| Objective: To investigate the impact of Qinghua Jiedu Decoction on HBV DNA of chronic HBV infection with a low viral load(LVL-CHB),and to uncover the underlying its molecular mechanism by network pharmacology.Methods: This study consists of two parts.The first part was a clinical retrospective cohort study.LVL-CHB Patients(HBs Ag and/or HBV DNA positive for more than 6 months,HBV DNA was larger than the detection limit but less than 2000 IU/m L)were recruited from the hepatitis clinic of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine(TCM)from October 2018 to March 2019.The patients fell into two groups: the TCM group(67 cases)and the control group(34 cases).The TCM group was given Qinghua Jiedu Decoction and the control group was not given any medicine.Both groups were observed for 96 consecutive weeks.The HBD-DNA negative conversion rate,decline in HBV-DNA titer,HBs Ag titer,HBs Ag negative conversion rate,HBs Ag serological conversion rate,HBe Ag titer,HBe Ag negative conversion rate,HBe Ag serological conversion rate,hepatic function,abdominal B-ultrasound and TCM syndrome score were contrasted between two groups.The second part is the network pharmacology research.Firstly,targets obtained from TCM database TCMSP and disease database Gene Card and OMIM were used to construct a network of "Qinghua Jiedu Prescription-Target – LVL-CHB".Secondly,the key nodes of Qinghua Jiedu Formula for treating LVL-CHB were screened through network analysis.Furthermore,by the enrichment analysis of GO function and KEGG signaling pathway,the signaling pathway involved in the treatment of LVL-CHB by Qinghua Jiedu Formula was obtained.Finally,the effects of Qinghua Jiedu Decoction on the screened key targets were observed at the cell and animal levels.Results: 1.Clinical study showed that at 24,48,72 and 96 weeks,HBV DNA titer in the TCM group was 2.07±1.22,1.70±1.22,1.65±1.24 and 1.44±1.20(lg IU/m L),respectively and in the control group was 2.64±0.55,2.63±0.55,2.69±0.68 and 2.69±0.79(lg IU/m L),respectively.HBV DNA titer in the TCM group was obviously lower than those in the control group at each time point(P < 0.05).At 24,48,72 and 96 weeks,HBV DNA negative conversion rate was 21.9%,29.7%,32.8% and 37.5% in the TCM group and 0%,0%,0%,0% and 2.8% in the control group,respectively.Compared with control group,the HBV DNA negative conversion rate in TCM group was obviously higher at every time point(P < 0.05).HBs Ag titer in the TCM group decreased significantly from 24 weeks,however,the HBs Ag titer between the two groups has no stactistically difference at every time point.At week 48,72 and 96,HBs Ag negative conversion rate in the TCM group and the control group was 1.6% vs 0%,3.1% vs 0% and 6.3% vs 0%,respectively,but the difference was not statistically significant.At week 48 and 96,HBs Ag serological conversion rate was 1.6% vs.0% in both TCM and control groups,but the difference was not statistically obvious.HBe Ag titer in the TCM group decreased with the extension of treatment time,however,there was no statistical difference in the HBe Ag titer between the two groups at each time point.At week 48,72 and 96,the negative conversion rate of HBe Ag in the TCM group was 8.3%,8.3% and 16.7%,respectively,and 0% in the control group,however,the difference was not statistically obvious.At week 72 and 96,the HBe Ag serological conversion rate was 8.3%,in comparison with 0% in the control group,however,the difference in the two groups has no statistically difference.2.At 24,48,72 and 96 weeks,the total bilirubin in the TCM group was 14.17±4.84,13.79±4.68,14.40±5.48 and 13.78±4.065 umol/L,respectively and in the control group was 16.48±5.94,17.92±9.98,16.45±7.80 and 15.22±5.801 umol/L,respectively.Compared with control group,the total bilirubin in serum of TCM group was obviously lower at every time point(P < 0.05).Abdominal B ultrasound score in the TCM group gradually decreased,however,the difference between the two groups was statistically significant at every time point.3.Compared with control group,the TCM syndrome score of the TCM group was obviously lower at every time point(P < 0.05).The effective rate of TCM syndromes of the TCM group and control group was 96.88% vs 36.11%(P < 0.05).4.There were no abnormal changes in blood routine,electrocardiograph and renal function related to the clinical trial.5.A total of 4035 target targets of Qinghua Jiedu Formula and 211 target targets of LVL-CHB were screened out through network pharmacology,and then 178 target targets of Qinghua Jiedu Formula for the treatment of LVL-CHB were obtained by taking the intersection.The candidate targets were statistically analyzed,and the top 10 targets were obtained: AKT1,ALB,IL6,VEGFA,CASP3,JN,MAPK8,EGFR,MAPK1,and MYC.KEGG enrichment analysis showed that the treatment of LVL-CHB by qinghua jiedu prescription was mainly related to MAPK signaling pathway,RAS signaling pathway,erb B signaling pathway,c AMP signaling pathway,calcium signaling pathway and RAP1 signaling pathway.6.Studies on mice and Hep G2 cells showed that Qinghua Jiedu Decoction significantly reduced the HBV DNA titer and the contents of HBs Ag and HBe Ag,while increased the HBs Ab content.Meanwhile,the expression of EGFR,which can promote HBV infection effectively,was decreased.Conclusion: Qinghua Jiedu Decoction significantly reduces the serological level of HBV DNA and increases HBD-DNA negative conversion rate in LVL-CHB patients likely by down-regulating the expression of EGFR.This provides a new idea for the application of traditional Chinese medicine in the treatment of LVL-CHB. |
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