| Objective: To investigate the mechanism of macrophage-derived exosome in HBV DNA replication and the correlation in patients with chronic hepatitis B.Methods:(1)In vitro,induce and identify mononuclear macrophages and exosome,and co-culture with HepG2.2.15 cells,and use real-time PCR to detect the HBV DNA changes in the supernatant;(2)Analysis of changes in the expression of functional miRNAs in exosome of macrophages;(3)Use of miRNA mimic and blockade assays to verify screening for inhibitory effects of functional miRNA on HBV DNA;(4)Combined with the clinical study cohort,patients with IA and IC period were enrolled,and healthy donors served as control groups,and to detect the expression of miRNAs in Serum-exosome,and to analysis the correlation with HBV DNA replication in patients with IA stage;(5)In vitro,to verify the effect of exogenous CD14-positive mononuclear cell exosome and serum-exosome on HBV DNA replication in HepG2.2.15 cells in patients with different immune status.Results:(1)Macrophage-derived exosome can effectively inhibit HBV replication;(2)The expression levels of miRNA-4284 and miRNA-1260 a in exosome of macrophages increased significantly;(3)miRNA-4284 and miRNA-1260 a could inhibit HBV replication;(4)The expression of miRNA-4284 and miRNA-1260 a in serum-derived exosome with IC stage was higher than IA stage;(5)The expression of miRNA-4284 and miRNA-1260 a in Serum-exosome was negatively correlated with HBV DNA in IA stage patients;(6)The serum and macrophage exosome in patients with IC stage inhibit HBV replication is stronger than IA stage.Conclusion:(1)In vitro,miRNA-4284 and miRNA-1260 a in macrophage-derived exosome inhibit HBV replication;(2)The expression of miRNA-4284 and miRNA-1260 a in serum-derived exosome of CHB patients with IC stage was higher than that in IA stage;(3)Serum-exosome and macrophage-derived exosome in IC stage patients should higher ability to inhibit HBV replication in HepG2.2.15 cells than IA stage patients. |
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