Experimental study on expression of ATP7B gene to restore copper transport in skin fibroblasts Me32aT22/2L cells
10.3760/cma.j.issn.1671-8925.2009.02.002
- VernacularTitle:Me32aT22/2L细胞内肝豆状核变性基因的表达恢复铜转运功能的实验研究
- Author:
Zhen-Wen YAN
1
;
Jiang-Ying CHEN
;
Xiu-Ling LIANG
Author Information
1. 中山大学附属第二医院
- Keywords:
Hepatolenticular degeneration gene;
Gene expression;
Me32aT22/2L cell;
Copper transport
- From:
Chinese Journal of Neuromedicine
2009;8(2):114-117
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observed hepatolenticular degeneration gene (ATP7B gene) expressed in Me32aT22/2L cell and detected whether A TP7B could transport redundant copper and reduce copper-induced cell apoptosis, which will make the basement for future gene therapy. Methods Piasmid pRc/CMV-WD containing A TP7B cDNA was transfected into Me32aT22/2L cell using lipofectamin2000 transfection methods. Intracellular distribution of ATP7B was detected by immunofluorescence histochemistry method and copper transportation function was studied using high-concentration copper incubation experiments, meanwhile, cell apoptosis induced by high-concentration copper incubation was observed. Results Expression of A TB7B gene could be detected and located around nuclei within the Me32aT22/2L cell. After incubation of high- concentration copper solution after 24, 48 and 72 hours, in empty vector group intracellular copper content was (600.60±69.71) ng/mg, (890.72±65.74) ng/mg and (1189.20±85.71) ng/mg respectively, however, intracellular copper content was (351.33±49.86) ng/mg, (427.38±30.95) ng/mg and (539.10±34.91) ng/mg in A TP7B group. Comparison of two groups has a statistical significance (P<0.01). Meanwhile, cell apoptotic rate was markedly decreased in A TPTB group compared with empty vector group (P<0.01). Conclusion Exogenous A TP7B gene could be expressed in Me32aT22/2L cell and A TPTB could transport intracellular redundant copper, subsequently reduced copper-induced cell apoptosis.
