Mechanism Of Modified Guilu Erxian Ointment Based On Targeted Metabolomics In Intervening IVF Outcome In Elderly Patients With Low Prognosis Of Kidney-qi Deficiency Type

Objective: To study the effect of modified Guilu Erxian ointment for tonifying kidney and benefiting essence on the outcome of in vitro fertilization embryo transfer(IVF-ET)in elderly infertile patients with low prognosis of kidney qi deficiency,and to verify and analyze the mechanism of traditional Chinese medicine on improving elderly infertile patients with low prognosis of kidney qi deficiency from the perspective of metabolomics by using targeted metabolomics technology,To provide metabolic biomarker basis and clinical treatment ideas for the improvement of elderly infertile patients with low prognosis.Methods: This study was a double-blind,randomized and placebo-controlled trial.A total of 119 infertile patients who underwent IVF-ET in Shandong Center for reproduction and genetics of integrated traditional Chinese and Western medicine were selected.80 elderly infertile patients with low prognosis receiving IVF were randomly divided into two groups according to the R language version 3.5.1 generation table.The elderly test group and the elderly placebo group,of which 40 were the treatment group treated with modified Guilu Erxian ointment and 40 were the placebo group treated with placebo.In addition,39 young healthy women who underwent IVF-ET or ICSI due to male factors were selected,Normal control group.The three groups were treated with flexible Gn RH antagonist regimen to promote ovulation,modified Guilu Erxian ointment and placebo from the third day of menstruation to before IVF.Main outcome measures: number of eggs retrieved,number of 2PN fertilization,total number of available embryos,total number of blastocysts and total number of high-quality embryos.Secondary outcome measures: GN days,total number of GN,estrogen on HCG day,progesterone on h CG day,cumulative pregnancy rate and cumulative abortion rate.Ultra high performance liquid chromatography triple quadrupole mass spectrometry(uhplc-qtrap MS)was used to detect the metabolites in three groups of samples,including 23 cases in the experimental group,21 cases in the placebo group and 21 cases in the normal control group.The results were analyzed by univariate statistical analysis,multidimensional statistical analysis,differential metabolite screening,correlation analysis of differential metabolites and KEGG pathway analysis.Results: 1 Finally,114 patients completed the trial,including 38 cases in the experimental group,37 cases in the placebo group and 39 cases in the normal control group.2.The total score of kidney qi deficiency syndrome in the treatment group was significantly lower than that in the placebo group(P < 0.001);The scores of "lumbosacral soreness,dizziness and tinnitus,fatigue,clear and long urination" in the single syndrome scores were lower than those before treatment,and the difference was statistically significant(P < 0.05).There was no statistically significant change in the scores of "more and less menstruation,irregular stool,insomnia and forgetfulness,dry mouth and throat,fear of cold and cold limbs"(P > 0.05).3.Compared with the placebo group,the number of eggs obtained,the number of 2pn fertilized,the number of high-quality embryos,the total number of available embryos,estrogen on h CG day and the cumulative pregnancy rate in the treatment group increased,and the cumulative abortion rate decreased,the difference was statistically significant(P < 0.05).There was no statistically significant difference in other indexes,but there was an improvement trend.4.The results of targeted metabolomics analysis showed that the follicular fluid in the treatment group gathered with the normal young group and deviated from the placebo group.A total of 55 significantly different metabolites were found in the follicular fluid of elderly patients with low prognosis and normal young patients with kidney qi deficiency,mainly including amino acids,fatty acids,bile acids,indoles,hydroxyl acids and dicarboxylic acids,mainly involving protein digestion and absorption,glycine,serine and threonine metabolism,biosynthesis of amino acids Nicotinate and nicotinamide metabolism.5.After the intervention of modified Guilu Erxian ointment,L-aspartic acid,glycine,L-serine,palmitoleic acid,palmitelaidic acid,L-Alanine,gamma linolenic acid,alpha linolenic acid,n-acetyltryptophan and other metabolites were down regulated,mainly involving amino acid metabolism and fatty acid metabolism.Conclusion: 1 Modified Guilu Erxian ointment can effectively improve the clinical symptoms and IVF outcome of elderly patients with low prognosis of kidney qi deficiency.2.There are differences in follicular fluid metabolites between elderly patients with low prognosis of kidney qi deficiency and normal women.Among them,the eight metabolites of L-aspartic acid,L-Alanine,aminoadipic acid,L-asparagine,L-arginine,L-serine,gamma linolenic acid,pentadecanoic acid and alpha linolenic acid are closely related to elderly patients with low prognosis of kidney qi deficiency,which may be inferred as biomarkers.3.The intervention mechanism of Jiawei Guilu Erxian ointment may be mainly through regulating amino acid metabolism and fatty acid metabolism.