| Objective:To observe the expression levels of regulatory T cells(Treg), T helper cell 17(Th17) transcription factors forkhead /winged helix trans- criptionfactor(Fox P3), Orphan nuclear receptorγt(RORγt) m RNA in peripheral blood mononuclear cells(PBMC) of chronic HBV infected patients with different stages, and to analyse the relationship with liver function alanine aminotransferase(ALT), aspartate aminotransferase(AST), total bilirubin(TBIL), HBs Ag, HBe Ag, HBV DNA. Discussing the possible role of Treg and Th17 cells may have played in the immune-tolerant phase in patients with chronic HBV infection. We tend to using the peripheral blood indicators to explore the state of immune-tolerance.Maybe we can immuno-therapy HBV infection early effective treatment through breaking immunetolerance.Methods:83 outpatients or hospitalization chronic HBV infected patients without anti-HBV therapy from Tianjin Second People’s Hospital from 2014 April to 2015 March were collected, including 24 cases of immunetolerance, 24 cases of immuneclearance,and 35 cases of inactive or low(non) copy. Detecting Fox P3, RORγt m RNA relative expression level in PBMC using real-time quantitative polymerase chain reaction(RT-q PCR). Detecting subjects’ ALT, AST, TBIL using automatic biochemical analyzer, and detecting HBV serological markers HBs Ag, HBe Ag, HBe Ab using electrochemical luminescence, HBV DNA quantification using real-time quantitative PCR. 14 cases of healthy people were collected as control group.Comparing Fox P3,RORγt m RNA relative expression levels in each group.Exploring its correlation with liver function, virological indicators and its role in the immunetolerant phase of chronic HBV infection maybe have played. Choosing 27 patients who had examined liver biopsy from the 83 subjects and analyzing their liver tissue fibrosis and inflammation. Studying the relationship of the Fox P3, RORγt m RNA relative expression levels with live biopsy.Result:1. The difference of Fox P3 m RNA relative expression between HBV infected patients groups and healthy controls group was statistically significant(P <0.05). All immunetolerance, immunecleaner and inactive or low(non) copy groups patients, the Fox P3 m RNA expression level were higher than healthy control group,and the difference were statistically significant(P <0.05). The Fox P3 m RNA relative expression levels in immunetolerant phase was higher than that in immuneclearance phase and non-active or low(non) copy group patients(0.252 ± 0.410 vs 0.195 ± 0.381,0.206 ± 0.498), and the difference was statistically significant(P<0.05).The difference was not statistically significant between the immuneclearance phase and non-active or low(non) copy phase(P = 0.737).2. The relative expression of RORγtm RNA in immuneclearance phase was higher than that in healthy control group, immunetolerance group, inactive or low(non) copy group(0.421 ± 0877 vs 0.102 ± 0.485,0.222 ± 0.743,0.230 ± 0.391), and the difference were statistically significant(P <0.01). The RORγtm RNA relative expression levels in immunetolerant phase, inactive or low(non) copy phase were higher than that in healthy control group(0.222 ± 0.743,0.230 ± 0.391 vs 0.102 ± 0.485)(P <0.05). The difference of RORγt m RNA relative expression level between immunetolerant phase and inactive or low(non) copy phase was not statistically significant(P = 0.831).3. Fox P3 m RNA relative expression levels were not significantly associated with ALT, AST,TBIL,but had a significant positive correlation with HBV DNA(P<0.05). RORγt m RNA relative expression levels were significantly correlated with ALT, AST(P<0.01),but there were no significant correlation with TBIL, HBV DNA. Fox P3 m RNA relative expression had a significant positive correlation with the relative expression of RORγt(r = 0.728, P<0.001).4. In the 83 sujects,the relative expression level of Fox P3 m RNA was higher in DNA positive group than that in DNA negative group(0.271 ± 0.341 vs 0.208±0.669), and the difference was statistically significant(P<0.05). The relative expression level of RORγtm RNA in DNA positive group was higher than that in DNA negative group(0.409 ± 0.693 vs 0.362 ± 0.406), the difference was not statistically significant(Z =-0.367,P= 0.714).5. In the 83 patients, the relative expression level of Fox P3 in HBe Ag positive group was lower than that in HBe Ag negative group(0.165 ± 0.328 vs 0.232 ± 0.507), but the difference was not statistically significant(P= 0.884). The relative expression level of RORγt m RNA in HBe Ag positive group was higher than that in HBe Ag negative group(0.361 ± 0.823 vs 0.240 ± 0.398), and the difference was statistically significant(P <0.05).6. Analysing the 27 patients who had recepted liver biopsy pathologic examination from the 83 research cases. 2 patients were in immune tolerance period(pathological diagnosis: G1S1-2, G1-2 S1), and whose liver tissue was mild inflammation damage and fibrosis. 14 patients were in immune clearance phase(pathological diagnosis :2 cases of liver cirrhosis, 3 cases with inflammatory grading G1, 9 cases with G≥2; 8 cases with fibrosis degre stage S1 and 4 caseswith S≥2). 11 patients were in non activity period(8 cases with inflammation degree G1, 3 cases with G2; 9 cases with the degree of fibrosis stage between S1 and S2, 2case with S≥2).7. In the 27 patients who had recepted live biopsy examination, the relative expression levels of Fox P3 m RNA in inflammation degree G2 group patients(14cases) was higher than that in G1 group(0.162 ± 0.280 vs 0.044 ± 0.066, P <0.05). RORγt m RNA relative expression levels in inflammation degree G2 group patients was higher than that in G1 group(0.489 ±1.135 vs 0.073 ± 0.062, P <0.05).In the patients that liver fibrosis for S> 1,RORγt m RNA relative expression level in PBMC was higher than that in S1 group(0.589 ± 1.273 vs 0.083 ± 0.076, P <0.05),but the Fox P3 m RNA relative expression levels was not statistically significant(P>0.05).Conclusion:1.Imbalance of Th17 / Treg cells is presence of immune tolerance phase of HBV infection.Monitoring the transcription factor Fox P3 m RNA,RORγtm RNA,which will help to evaluate the body’s immune state and may provide reference for antiviral treatment2.In the immune tolerance phase the advantage lay with Treg,which maybe one of the reasons leading to the body’s immune tolerance state. Th17 cell dominate in the immune clearance phase. Th17 cell have promote inflammation effect and is good for the removal of the virus.3. Fox P3 m RNA is positive related with HBV DNA loads. Fox P3 m RNA suppress the immune response,which make HBV DNA high replicate. RORγtm RNA promote inflammation.Inflammatory response returned to normal and RORγtm RNA decrease expression after the HBe Ag seroconversion occurred,4. RORγtm RNA may play an important role in the fibrosis process of liver cells,which can help to judge the degree of fibrosis or assess liver cells fibrosis presentence or not.5. In addition to dynamic testing liver function and virological indicators in patients with chronic HBV infected on time,monitoring patients’ Treg, Th17 cells and their associated transcription factors are also important.we can understand the body’s immune status and change, which may provide reference value for choosing anti-HBV treatment time. |
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