| Objective:Chronic hepatitis B(CHB)is one of the major causes of serious liver diseases,including liver cirrhosis,liver failure and hepatocellular carcinoma,and has a serious effect on the survival quality of CHB patients.The fully understanding of pathogenesis,and analysis of the related factors of CHB are significant to guide the clinical treatment.Previous studies have showed that the persistent HBV infection is associated with the functional exhaustion of T lymphocytes.The inhibitory microenvironment,including various inhibitory receptors,cells and cytokines,plays an important role in mediating the functional exhaustion of T cells.Myeloid-derived suppressor cells(MDSCs)are a heterogeneous population of immature myeloid cells,play immunosuppressive role through various mechanisms.However,the role of MDSCs in the development of CHB is still not clear.This study aimed to investigate the associations between the expression of MDSCs and the levels of other immune cells or clinical indicators,further to explore the clinical significance of MDSCs in the progression of CHB,by observing the frequency modulation of MDSCs,Treg,Th1 and Tc1 cells and detecting the levels of IL-10,TGF-? and IFN-? in the peripheral blood of patients with CHB.Finally,we hope our findings can provide new clues and experimental basis for clinical immunotherapy of CHB.Patients and Methods:A total of 39 patients with CHB were recruited of Shijiazhuang Fifth Hospital from December 2015 to December 2016.The diagnosis was based on clinical and laboratory findings according to the Guideline of Prevention and Treatment for chronic hepatitis B(2015 version).Twenty-six healthy volunteers,who came to the hospital for physical check-ups,were recruited asthe control group.The CHB patients were comparable to healthy controls with respect to sex and age.The basic information of all subjects was investigated,and the blood samples were collected.The frequencies of CD33+CD11b+HLA-DR-MDSCs in the peripheral blood were detected by flow cytometry.According to the expression of CD14 and CD15,CD14+CD15-MDSCs were defined as monocytic MDSCs(M-MDSCs),CD14-CD15+ MDSCs as granulocytic MDSCs(G-MDSCs),then the ratios of two subpopulations in MDSCs were evaluated.Peripheral blood mononuclear cells were isolated by ficoll density gradient centrifugation,the frequencies of Treg,Th1 and Tc1 cells were analyzed using flow cytometry.Treg,Th1 and Tc1 cells were defined as CD4+CD25+Foxp3+,CD4+IFN-?+ and CD8+IFN-?+cells,respectively.The serum levels of IL-10,TGF-? and IFN-? in CHB patients and healthy controls were detected by ELISA.Clinical and laboratory indicators,including liver function,HBV DNA load and HBeAg were measured.All statistical analyses were performed by SPSS 21.0 software.The expression of immune cells and the levels of cytokines were compared by Student's t test or rank sum test.Pearson correlation tests were performed for correlation analysis,including the associations between the expression of MDSCs or its subpopulations and the levels of other immune cells or clinical indicators.Multiple linear regression analysis was used to analyze the major related factors of MDSCs in treatment-na?ve CHB patients.A two-sided P value of ?0.05 considered to be significant.Results:1 The frequency of MDSCs in the peripheral blood of patients with CHB was significantly higher than that in healthy controls(1.19% vs 0.33%),and the difference was statistically significant(Z=-4.071,P<0.001).Compared with healthy controls,CHB patients had an increased percentage of G-MDSCs in MDSCs(Z=-2.015,P=0.044).However,no significant difference was observed with respect to the percentage of M-MDSCs in MDSCs between the two groups(Z=-1.614,P=0.107).2 The frequency of Treg in the peripheral blood of patients with CHBwas also significantly higher than that in healthy controls(3.28% vs 1.33%),and the difference was statistically significant(Z=-3.065,P=0.002).The ratios of Th1 and Tc1 cells in the CHB group were slightly higher than the healthy control group,however,the differences were not statistically significant(P>0.05).3 According to whether received antiviral treatment,CHB patients were divided into treatment-na?ve group and treated group.The frequencies of MDSCs and Treg in treatment-na?ve group or treated group were significantly higher than the healthy control group(MDSCs: treatment-na?ve group vs treated group vs healthy control group: 1.20% vs 1.20% vs 0.33%,H=16.70,P<0.001;Treg: treatment-na?ve group vs treated group vs healthy control group: 3.24% vs 3.28% vs 1.33%,H=9.985,P=0.007),however,the difference between treatment-na?ve group and treated group was not significant(P>0.05).The percentages of G-MDSCs,M-MDSCs,Th1 and Tc1 cells among the three groups had no statistically differences(P>0.05).4 In the treatment-na?ve CHB patients,the level of MDSCs was negatively correlated to Th1 cells(r=-0.645,P=0.013),but it was not associated with Treg and Tc1 cells(P>0.05).The percentages of G-MDSCs or M-MDSCs were not correlated with the levels of Treg,Th1 and Tc1 cells(P>0.05).5 The frequency of MDSCs was positively associated with the levels of ALT and HBV-DNA in serum of the treatment-na?ve CHB patients(r=0.436 and r=0.542,respectively),but the frequency of MDSCs was not correlated with the levels of AST,TBIL,DBIL(P>0.05).The percentages of G-MDSCs or M-MDSCs were not correlated with the levels of ALT,AST,TBIL,DBIL and HBV-DNA(P>0.05).In the treatment-na?ve CHB patients,the mean frequency of MDSCs in the HBeAg(+)group [(1.36 ± 0.79)%] was higher than that in the HBeAg(-)group[(0.82 ± 0.59)%],similarly,the percentages of G-MDSCs or M-MDSCs in the HBeAg(+)group were slightly higher than which in the HBeAg(-)group,but the differences were not statistically significant(P>0.05).6 Compared with the health,the level of IL-10 was significantly increased in CHB patients.However,the levels of TGF-? and IFN-?significantly decreased,and the differences were statistically significant(P<0.05).7 The levels of IL-10,TGF-? and IFN-? were not correlated with the percentages of MDSCs and its subpopulations(P>0.05).8 The amounts of IL-10 was positively associated with the levels of ALT,AST,TBIL,DBIL and HBV-DNA in serum of the CHB patients(P<0.05),the correlation coefficients were 0.487,0.757,0.487,0.486 and 0.664,respectively.However,the levels of TGF-? and IFN-? were not correlated with the clinical parameters(P>0.05).9 Multiple linear regression analysis showed that,the levels of HBV-DNA and Th1 cells were the major related factors of MDSCs in treatment-na?ve CHB patients.Conclusions:1 Compared with healthy controls,the frequency of MDSCs in the peripheral blood of patients with CHB was significantly increased,the population of G-MDSCs was also significantly increased,while no significant change was found about M-MDSCs.Similarly,the percentage of Treg was obviously increased,whereas no significant difference was observed with respect to the percentages of Th1 and Tc1 cells between the two groups,suggested that both MDSCs and Treg participate in the process of immune tolerance in the CHB patients,and the immune response of CHB patients is affected by the increased immune suppressive components due to HBV persistent infection.2 Compared with healthy controls,the level of IL-10 was significantly increased in CHB patients,and was positively associated with the clinical parameters,showed that MDSCs may prompt immune escape by producting IL-10.3 After accepted the antiviral treatment(Entecavir),the expression of MDSCs in the patients with CHB was no significantly changed,indicated thatalthough the antiviral treatment can effectively inhibit the viral replication and reduce the inflammation symptom,the expression of MDSCs have not been affected.4 The frequency of MDSCs in the peripheral blood of patients with CHB was negatively correlated to Th1 cells,however,MDSCs was not correlated to Treg or Tc1 cells.MDSCs may play immunosuppressive role by inhibiting the function of Th1 cells in the progression of chronic hepatitis B.5 The frequency of MDSCs were positively associated with the levels of ALT and HBV-DNA in serum of patients with CHB,implying that MDSCs take part in the progression of chronic hepatitis B.MDSCs may represent a potential target for developing effective immunotherapy for HBV infection. |
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