT co-transfec- tion without deleterious effects on the chromosomal profile of the transgenic cell.The polycarbohydrate structure of Galα1- 3Ga1β1-4GluNAc-R （known as αGal epitopes of xenoantigen）, produced by α1-3-galactosyltransferase （α1,3-GT） in the course of animal development, is the major xenoantigen on the cell surface of porcine which causes hyperacute rejection in pig-to-human xenotransplantation. Alpha-1,3-galactosidase （AGL）, a hydrolytic enzyme, can remove the terminal α1,3-galactosyi from the Galα1-3Galβ1-4GluNAc-R structure resulting in cleaning αGai epitopes from the porcine cells. Aipha-1,2-fucosyitransferase （HT） can modify the surface carbohydrate phenotype of porcine cells, bringing about reduction of αGai epitopes expression. In this study, human AGL and HT gene were co-transfected to porcine fetal fibroblast （PFFb） in equimolar concentration to reduce the xenoantigen. Gene and protein of hAGL and HT were both detected to express at high level by RT-PCR and Western blot, respectively. There was an 84% reduction in αGai xenoantigen and an 82% increase in H antigen as assayed by flow cytometry in the AGL and HT gene co-transfected PFFb. The number and morphology of transgenic PFFb chromosome were normal. Findings indicate that Galα1-3Gal epitopes of PFFb could be down regulated by AGL and HT co-transfection without deleterious effects on the chromosomal profile of the transgenic ceil.

关 键 词：异种移植术 αGal抗原决定基 减量调节 聚多糖 α1-3-半乳糖苷酶 α1-2-墨角藻糖基乳糖 

分 类 号：R617[医药卫生—外科学]