Study On The Relationship Between Regulatory T And Nk Cells In Chronic Liver Diseases

Objective:During the progression of chronic liver disease,there is a phenomenon of NK cell depletion,but the specific mechanism is unclear.This study aims to explore(1)the relationship between CD4~+regulatory T cells and NK cell depletion in patients with chronic hepatitis B(CHB).(2)CD8~+regulatory T cells and CD4~+regulatory T cells and their relationship with NK cells in patients with hepatocellular carcinoma(HCC).Methods:(1)Collect the peripheral blood of CHB patients and healthy controls from clinical specimens,and analyze their biochemical indicators and ALT,AST,HBV DNA,HBs Ag,HBe Ag;(2)Peripheral blood of CHB patients,HCC patients or healthy people Blood mononuclear cells(PBMCs)for phenotype detection of NK cells and Treg cells;(3)PBMCs are separated and tested in vitro using flow cytometry to detect functional changes of NK cells in patients with CHB;(3)PBMCs will be isolated After culturing in vitro using CHB patient or healthy human serum,flow cytometry was used to detect changes in cell phenotype and function;(4)NK cells and Treg cells were separated and purified using magnetic beads,and co-culture was performed in vitro to study Treg cells and Possible mechanism of NK cell interaction;(5)CBA cytokine analysis kit was used to detect cytokine content in serum or culture supernatant.Results:In the study of CHB patients,it was found that(1)the expression ratio of NK cell inhibitory molecule NKG2A in peripheral blood of patients was increased,and was positively correlated with HBV-DNA load;(2)NK cell function was significantly reduced,and blocking NKG2A significantly restored NK cells(3)The expression ratio of CD4~+Treg cells and the secretion of IL-10 in CHB patients were significantly increased,and the peripheral blood NKG2A~+NK cells in CHB patients were significantly positively correlated with IL-10+Treg cells;(4)CHB patients were found in in vitro experiments Serum and HBe Ag can induce NKG2A~+NK cell expression through IL-10,and blocking IL-10 can restore NK cell exhaustion.Studies in HCC patients found that(1)the proportion of CD8~+regulatory T cells and CD4~+regulatory T cells in HCC patients is higher than that in healthy people,and CTLA-4 expression in cancer tissues is significantly higher than in adjacent tissues;The proportion of NK cells in peripheral blood and cancer tissues was significantly lower than that in healthy people and adjacent tissues;(3)CD8~+regulatory T cells and CD4+regulatory T cells were negatively correlated with NK cell expression.Conclusions:The depletion of NK cells and the increase of CD4~+Treg cells exist in CHB patients.In vitro experiments found that HBe Ag can increase the expression of NKG2A and decrease the function of NK cells by inducing IL-10 secretion from CD4~+Treg cells.In patients with HCC,regulatory T cells may also be involved in negatively regulating NK cells.