Clinical And Metabolomic Study On Bushen Culuan Recipe In Treatment Of Ovulatory Infertility Caused By Polycystic Ovary Syndrome

1.Clinical study on treatment of ovulatory infertility caused by polycystic ovary syndrome with Bushen Culuan Recipe.1.1 Objective To observe the clinical efficacy and safety of Bushen Culuan Recipe in the treatment of ovulatory infertility caused by polycystic ovary syndrome(PCOS).1.2 Methods The clinical trial method of multi center,random,double-blind,double simulation and positive drug control was adopted.According to the diagnostic criteria and inclusion and exclusion criteria of traditional Chinese medicine and Western medicine,94 patients with PCOS infertility of kidney deficiency and blood stasis type who met the entry conditions were strictly screened.The drugs were given successively according to the drug random number,47 in the treatment group and 47 in the control group.Both groups began to take the test drug on the fifth day of menstrual cycle or the fifth day of withdrawal and bleeding of progesterone and estradiol valerate+progesterone.Treatment group:take Bushen Culuan Recipe,3 times/day,1 bag/time,for 14 days;At the same time,take clomiphene placebo,50mg/tablet,once a day,one tablet/time,for 5 consecutive days.Control group:take clomiphene,50mg/tablet,once a day,one tablet/time,for 5 consecutive days;At the same time,take Bushen Culuan Recipe placebo,3 times/day,1 bag/time,for 14 consecutive days.Continuous treatment for 1-2 courses(each course is 3 menstrual cycles),and follow-up for 12 months after treatment.The basal body temperature(BBT)was monitored,the number of small follicles,the volume of ovary,the thickness of endometrial EM and whether ovulation were monitored by B-ultrasound,the changes of serum sex hormones and ovarian function(LH,FSH,PRL,E2,P,T,F-T,INHB,AMH)were detected,the score of TCM syndrome and signs before and after treatment were recorded,and the safety indexes were detected.SPSS statistical software was used to analyze the data by using t-test,chi square test and rank sum test.The effective rate,ovulation rate,pregnancy rate,changes of sex hormone and ovarian function,changes of B-ultrasound,TCM syndrome score and sign score of the two groups after treatment were compared to determine the curative effect and evaluate the safety.1.3 Results The results of treating PCOS infertility of kidney deficiency and blood stasis type with Bushen Culuan Recipe are as follows:(1)Total curative effect:in the treatment group,the curative ratio was 53.19%,the actovoty ratio was 25.53%,the efficiency was 14.88%,the ineffective rate was 6.38%,and the overall response rate was 93.62%;while control group,the recovery rate was 12.76%,the effective rate was 23.40%,the actovoty ratio was 42.55%,the inefficiencies was 21.27%,and the total effective rate was 78.72%.P<0.05,the difference between the two groups was significant,which showed that the total effective rate of Bushen Culuan Recipe was higher than that of clomiphene.(2)Ovulation probability:the probability in the therapy group was 48.8%;The probability of control group was 53.37%.The ovulation rate of clomiphene in the control group was higher than that of Bushen Culuan Recipe,and the difference was not significant(P>0.05).(3)Pregnancy rate:in the treatment group,24 cases were pregnant after treatment,23 cases were not pregnant,and the pregnancy rate was 51.06%;In the control group,there were 5 cases of pregnancy and 42 cases of non pregnancy after treatment,and the pregnancy rate was 10.64%.Chi square test was used for statistics,P<0.05.,make clear that the pregnancy rate of Bushen Culuan Recipe was higher than that of clomiphene.(4)Sex hormone and ovarian function:the therapy group was evidently so much the better than the control group in reducing LH,T,INHB,LH/FSH ratio and increasing P level.There were significant differences by rank sum test(P<0.05);After treatment,FT in the two groups was tested by t-test.FT in the treatment group decreased significantly,with significant difference(P<0.05);There was no obvious distinction in PRL,E2 and AMH between two groups.After treatment,the levels of LH,T,FT,INHB,AMH,LH/FSH and E2 and P in the treatment group had significant improvement by comparison with before;The control group had significant improvement in reducing LH,AMH and LH/FSH after treatment(P<0.05),and there was no significant difference in other indexes before and after treatment(P>0.05).(5)B-mode ultrasonography:statistics showed that after treatment,there were significant differences in reducing the number of 2-9mm follicles,ovarian volume and increasing the thickness of endometrium between the treatment group and the control group(P.Before and after treatment,the number of small follicles and ovarian volume in the treatment group were significantly reduced,and the thickness of endometrium was significantly thickened(P<0.05).Before and after treatment,the number of follicles in the control group was significantly improved(P<0.05),and there was no significant change in ovarian volume and endometrial thickness(P>0.05).(6)BBT of basal body temperature:comparing the BBT of the two groups before and after treatment,the BBT normalization rate of 47 patients in the treatment group was 70.21%and that of 47 patients in the control group was 40.43%.Chi square test was used to count the two groups,P<0.05.The BBT normalization result of the therapy group was sharp better.(7)TCM symptom score:after treatment,the symptoms of amenorrhea,late menstruation,less menstruation,weak waist and knee,mental fatigue,sluggish menstruation,multi color and dark menstrual blood,menstrual blood block,menstrual abdominal pain,ecchymosis on tongue,ecchymosis,fine pulse and so on in the treatment group were significantly improved compared with the control group.There was no distinguish between the two groups in the number of times of urination and tinnitus.(8)physical traits:post-therapy,it is vary widely in BMI,hairiness score,acne score and black acanthosis score between the two groups(P<0.05).All signs in the treatment group were all improved significantly,BMI and acne scores in the control group were all improved than before(P<0.05),and hairy and black acanthosis had no significant changes.(9)Safety evaluation:the patients in the treatment group and the control group were made blood,urine,liver,kidney routine test,and ECG.No obvious abnormalities were found.No adverse reactions or events was found.(10)Follow up results:94 patients out of the treatment group were followed up.In the treatment group,there were 7 cases of spontaneous labor,5 cases of cesarean section and 2 cases of biochemical abortion;In the control group,there were 1 case of cesarean section and 1 case of biochemical abortion.1.4 Conclusion After treatment,the total effective rate and pregnancy rate of Bushen Culuan Recipe were significantly higher than those of clomiphene.It had an obvious regulatory effect on sex hormone and ovarian function.It could significantly reduce LH,LH/FSH ratio,T,FT,INHB,and significantly increase the levels of P and E2,so as to improve the polycystic state of ovary,reduce the number of small follicles and ovarian volume,and increase the thickness of endometrium;Bushen Culuan Recipe has curative effect is good and there are no toxic and side effects.2.Clinical characteristics and risk factors of thyroid function and glucose and lipid metabolism in infertile PCOS patients with kidney deficiency and blood stasis2.1 Objective To observe the clinical characteristics of thyroid function and glucose and lipid metabolism in infertile PCOS patients with kidney deficiency and blood stasis,and to analyze the risk factors affecting glucose and lipid metabolism.2.2 Methods The subjects were 55 infertile PCOS patients with kidney deficiency and blood stasis who met the standard of natriuretic peptide I.Research Indicators:height,weight and BMI were measured;Sex hormones(FSH,LH,E2,P,T),thyroid function(TSH,Anti TG,Anti TPO),blood lipids(CHOL,TG,HDL-C,LDL-C),oral glucose tolerance test and insulin release test were measured,and IR index was calculated.They were grouped according to glucose tolerance,BMI,blood lipid,IR and age;The data statistical software was SPSS 26.0.The risk factors related to glucose and lipid metabolism were analyzed "by univariate and multivariate logistics.2.3 Results 55 cases of PCOS infertility patients with kidney deficiency and blood stasis accounted for 52.7%and 47.3%respectively;50.9%had normal BMI,49.1%were overweight and obese;TSH<2.5 mIU/L accounted for 65%,TSH group>2.5 mIU/L accounted for 35%,insulin sensitivity accounted for 49%,IR accounted for 51%;70%had normal glucose tolerance and 30%had IGT;55%of the patients had normal blood lipid and 45%had abnormal blood lipid.The analysis showed that thyroid function,IR,IGT and dyslipidemia had no significant effect on sex hormones in PCOS infertility patients with kidney deficiency and blood stasis;According to the clinical analysis of the characteristics of nail work and glucose and lipid metabolism,it was found that with the increase of age,BMI and IR increased significantly,while HDL-C and T promoting liver gluconeogenesis decreased significantly;Homo-IR,fasting and 1-2 hours postprandial blood glucose,insulin level and blood lipid in overweight and obese patients were significantly higher than those with normal BMI;Univariate analysis showed that dyslipidemia,IGT and BMI were significant risk factors for IR,and IR was also a significant risk factor for IGT;Abnormal BMI and TSIR are significant risk factors.Logistic regression analysis showed that dyslipidemia was an independent risk factor for IR,while IR was an independent risk factor for IGT;At the same time,BMI and IR are independent risk factors of dyslipidemia.2.4 Conclusion (1)Clinical characteristics of 55 PCOS infertility patients with kidney deficiency and blood stasis type:47.3%were elderly,35%had abnormal TSH,51%had IR,29%had IGT,and 45%had dyslipidemia.(2)BMI and IR were positively correlated with age in PCOS infertile patients with kidney deficiency and blood stasis type;(3)TSH and FSH in infertile patients with kidney deficiency and blood stasis type were negatively correlated,while IR,IGT,dyslipidemia and sex hormones were not significantly correlated;(4)The BMI of infertile patients with PCOS due to kidney deficiency and blood stasis is a significant influencing factor for HOMO-IR,fasting and 1-2 hours postprandial blood glucose and insulin levels,and blood lipid levels,suggesting that clinical attention should be paid to monitoring blood sugar in patients,although it does not reach Diabetes diagnostic criteria,but potentially higher risk;(5)TSH,BMI and IR in infertile patients with kidney deficiency and blood stasis type PCOS are independent risk factors for dyslipidemia;IR is an independent risk factor for IGT;IR and dyslipidemia are independent risk factors for each other.(6)In conclusion,IR in infertile patients with kidney deficiency and blood stasis type PCOS is directly and positively correlated with BMI,age factors,and TSH,and is also an independent risk factor for IGT and dyslipidemia,and IR can lead to hyperandrogenism and ovulation.obstacle.3.Metabonomics Study of Bushen Culuan Recipe in the.treatment of PCOS infertility with kidney deficiency and blood stasis3.1 Objective Non targeted metabolomics was used to compare the differences of serum metabolites between the treatment group and the control group of PCOS infertility patients with kidney deficiency and blood stasis before and after treatment,find the relevant KEGG signal pathway of the different metabolites,and analyze the relationship between the different metabolites and the sexual hormone and ovarian function of the subjects,so as to explore the mechanism of Bushen Culuan Recipe in the treatment of PCOS infertility with kidney deficiency and blood stasis from the metabolomic level.3.2 Methods 26 patients(13 cases in the treatment group and 13 cases in the control group)who met the diagnosis and discharge criteria of PCOS infertility of kidney deficiency and blood stasis type in clinical study 1 were selected.Group Z is the experimental group,Z1 is before treatment and Z2 is after treatment;Group D is the control group,D1 is before treatment and D2 is after treatment.The metabolites in the sera of all subjects were analyzed by liquid chromatography-mass spectrometry.After extracting the data,the multivariate statistical analysis was carried out by partial least squares discriminant analysis and cluster analysis,and the differential metabolites were compared according to zl-z2,dl-d2 and d2-z2.Finally,all differential metabolites were summarized and analyzed to find the rise and fall trend and metabolic pathway of differential metabolites after treatment with Bushen Culuan Recipe.Finally,the relationship between the differential metabolites and the subjects’ sex hormones and ovarian function was analyzed.3.3 Results Metabonomic analysis display that there were great changes between zl-z2 groups.The metabolites were ornithine,tryptophan,azelaic acid,4-pyridoxine,glutaric acid,cholic acid,palmitoleic acid,linoleic acid,lauroyl carnitine and triglyceride;There was little difference in the content of metabolites between dl-d2 groups;There were significant differences in amino acids,acyl carnitine and oleic acid in d2-z2 group.A total of 24 kinds of foreign bodies with poor total metabolism were selected.After treatment with Bushen Culuan Recipe,the up-regulated substances were cholic acid,phosphatidylcholine,4-pyridoxine β Sitosterol;Down regulated include glutaric acid,palmitoleic acid,azelaic acid,linoleic acid,aspartic acid,4,5-dihydrowhey acid,phenylalanine,5,6-dihydrouridine,lactic acid,Decanoyl carnitine,triglyceride,lauroyl carnitine,cholesterol,uric acid,tyrosine,tryptophan,serine,threonine,ornithine and citric acid,involving lipid metabolism,amino acid metabolism,carnitine metabolism and other pathways.KEGG signaling pathway involves 66 pathways,including bile secretion,aminoacyl tRNA biosynthesis,fat digestion and absorption,fat digestion and absorption,citric acid cycle and so on.The correlation analysis of differential metabolites with sex hormones and ovarian function showed that E2,P and cholic acid β-Sitosterol,4-pyridoxine and phosphatidylcholine(hemolysin)were positively correlated;There was no significant correlation between PRL and different metabolites.FSH,and,AMH,inhb and other indicators are related to cholic acid β-Metabolites such as sitosterol,4-pyridoxine and phosphatidylcholine(hemolysin)showed atypical negative correlation,and atypical positive correlation with various amino acids and lipid metabolites(all P>0.05).LH,t,FT and cholic acid β-Sitosterol,4-pyridoxine and phosphatidylcholine(hemolysin)were strongly negatively correlated and positively correlated with various amino acids and lipid metabolites.3.4 ConclusionThe study of non targeted metabolomics showed that BBushen Culuan Recipe acted on 66 related KEGG metabolic pathways through 24 different metabolites to achieve the purpose of treating PCOS infertility of kidney deficiency and blood stasis type.The action process involves amino acid metabolism,lipid metabolism,carnitine metabolism,purine metabolism,energy metabolism and so on.Moreover,there is a correlation between sex hormones and foreign bodies with poor metabolism,indicating that the change of metabolic results has practical significance for PCOS infertility of kidney deficiency and blood stasis type.