The porcine alpha1, 3 galactosyltransferase gene siRNA targeted heterozygous hepatocyte negative express GT.
- Author:
Quan-jian YAN
1
;
Yuan-fang ZHANG
;
Jie YANG
;
Qiang DING
;
Ji-gong WANG
;
Hao-wen JIANG
;
Hong ZHAO
;
Ke XU
;
Jian GONG
;
Liu-jin LI
;
Cheng-hai LIU
;
Jin-long GUO
;
En-jing LIU
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Cells, Cultured; Cloning, Molecular; Cytotoxicity, Immunologic; genetics; Galactosyltransferases; biosynthesis; genetics; Gene Silencing; Gene Targeting; methods; Hepatocytes; cytology; metabolism; Heterozygote; Immune Tolerance; genetics; Killer Cells, Natural; immunology; Mutation; RNA, Small Interfering; biosynthesis; genetics; Swine; Transfection
- From: Chinese Journal of Hepatology 2004;12(8):482-484
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo study whether the porcine alpha1, 3 galactosyltransferase gene siRNA targeted heterozygous hepatocyte negatively expresses GT mRNA and resists to the cytotoxicity of nature antibody in human serum.
METHODSThe porcine alpha1, 3 galactosyltransferase gene siRNA targeted vector (pPNTloxPGTsiRNA) were construct with pPNTloxPGT and pMXSV/U6 vector. Positive-negative selection was used to produce a heterozygous pPNTloxPGTsiRNA knockout (+/-) clone. The GT mRNA expressions were detected with northern blot. Complement-mediated NAb cytotoxicity after incubation of hepatocytes with NAbs and complement was determined using 3- (4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium(MTS, tetrazolium salt) colorimetric assay.
RESULTSThe pPNTloxPGTsiRNA targeted porcine hepatocyte (+/-) negative express GT mRNA. Only 14% to 18% cytotoxicity can be detected at the highest serum concentration. The pPNTloxPGT targeted porcine hepatocyte (+/-) express GT mRNA just as the wild type porcine cells and the cytotoxicity are 77% to 83%.
CONCLUSIONThe porcine a1, 3 galactosyltransferase gene siRNA targeted heterozygous hepatocyte (+/-) negative express GT and resisted to nature antibody in human serum.
