Eukaryotic expression of human Arresten gene and its effect on the proliferation and migration of vascular smooth muscle cells
- VernacularTitle:人Arresten基因的真核表达及其对血管平滑肌细胞增殖和迁移的影响
- Author:
Dan SHANG
;
Qichang ZHENG
;
Zifang SONG
;
Xiedan WANG
;
Qinggang HU
;
Xingjun GUO
- Publication Type:Journal Article
- Keywords:
Eukaryotic expression;
Arresten;
Vascular smooth muscle cells;
Cell proliferation;
Cell movement
- From:
Chinese Journal of Pathophysiology
2000;0(10):-
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To express human Arresten gene in eukaryotic cell,and to investigate its effect on the proliferation and migration in vitro of rat primary cultured thoracic aortic vascular smooth cells (VSMCs).METHODS: COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome.48 hours after transfection,polymerase chain reaction(RT-PCR) was used to detect the expression of Arresten mRNA in the cells,while Western blotting assay was applied to detect expressed Arresten protein in concentrated supernatants.VSMCs were then co-cultured with the concentrated supernatants;and its proliferation was detected using cell counting kit-8(CCK-8) in vitro.Migration of VSMCs was assayed by a microchemotaxis chamber and a polycarbonate filter (Transwell's chamber) with pores of 8 ?m in diameter.RESULTS: RT-PCR revealed that the genome of Arresten-transferred cells contained a 449bp specific fragment of Arresten gene.Successful protein expression in supernatants was confirmed by Western blotting.CCK-8 assay showed that the proliferation of VSMCs was inhibited significantly by Arresten protein as compared with control group(P
