HBVx Gene Influnces In Hepatic Progenitor Cells And Animal Models

Objective construction expression HBVx gene lentivirus vector and make it stability in 14-19 cells,establish a mouse model by portal vein injection of the cells.In the mice models,detected each group HBVx gene expression in different time.To observe their effects on the liver in the model,and a series of biological changes.Methods(1)HBVx gene was amplified by PCR And connect with the lentivirus expression vector(p Lenti-CMV-HA-3FLAG-PGK-EGFP-F2A-Puro),then using by restriction enzyme digestion,colony PCR and sequencing to verify whether the plasmid was successfully constructed.Then build a successful cell HBVx target gene detection after carrying the HBVx infection lentivirus 14-19 hepatic progenitor cells,and puromycin continuous selection for 4 weeks.(2)To construct anmial models.At the same time,respectively at 3,7,14,30,60 days to detect the expression of various tissues and organs of mice HBVx to use PCR,and use Elisa analysis the expression level of AFP and ALT in serum in mice.Results(1)Successfully constructed lenti-virus carrying HBVx expression plasmid;and constructed stable expression HBVx in 14-19 hepatic progenitor cells,and use puro(puromycin)for four consecutive screening.(2)successful construct animal models.And in 3,7,14,30,60 days simultaneous detection of the expression of various tissues and organs of HBVx by PCR methods,we found HBVx only express in the liver,is not expressed in the other organs.And through use Elisa method,respectively text 3,7,14,30,60 days in mice serum the expression level of AFP and ALT,compared with control group,mice serum AFP,and ALT levels were elevated.ConclusionHepatic progenitor cells carrying HBVx can be stably expressed in vivo.By detecting the expression of HBVx in tissues,HBVx has hepatophilic and does not involve other organs.The animal model of long-term expression of HBVx was constructed,which laid a good foundation for the study of the mechanism of HBVX in the development and progression of hepatocellular carcinoma.