Construction of a recombinant lentiviral vector for VHL and VHL shRNA and its effect on proliferation and apoptosis of renal cell carcinoma cells.
- Author:
Donglai SHEN
1
;
Xin MA
;
Yu ZHANG
;
Yu GAO
;
Xingtao LI
;
Liangyou GU
;
Huijie GONG
;
Shaoxi NIU
;
Xu ZHANG
Author Information
- Publication Type:Journal Article
- MeSH: Apoptosis; Carcinoma, Renal Cell; pathology; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Genetic Vectors; Humans; Lentivirus; Plasmids; RNA, Messenger; RNA, Small Interfering; Transfection; Von Hippel-Lindau Tumor Suppressor Protein; genetics
- From: Journal of Southern Medical University 2015;35(3):348-354
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct a lentiviral expression vector for human VHL and its shRNA vector, and study the effect of VHL on proliferation and apoptosis of renal cell carcinoma cell lines.
METHODSLentiviral vectors pZsGreen1-VHL and pLL3.7-shVHL were constructed and transfected into 293T cells with 3 packaging plasmids by Lipofectamine(TM) 2000 reagent. The supernatant was collected to infect A498 and Caki-1 cells, respectively. VHL mRNA and protein levels were detected by RT-PCR and Western blotting, respectively. The effect of VHL on the proliferation, cell cycle and cell apoptosis were analyzed by MTS and flow cytometry.
RESULTSThe recombinant lentiviral vectors were successfully constructed. The proliferation of A498 cells with reconstituted wild-type VHL was significantly inhibited, while the proliferation of Caki-1 cells with VHL knockdown was significantly enhanced as compared with the control cells (P<0.05). VHL induced G0/G1-S cell cycle arrest. The apoptosis rate of A498 cells with reconstituted wild-type VHL was significantly increased while that of Caki-1 cells with VHL knockdown was significantly lowered compared with the control cells (P<0.05).
CONCLUSIONVHL can inhibit the proliferation and induce apoptosis of renal cell carcinoma cells.
