Dynamic Changes And Clinical Significance Of Treg Cells Frequencies And The Frequency Treg Cells Expressing TGF-β Of Patients With Hepatitis B Cirrhosis

Research background and purposeLiver cirrhosis is one of the common clinical outcomes of chronic HBV infection in China,which leads to high mortality and poor prognosis due to complications.The pathogenesis of hepatitis B cirrhosis is closely related to the role of immune cells in patients.Regulatory T(CD4+CD25+FOXP3+T)cells are a kind of immune cells that can induce immune tolerance and regulate immune overreaction.Many studies have reported the role of regulatory T cells in the progression of the diseases with abnormal immune response and chronic viral infection.Transforming growth factor-β(TGF-β)has been proved to activate hepatic stellate cells in an inflammatory environment and induce them to produce a large amount of extracellular matrix,which is closely related to the progression of liver cirrhosis.In addition,TGF-β can be secreted by Treg cells.It plays an important role in regulating epithelial to mesenchymal transition(EMT).EMT signaling is up-regulated to induce extracellular matrix deposition.We will observe the dynamic changes of Treg cells frequencies and the frequency Treg cells expressing TGF-β in patients with hepatitis B cirrhosis before and after treatment.Study members and methods1.Study membersA total of 105 HBsAg-positive with chronic HBV infection who were admitted to the Department of Infectious Disease of the First Affiliated Hospital of Zhengzhou University from September 2019 to September 2020 were enrolled.All patients informed consent got a biopsy.All tissue specimen sent to pathological examination.50 patients confirmed as hepatitis B cirrhosis were enrolled in the treatment group.The peripheral blood samples from 50 patients diagnosed as chronic HBsAg carriers and 50 healthy subjects were collected.Patients with hepatitis B cirrhosis and chronic HBsAg carriers were defined according to the criterions of diagnosis established by Hepatology Branch and Lemology Branch of Chinese Medicine Academy.Inclusion criteria:① HBsAg positive for more than 6 months,and liver biopsy pathology report confirmed liver fibrosis grade(S≥3);② HBV DNA was positive;③ No immunomodulators or antiviral drugs have been received in the last 6 months.Exclusion criteria:① Patients who did not addhere to the drug regimen for 72 weeks;②Patients with abnormal alpha-fetoprotein(AFP)or imaging evidence of primary liver cancer;③Patients with co-infection of HAV,HBV,HDV,HEV and HIV,combined with decompensated liver disease,alcoholic hepatitis,poorly controlled diabetes,autoimmune diseases or autoimmune mediated related diseases caused by other causes.2.MethodsAccording to the research strategy,the study subjects were divided into three groups:healthy control group,HBsAg carrier group and treatment group.(1)Compare the clinical data and serum markers of patients from the three groups.(2)Compare peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β among the three groups.(3)Patients with hepatitis B cirrhosis were divided into further two groups:normal ALT group and abnormal ALT group according to ALT>40U/L;Peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β between two groups were compared.(4)Hepatitis B cirrhosis patients received antiviral therapy;The clinical markers,peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β were compared at different time points(baseline,48 weeks and 72 weeks).(5)The analysis of Spearman rank correlation was used to analyze the correlation between Treg cells frequencies in peripheral blood and the frequency Treg cells expressing TGF-β and liver fibrosis staging.(6)Compare peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β between the different liver fibrosis extent.Results1.There was significant difference in the age、Lymphocyte、PLT、ALT、AST、GGT、ALB、TBIL and PT among the three groups(P<0.05).2.At baseline,the frequency of peripheral blood Treg cells in the treatment group increased significantly(t=6.934,P<0.05;t=3.905,P<0.05),and the expression of TGF-β increased significantly compared with the healthy control group and chronic HBsAg carriers.(t=10.270,P<0.05;t=6.880,P<0.05).3.In the treatment group,the frequency of Treg cells(t=2.220,P=0.031)and the frequency Treg cells expressing TGF-β of normal ALT group increased significantly compared with the abnormal ALT group(t=2.889,P=0.017).4.Compared with baseline,peripheral blood Treg cells frequencies decreased in the treatment group at 48 weeks(t=2.204,P<0.05),but there was no significant difference between weeks 48 and weeks 72(t=0.230,P=0.819).The frequency Treg cells expressing TGF-β also showed a decreasing trend at 48 and 72 weeks(48 weeks:t=2.089,P<0.05;72 weeks:t=4.446,P<0.05).5.The pathologic results of liver biopsy at 72 weeks,showed that the stage of liver inflammation and the rank of liver fibrosis in the treatment group were significantly improved compared with the treatment baseline.(inflammation:Z=-8.190,P<0.05;fibrosis:Z=-7.852,P<0.05).6.At baseline and 72 weeks,peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β showed the positive correlation with liver fibrosis(baseline:r=0.424,P<0.05,r=0.317,P=0.025;72 weeks:r=0.417,P<0.05,r=0.466,P<0.05).7.There were significant difference in the peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β between stage 0 and stage 1、stage 2 and stage 3、stage 3 and stage 4(P<0.05).ConclusionTreg cells frequencies in the peripheral blood and the frequency Treg cells expressing TGF-β in patients with hepatitis B cirrhosis decreased significantly after effective antiviral therapy.The dynamic changes of peripheral blood Treg cells frequencies and the frequency Treg cells expressing TGF-β could be used as biological markers to predict the extents of improvement of inflammation and cirrhosis in patients with hepatitis B cirrhosis.