The Role Of RORγt And Foxp3in The Pathogenesis Of HBV-related Acute-on-chronic Liver Failure

Objective: Hepatitis B Virus(HBV)-related acute-on-chronic liver failure(ACLF)is an emergent, life-threatening syndrome that is characterized bysevere jaundice, hepatic encephalopathy and other variant complicationsowing to excessive liver necrosis and apoptosis due to HBV infection. Themechanisms of HBV related ACLF are extremely complex. Interactionbetween virus and the host is considered to be important factors. And virusreplication serves as requirement and primary cause leading to HBV relatedACLF. The damage of host immune responses may play an important role inits pathogenesis．Effector CD4+T cell plays an important part in immunesystems and immunoregulation. Previously some studies have suggested thatthe imbalance of Th1cells and Th2cells has play a important role in HBVrelated ACLF; Recently it is confirmed that Th17cells also take part inHBV-ACLF pathogenesis as a newcomer in CD4+cell family. In this study, wedynamically observed changes of the expressions of FOXP3, Th17cell, IL-21in peripheral blood and the balance and change rule of Treg/Th17in order toinvestigate the correlation between the pathogenesis and prognosis ofHBV-related ACLF and the balance of Treg/Th17and IL-21.Methods: Subjects in the experimental group are patients in the ThirdHospital of Hebei Medical University and the fifth Hospital of Shijiazhuangcity.45cases of HBV-related ACLF as ACLF group,50cases with chronichepatitis B (CHB) as CHB group,20cases with decompensated liver cirrhosis,10cases with compensated stage liver cirrhosis and14healthy cases asnormal control group. Liver and renal function parameters, coagulationfunction parameters, serum virology and HBV DNA loads were determined.The expression level of FOXp3, RORγt and IL-21mRNA in PBMCs weredetected by Real-time quantitative polymerase chain reactions(RT-PCR). The serum IL-23concentrations were determined by enzyme-linked immunosorb-ent assays(ELISA). In addition, we dynamically observed the changes ofFOXp3、RORγt and IL-21levels in HBV-related ACLF patients.Results:1The demography and baseline condition in clinicalThe serum alanine transaminase (ALT) levels, TBIL and INR in ACLFgroup were significant higher than those in other groups（P<0.05）.Withrespective to the occurrence of severe complications in total ACLF patients,there were9having pulmonary fungal infection,12having spontaneousbacteria peritonitis,4having hepatorenal syndromes, and7having hepaticencephalopathy.2The expression level of FOXP3mRNA in PBMCs:The relative expression level of FOXP3mRNA in PBMCs in ACLFgroup was observably higher than other five groups(P<0.05), as follows:ACLF group(0.27010-20.01810-2), HC group(0.04110-20.00410-2),CHB-m group(0.12010-20.01810-2), CHB-s group(0.18210-20.02510-2), LC-c group(0.13810-20.01810-2), LC-d group(0.17810-20.02710-2).Except for LC-d and CHB-s group(P=0.569), there wassignificant difference between any two groups (P<0.05).3The expression level of RORγt mRNA in PBMCsThe expression level of RORγt mRNA in PBMCs in ACLF group wasthe highest among all groups (P<0.05), as follows: ACLF group(0.29910-20.03310-2), HC group(0.03110-20.00510-2), CHB-m group(0.07910-20.02010-2), CHB-s group(0.13910-20.02310-2), LC-c group(0.09310-20.01110-2), LC-d group(0.14610-20.01310-2).There wassignificant difference between any two groups except LC-d and CHB-sgroups(P=0.412).4The expression level of RORγt/Foxp3mRNA was on the rise along with thesevere state of illness with HBV-related liver disease. Compared with othergroups, it was the highest in ACLF group (1.1050.055). In liver cirrhosis groups, the expression level of RORγt/Foxp3mRNA in LC-d group wassignificantly higher than LC-c group (P<0.05).5The expression level of IL-21in PBMCsThe expression level of IL-21in PBMCs was gradually increasing in HC,CHB-m, CHB-s and ACLF group. And it achieved the highest level in ACLFgroup(0.32610-20.06010-2). There is significant difference between anytwo groups.6Change of serum IL-23levelsThe levels of serum IL-23in ACLF group (38.932.4pg/ml)weresignificantly higher than that in CHB-m group (16.353.24pg/ml), CHB-sgroup (23.433.5pg/ml) and HC group (12.321.81pg/ml)(P<0.05).7Linear correlation of RORγt and Foxp3expression with disease severitymaikers in ACLF patientsIn ACLF patients, RORγt was positively correlated with PTA (r=-0.373，P=0.012), TBil (r=0.303, P=0.043), HBV DNA load (r=0.297, P=0.047) andMELD score (r=0.328, P=0.023). And there were no correlated betweenRORγt and ALT (r=-0.279, P=0.064). We also find that Foxp3was positivelycorrelated with PTA (r=-0.474, P=0.001), TBil (r=0.357, P=0.023), HBVDNA load (r=0.352, P=0.018) and MELD score (r=0.376, P=0.011), but nocorrelated between RORγt and ALT (r=-0.285, P=0.057).8Changes of RORγt, Foxp3and RORγt/Foxp3levels in PBMCs in differentprognostic ACLF groupsA total of28ACLF patients received dynamic RORγt and Foxp3mRNAlevels detection. According to the conditions of4-week observation, allpatients were divided into survival group (n=12) and non-survival group (n=6). The non-survival patientshad an obvious higher RORγt and Foxp3mRNAlevels compared with the survivors (P <0.05). Respectively at7-day,14-day,21day and21-day, all ACLF patients received dynamic RORγt and Foxp3mRNA levels detection. The RORγt and Foxp3mRNA levels showed aincreasing trend in survivors group, and the ratio of RORγ/Foxp3was on therise. In contrast, the non-survival group exhibited an decreasing trend.And there was significant difference on the RORγt and Foxp3mRNA levelsbetween non-survival group and survival group on the same point (P <0.05).Conclusions:1The expression level of RORγt/Foxp3mRNA in PBMCs graduallyincreased along with the severe state of illness with HBV-related liver disease.And it had achieved the peak in ACLF group.2The balance of RORγt/Foxp3maybe play an important role in theaggravation of HBV infection.3There was a positive correlation between the expression level of IL-21mRNA and RORγt mRNA. And the level of L-21mRNA closely related to theseverity of disease. IL-21may be an important effector of Th17cell in theaggravation of HBV infection.4The baseline and dynamic change of Foxp3, RORγt mRNA and IL-21in ACLF patients could evaluate the short-term prognosis.