Establishment of a cell co-culture system in accordance with the immunological characteristics of chronic HBV infection
10.3760/cma.j.issn.1007-3418.2016.12.003
- VernacularTitle:符合慢性乙型肝炎病毒感染免疫特点的细胞共培养体系的建立
- Author:
Haijun LI
1
;
Xinying YANG
;
Fubiao KANG
;
Fengxin LIU
;
Dianxing SUN
Author Information
1. 解放军白求恩国际和平医院肝病传染科
- Keywords:
Hepatitis B,chronic;
Monocytes;
Immunology
- From:
Chinese Journal of Hepatology
2016;24(12):892-896
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate whether the co-culture of Huh7.93 cells and peripheral blood mononucleated cells from chronic hepatitis B patients (cPBMCs) can simulate the replication features of hepatitis B virus (HBV) and immune function in chronic hepatitis B (CHB) patients,and to provide an in vitro cell co culture system for the research on immune clearance in chronic HBV infection.Methods Huh7.93 cells were cultured alone or co-cultured with peripheral blood mononucleated cells from healthy people who underwent physical examination (nPBMCs) or cPBMCs for 7 days.The CCK8 assay was performed to measure the proliferative activity of Huh7.93 cells,and quantitative real-time PCR and Southern blot were used to measure HBV replication in cPBMCs and co-cultured cells.The independent samples t-test was used for comparison between two groups.Results When Huh7.93 cells were co-cultured with peripheral blood mononucleated cells (PBMCs) at a certain ratio,Huh7.93 cells had a high proliferative activity and good cell growth.HBV could not infect or replicate in cPBMCs.HBV DNA in the supematant of Huh7.93 cells co-cultured with cPBMCs showed significant increases and significantly higher than that in the supernatant of Huh7.93 cells cultured alone on day 4 (6.01 ± 0.20 log10copies/ml vs 4.99 ± 0.08 lOg10copies/ml,P =0.000) and day 7 (7.82 ± 0.24 log10copies/ml vs 6.96±0.09 log10copies/ml,P =0.000).On day 7 of culture,the cell lysis buffer of Huh7.93 cells co-cultured with cPBMCs had a significant increase in the level of HBV replicative intermediate compared with that of Huh7.93 cells cultured alone.After HepG2.2.15 cells were co-cultured with cPBMCs,there was no significant increase in HBV replication.Conclusion The co-culture of Huh7.93 cells and cPBMCs produces similar viral replication as human body infected with HBV and can well simulate the liver microenvironnent and immune function in CHB patients,which provides a new method for the research on immune clearance in chronic HBV infection.
