Exploring The Mechanism Of Cangfu Daotan Decoction On Phlegm-Dampness Type Of Polycystic Ovary Sydrome Based On MiR-17-5p/ATG7 Mediated Autophagy

Objective:Abnormal activation of autophagy is considered to be an important mechanism of polycystic ovary syndrome.Our team found that the expression of autophagy related gene ATG7 increased in granulosa cells of patients with phlegm-dampness type of polycystic ovary sydrome.This study explored the pathogenesis of PCOS based on autophagy and revealed the correlation between autophagy and the clinical phenotype of PCOS.Intervention with Cangfu Daotan Decoction was used to explore the efficacy of traditional Chinese medicine in the treatment of PCOS and the possible microscopic efficacy mechanism.Methods:Part 1:According to the prediction results of bioinformatics software,the associated micro RNA of ATG7 was verified by double luciferase experiment.Part 2:76 patients who met the diagnostic criteria of phlegm-dampness type of polycystic ovary sydrome were randomly divided into experimental group（EG,Cangfu Daotan Decoction+IVF/ICSI）and observation group（OG,placebo+IVF/ICSI）.Another 38 patients who underwent assisted reproduction due to male factors were selected as the control group（CG,IVF/ICSI）.The purpose of this study was to observe the improvement of clinical symptoms and the changes of detection indexes after adjuvant Chinese medicine treatment.The three groups of patients were treated with antagonist regimen for controlled ovarian stimulation.The changes of TCM phlegm-dampness syndrome score and body mass index（BMI）in the EG and the OG before and after treatment were analyzed.Clinical indicators such as the use days of gonadotropin（Gn）,the dosage of Gn,the level of estradiol（E₂）、progesterone（P）、and luteinizing hormone（LH）on the day of ovulation triggering,the number of oocytes retrieved,2pn fertilization rate,high-quality embryo rate and other indicators of the three groups were analyzed.q PCR and Western Blot were used to detect the expression levels of ATG7 and its associated micro RNA in granulosa cells of the two groups of patients.Results:Part 1:miR-17-5p is a micro RNA associated with ATG7.miR-17-5p can bind to the 3’UTR of target gene ATG7 and inhibit its expression.Part2:1.TCM phlegm-dampness syndrome score:The score of phlegm-dampness syndrome decreased in the EG after treatment（P<0.05）;The integral difference of phlegm-dampness syndrome before and after treatment in the EG was higher than that in the OG（P<0.05）.2.BMI:the BMI decreased in EG after treatment（P<0.05）;The difference of BMI before and after treatment in the EG was greater than that in the OG（P<0.05）.3.Gn days and Gn dosage:Gn days in the EG was lower than that in the OG（P<0.05）,and both the EG and the OG were higher than that in the CG（P<0.05）;The dosage of Gn in the EG and the OG was higher than that in the CG（P<0.05）.There was no significant difference between the EG and OG（P>0.05）,but the dosage of the EG has a decreasing trend.4.E₂,P and LH levels on trigger day:the E₂levels on the day of ovulation triggering in the EG were higher than those in the OG and the CG（P<0.05）;There was no significant difference in LH level and P level among the three groups（P>0.05）.5.The number of eggs retrieved:the number of eggs retrieved in the CG was lower than that in the EG and the OG（P<0.05）;There was no significant difference between the EG and the OG（P>0.05）,but the EG has an increasing trend.6.2pn fertilization rate and high-quality embryo rate:there was no significant difference in 2pn fertilization rate among the three groups（P>0.05）,but the EG showed an increasing trend compared with the OG;The rate of high-quality embryos in the EG and the CG was higher than that in the OG（P<0.05）.7.Comparison of the expression of miR-17-5p,ATG7 in the three groups:the relative expression of miR-17-5p in the EG and the OG was significantly lower than that in the CG（P>0.05）,and that in the OG was lower than that in the EG（P<0.05）;The relative expression of ATG7 in the EG and the OG was higher than that in the CG（P<0.05）.There was no significant difference between the EG and the OG,but the relative expression of ATG7 in the EG tended to decrease.The ATG7protein expression level in the OG was higher than that in the EG and the CG（P<0.05）,and that in the EG was higher than that in the CG（P<0.05）.Conclusions:1.In PCOS patients,low expression of miR-17-5p may lead to excessive autophagy of granulosa cells by up regulating the expression of ATG7,thus participating in the occurrence of PCOS;2.Cangfu Daotan Decoction can significantly improve the clinical symptoms of patients with phlegm-dampness PCOS,improve the 2pn fertilization rate and high-quality embryo rate,up regulate the expression of miR-17-5p and down regulate the expression of ATG7;3.The improvement of eggs and embryo quality of PCOS patients by Cangfu Daotan Decoction may be related to Cangfu Daotan Decoction inhibiting the expression of ATG7 by up regulating the expression of miR-17-5p,so as to weaken the excessive autophagy of granulosa cells in PCOS patients.