The comparative study on the regulation of apoptosis by hepatitis B virus X protein between B and C genotype
10.3760/cma.j.issn.1003-9279.2013.05.008
- VernacularTitle:B、C基因型乙型肝炎病毒X蛋白调控细胞凋亡的比较研究
- Author:
Yue MING
1
;
Qi-Feng XIE
;
Lin YANG
Author Information
1. 番禺中心医院
- Keywords:
Hepatitis B virus;
Genotype;
Apoptosis;
Oncogeme proteins,viral
- From:
Chinese Journal of Experimental and Clinical Virology
2013;27(5):344-347
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the apoptosis regulation on hepatoma cells by HBx between genotype B and C.Methods Genotype B and C HBx gene fragments were amplified and inserted into green fluorescent protein(GFP) eukaryotic expression vector pEGFP-C1 to construct recombinant pGFP-XB and pGFP-XC.The pEGFP-C1,pGFP-XB and pGFP-XC were introduced into Bel-7402 cells by Fugene HD to obtain Bel-7402 cells expressing GFP.The transcription and expression of HBx gene were demonstrated by RT-PCR and Western Blot analysis.Bel-7402,Bel-7402/GFP,Bel-7402/GFP-XB,Bel-7402/GFP-XC cells were treated with adriamycin(2.5 μg/ml),and the apoptosis of the cells was determined by trypan blue exclusion,and flow cytometry analysis.Results RT-PCR and Western Blot analysis showed that HBx genes of genotypes B and C were transcribed and expressed in Bel-7402/GFP-XB,Bel-7402/GFP-XC cells.Trypan blue exclusion showed adriamycin induced time-dependent cell death in Bel-7402,Bel-7402/GFP cells while no significant cell death was observed in Bel-7402/GFP-XB,Bel-7402/GFP-XC cells.Flow cytometry analysis indicated that no significant differences of apoptosis rates of Bel-7402/GFP-XB (3.87％) and of Bel7402/GFP-XC(4.01％) were observed (P ＞ 0.05),moreover,no significant differences of Bel-7402/GFP-XB(3.87％),Bel7402/GFP-XC (4.01％)and of the untreated cells.Apoptosis rates in Bel-7402/GFP-XB (3.87％),Bel-7402/GFP-XC (4.01％)cells were significantly lower than those in Bel-7402 (27.05％) and Bel-7402/GFP (29.14％) cells at 48 hours after the adriamycin treatment (P ＜ 0.01).Conclusions Bel-7402 cell lines expressing GFP,GFP-XB and GFP-XC fusion proteins were successfully established.HBV X protein blocks adriamycin-induced apoptosis of Bel-7402 cells.There is no difference between HBx of genotype B and C in inhibiting apoptosis induced by adriamycin.
