| Objective: To evaluate the clinical efficacy of Nourishing Liver and Kidney and Cholagogic Decoction in the treatment of primary biliary cholangitis(PBC).While the effect of Nourishing Liver and Kidney and Cholagogic Decoction on Th17 cell expression,the levels of IL-6,IL-17,IL-21,IL-23,TGF-beta,Act1 and expression of key proteins(TRAF6,NF-κBp65 and IκBɑ)of IL-17/Act1/NF-κB signaling pathway were observed.The possible mechanism of Nourishing Liver and Kidney and Cholagogic Decoction in the treatment of PBC was explored to provide experimental basis for the clinical application.Methods 1.Clinical research: 76 patients with PBC of deficiency of liver and kidney were randomly divided into test group and control group.The test group received Nourishing Liver and Kidney and Cholagogic Decoction combined with UDCA for 24 weeks,while the control group received UDCA only for 24 weeks.Through the 24-week clinical treatment of the two groups,the changes of Traditional Chinese Medicine(TCM)syndromes,liver function,immunoglobulin,blood lipid and liver fibrosis markers were observed.2.Experimental research: Fasting blood of enrolled PBC patients was drawn before treatment and 24 weeks after treatment.The expression ratio of Th17 cells in peripheral blood of the two groups was analyzed by flow cytometry(flow cytometry,FCM).The levels of IL-6,IL-17,IL-21,IL-23,TGF-β and Act1 in peripheral serum of the two groups were measured by enzyme-linked immunosorbent assay(Elisa).The expression of TRAF6,NF-κBp65 and IκBɑ protein in peripheral blood of the two groups was detected by Western Blot and compared with that of normal control group.Pearson correlation test was used to analyze the correlation between immune indicators and TCM syndrome score,GGT,ALP and Internal relationship of immune indicators.To investigate the possible mechanism of Nourishing Liver and Kidney and Cholagogic Decoction,the expression of Th17 cells,the levels of related cytokines and the expression of key protein in IL-17/Act1/NF-κB signaling pathway in PBC patients of test and control group were analyzed before and after treatment.Results 1.clinical research results:(1)Comparison of clinical effective rates: After 24 weeks of treatment,FAS set analysis showed that the effective rate was 84.21% in the test group and 63.16% in the control group.There was significant difference between the two groups(P < 0.027);PPS set: the effective rate in the test group was 86.49%,and the 66.67% in the control group.PPS set in the control group was consistent with the result of FAS set analysis.(2)Comparison of TCM syndrome scores: after 24 weeks of treatment,the scores of TCM syndromes in the test group and the control group were lower than those before treatment,but the scores in the test group were significantly lower than those in the control group(P = 0.014).The results of FAS set and PPS set analysis were consistent.(3)Comparison of biochemical indexes: the two groups were detected at four time points(before treatment and 8 weeks,16 weeks,24 weeks after treatment).The results showed that the ALP,GGT,ALT of the test group decreased faster and more significantly than that of the control group,and there was significant difference between the FAS sets(P = 0.000,P = 0.001,P = 0.036).The results of FAS set were consistent with those of the PPS set analysis.(4)Comparison of liver Fibrosis Indexes:After 24 weeks of treatment,the changes of HA,CIV in the test group and the control group were statistically significant(P=0.027,P=0.006),FAS set and PPS set analysis.(5)TC comparison: the TC level of the tset group and the control group decreased gradually at four time points(0,8,16,24 weeks),with significant difference(P< 0.05).After 24 weeks of treatment,the TC level of the test group was significantly lower than that of the control group(P=0.032).The results of FAS set and PPS set analysis are consistent.(6)Ig M comparison: after 24 weeks of treatment,there was significant difference in Ig M level between the test group and the control group(P = 0.010).The results of the Ig M set and those of the PPS set analysis were consistent.2.Experimental research results:(1)Comparison of expression ratio of Th17 cells in peripheral blood between PBC patients and healthy groups:The expression ratio of Th17 cells in peripheral blood of patients with PBC was significantly higher than that of healthy patients(P < 0.05).Correlation analysis showed that the expression rate of Th17 cells was not significantly correlated with TCM syndrome score and GGT,ALP.(2)Comparison of Th17-related cytokine levels between PBC patients and healthy groups:The levels of cytokines IL-17,IL-6,IL-21 and IL-23 in PBC patients were significantly higher than those in healthy controls,while the level of TGF-β1 was significantly lower than that in healthy controls(P < 0.05).Correlation analysis showed that serum IL-6,IL-17 and IL 21 levels were correlated with TCM syndrome score,IL-17,IL-6,IL-21 level was positively correlated with GGT level in PBC patients,IL-17,IL-21 level was positively correlated with ALP level in PBC patients,and Th17 cell expression ratio was positively correlated with IL-17,IL-6 level(P < 0.05).(3)Comparison of main members of IL-17/Act1/NF-κB signaling pathway in peripheral blood between PBC patients and healthy groups:Compared with the healthy group,the Act1 level of PBC patients was increased,expression of TRAF6,NF-κBp65 protein was obviously increased,and the expression of IκBɑ protein was significantly decreased.Correlation analysis showed that TRAF6 was positively correlated with NF-κBp65 and Act1,IL-6,IL-17,IL-21,and NF-κBp65 was positively correlated with TRAF6,Act1,IL-6,IL-17,IL-21(P < 0.05).There was a negative correlation between NF-κBp65 and TRAF6,NF-κBp65 and IL-6,IL-17,IL-21(P < 0.05).(4)24 weeks after treatment,the expression rate of Th17 cells in PBC patients in the test group was significantly lower than that in the control group(P=0.038),and the level of IL-6,Act1,IL-17 in PBC patients in the test group was significantly lower than that in the control group(P = 0.000,P = 0.025,P = 0.037).The expression of TRAF6,NF-κBp65 protein in the test group was obviously lower than that in the control group(P= 0.000,P = 0.009),IκBɑ protein expression was significantly higher than that in the control group(P = 0.035).Conclusion 1.Compared with UDCA alone,Nourishing Liver and Kidney and Cholagogic Decoction combined with UDCA has more advantages in the treatment of primary biliary cholangitis with deficiency of liver and kidney.2.The expression ratio of Peripheral blood Th17 cell and the levels of cytokines IL-17,IL-6,IL-21 and IL-23 in PBC patients were significantly higher than those in healthy controls,while the level of TGF-β1 was significantly lower than that in healthy controls.Symptoms in PBC patients may be associated with levels of IL-6,IL-17,IL-21.IL-17,IL-6,3.IL-21 may be more closely related to the severity of biliary epithelial cell damage and fibrosis formation and progression in PBC patients.the correlation between IL-6,IL-17,and IL-21 may be related to the feedback regulation of these cytokines.4.In this study,the expression of Act1,TRAF6,NF-κB p65 and IκBɑ in peripheral blood of PBC patients and healthy groups was detected,and it was proved that IL-17/Act1/NF-κB signaling pathway might play a very important role in the pathogenesis of PBC.4.Nourishing Liver and Kidney and Cholagogic Decoction can down-regulate the synergistic effect of IL-6,which could reduce the Synergistic Effects with TGF-β.When the transformation and overexpression of Th17 cells was down-regulate,it makes the secretion of downstream cytokines,especially the secretion of IL-17,decrease accordingly.Finally,the transmission of IL-17/Act1/NF-κB signal pathway could be down-regulate and the occurrence of downstream inflammation will be alleviated.Meanwhile,Nourishing Liver and Kidney and Cholagogic Decoction may also reduce the inflammation and cholestasis of the liver by promoting the synthesis of IκBɑ protein,re-inhibiting the activity of NF-κB and promoting the formation of spontaneous negative feedback loop. |
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