Osteopontin Regulates The Reaction Of Dendritic Cell On Hepatitis B Virus

Background and objectiveCurrently, the infection of hepatitis B virus (HBV) is still an important public health problem of influencing worldwide human health. However, so far the generated mechanism of HBV is still not clear.The generation and development of virus hepatitis are closely associated with the activation of immune cells and the production of cytokines.Dendritic cell (DC) plays a vital role in the recognition and presentation of HBV antigens, and it also influences the activations of Thl and Th2cells in the pathological and physiological process of HBV. Recent researches showed that DC induced not only immune response of hepatitis virus,but also immune tolerance, which was determined by the activation condition of DC.Moreover, some researches reported that DC function was impaired in HBV patients, but DC function was recovered to a certain extent after anti-virus treatment of HBV patients, along with functional restoration of T cell.Osteopontin(OPN) is a secretory phosphoric glycoprotein.As an immune regulatory factor, OPN plays an important part in the differentiation, maturity, and survival of DC.Our previous research proved that a variety of hepatic diseases are associated with high osteopontin (OPN) expression, and it also was closely related to the development and prognosis of hepatitis, hepatocellular carcinoma, and hepatic fibrosis.This research aims to explore the influence of OPN on the maturation and secretory function of DC during the process of HBV infection, thereby study the regulatory function of OPN on DC in this process.Material and methodsWe detected the expression of cluster of differentiation antigen (CD) molecular on DC surface in wild type mice and OPN knock-out mice after HBV antigen stimuli by flow-cytometer. We also measured the cytokines levels of culture supernatant of DC after HBV antigen stimuli by the method of ELISA. Moreover, the cytokines levels of co-culture supernatant of both DC and CD4+T cells after HBV antigen stimuli were detected by the method of ELISA.In addition, we collected the anticoagulant blood from the healthy people in outpatient service, separated the peripheral blood mononuclear cell (PBMC) with or without OPN antibody to culture, and then provided HBV antigen stimuli.And then we detected the expression levels of CD molecular on DC surface via flow-cytometer, and cytokines levels of culture supernatant by ELISA assay.ResultsThere is no significant difference in the expression of CD80,CD86and MHC-Ⅱ molecular in the DC without HBV antigens stimuli between wild type mice and OPN knock-out mice. However, the expression of CD80, CD86and MHC-Ⅱ molecular was remarkably decreased in the DC of OPN knock-out mice after HepG2.2.15supernatant stimuli and HBsAg stimuli, respectively. These data suggested that DC maturation was inhibited in OPN knock-out mice.On the contrary, the expression of CD80, CD86and MHC-II molecular was increased in the DC of OPN knock-out mice after HBcAg stimuli.Through the analysis of the IFN-a and IL-12secreted by DC,we found that without antigen stimuli,the secretory level of IFN-a was significant reduced in DC from OPN knock-out mice versus DC from wild type mice, but the IFN-a level in DC from OPN knock-out mice after antigen stimuli was still lower than those from wild type mice without antigen stimuli.The similar result was also found in the IL-12expression. Furthermore, we also found that even though the CD molecular expression was elevated in DC from OPN knock-out mice after HBcAg antigen stimuli,the secretory function of DC was decreased.Through the co-culture of DC from different sources and CD4+T cells from different sources, and the detection of Thl and Th2cytokines,we found that compared to the co-culture of both DC and CD4+T cells from wild type mice,the level of Thl cytokines including TNF-a was decreased, while the levels of Th2cytokines including IL-4,IL-6and IL-10was elevated in the co-culture of DC from OPN knock-out mice and CD4+T cells from wild type mice.Meanwhile, through the co-culture of DC from different sources and CD4+T cells from different sources,we found there were no significant difference in cytokines levels in co-culture supernatant of DC from same sources co-cultured with CD4+T from different sources, which further prove that OPN secreted by DC played a determined part in the process of T cells response induced by DC.Moreover,through human PBMC culture with OPN antibody to block OPN function, we found that the expression of CD80and CD86on DC surface was decreased, and the same trend was observed after HBV antigens stimuli.These data indicated that DC maturity was suppressed after blocking OPN function.We also found that the level of IL-12mainly secreted by DC was reduced. Meanwhile, the level of IFN-a was significantly decreased, while the levels of IL-4and IL-6were obviously elevated.ConclusionDuring the process of development and maturity, DC secreted massive OPN,and OPN performed a critical role in this process.The shortage of OPN could inhibit DC maturity via suppressing the expression of CD80and CD86on DC surface, and the level of cytokines secreted by DC after HBV antigen stimuli.The defect of DC in phenotype and function inhibited the activation of Th1induced by DC after HBV antigens stimuli, leading to the imbalance of Th1/Th2cytokines,thereby influencing DC function in the process of HBV infection.