| Objective:Objective to evaluate the clinical efficacy of “Xiao Yi Yu Si Decoction” in the treatment of IVF caused by EMs of kidney deficiency and blood stasis type,and to explore its possible targets and pathways from the perspective of proteomics.Method:In the clinical observation part,a total of 84 patients were included,including 54 cases of infertility patients with kidney blood type of EMs who underwent IVF-ET in the reproductive center of Affiliated Hospital of Shandong University of traditional Chinese medicine(TCM)from August 2019 to October 2020.They were randomly divided into the TCM intervention group(24 cases)and the non intervention group(30 cases);in addition,30 women who underwent IVF-ET solely for male factors in the same period were selected as the blank control group.The three groups were treated with luteal phase short acting long protocol for ovarian hyperstimulation,subcutaneous injection of triptorelin acetate 0.05 mg daily from the middle luteal phase(20th day of menstruation),and the TCM intervention group was treated with TCM from the down-regulation day to the day trigger.Observe the changes of kidney deficiency and blood stasis syndrome score before and after treatment in TCM intervention group and non intervention group;record the number of eggs,normal fertilization rate,effective embryo number,high-quality embryo rate,clinical pregnancy rate,and cumulative pregnancy rate of embryos obtained in this cycle.In the experimental study,5ml follicular fluid samples from 10 women in EM-TCM group,10 women in EM-NON group and 10 women in EM-CON group were randomly selected by using label-free quantitative proteomics technology for mass spectrometry experiment and data analysis;the data were collected and analyzed by protein extraction,peptide enzymolysis,chromatographic grading,LC-MS/MS.The original differential protein information was obtained by protein database retrieval and other steps,and the differential proteins and pathways that may be related to the infertility of kidney blood type EMs and the curative effect of TCM were screened by the screening criteria of multiple change > 1.2 times and P value < 0.05(t test or other);the differential proteins and pathways with the best specificity and sensitivity were determined by bioinformatics analysis and ROC curve drawing.Results:(1)The final completion of the experiment was 84 cases,24 cases in EM-TCM group;30 cases in EM-NON group and 30 cases in EM-CON group.The results of clinical experiments showed that there was no significant difference in baseline characteristics among the three groups(P>0.05);After the intervention of TCM,the score of kidney deficiency and blood stasis syndrome was significantly lower than that before treatment and before intervention,and the difference was statistically significant(P <0.01);Total Gn,HCG day E2 and P levels were not significantly different between the three groups(P > 0.05);The number of oocytes,normal fertilization rate and high-quality embryo rate in EM-TCM group were significantly higher than those in EM-NON group(P<0.05);The comparison of cumulative pregnancy rate among the three groups had no significant difference(P > 0.05),but the EM-TCM group showed an increasing trend compared with EM-NON group and EM-CON group(50.00% vs.43.44% vs.33.33%);There was no significant difference in the clinical pregnancy rate of fresh embryo transfer and frozen embryo transfer,and live birth rate among the three groups(P>0.05).(2)The results showed that a total of 256 proteins were identified in the three groups,including 203 quantifiable proteins,EM-TCM vs.EM-NON group,five differential proteins were found;EM-NON vs.EM-CON group,a total of 15 differential proteins were found;EM-TCM vs.EM-CON group,a total of 20 differential proteins were found.(3)The results of bioinformatics and statistical analysis showed that the three groups of differential proteins were mainly from extracellular;EM-NON vs.EM-CON group,AUC of IGLV7-46、IGHG2 and IGKV2-30 were both greater than0.8,P<0.05,AUC of IGLV7-46 was 0.852,the best sensitivity was 89%,the best specificity was 78%;AUC of IGHG2 was 0.815,the best sensitivity was78%,the best specificity was 89%;AUC of IGKV2-30 was 0.864,the best sensitivity was 78%,the best specificity was 89%;EM-TCM vs.EM-NON group,five differential proteins were found: APOE、HRG、ATRN、A1AG2 and SAMP,among which ATRN was expressed in both EM-NON vs.EM-CON group and EM-TCM vs.EM-NON group,KEGG analysis showed that the cholesterol metabolism pathway changed relativly significant after the TCM intervention.Conclusion:TCM “Xiaoyiyusi Decoction” can significantly improve the clinical symptoms of EMs-related infertility women with kidney deficiency and blood stasis,and relatively improve the outcome of IVF treatment.IGLV7-46,IGHG2 and IGKV2-30 can be used as potential diagnostic markers for EMs-related infertility.ATRN may be the main target of “Xiaoyiyusi Decoction”,and Cholesterol metabolism may be one of the main pathway of“Xiaoyiyusi Decoction”. |
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