| Clinical study of Modified Yijing Decoction on diminished ovarian reserveObjectiveTo study clinical efficacy and mechanism of the treatment on patients who had diminished ovarian reserve(DOR),withing the modified Yijing Docoction(mYJD).Methods73 DOR patients were randomly divided into the observation group and the control group,37 cases were randomly divided into Chinese medicine group(CMG)with 36 cases in western medicine group(WMG).4 cases were excluded from CMG,while 3 cases were excluded from WMG.Each group was actually completed 33 cases.CMG:at the first day after menstruation,CMG was given mYJD,until the next menstruation;Then take the next cycle of mYJD at the first day after the second menstruation(if more than 40 days of menstruation was not tide,stop taking mYJD,just take dydrogesterone tablets 20mg/d x 10d).WMG:At the fifth day of menstruation,the WMG was start to be given sequential therapy of estrogen and progesterone(progyn ova 2mg,for 21 days,plus dydrogesterone tablets 10mg,in the twelfth day,for 10 days).Then the fifth days of the next menstruation start to take the next cycle of the medicine with the same way as before.All the medicines were being taken for 6 cycles.The serum bFSH,bLH,bE2,AMH,INHB,ACTA,FS,AFC,ovarian volume,TCM syndrome score,and the pregnancy rate of the two gr oups were observed and compared between the two groups before and after t reatment.Results1.Comparing the distribution of the age,gravidity,production,course of disease,TCM symptom score before treatment,there were no differences between the two groups.Comparing the two groups,the results of serum and ultrasound examination results,also had no significant differences before treatment(P>0.05).2.Comparison of bFSH,bLH and bE2 in two groups before and after the treatment:Comparing of the level of bLH、bFSH and bE2 between the two groups before treatment,there were not significantly differences(P>0.05).Comparing with before treatment,the level of bLH and bFSH of the two groups were both significantly decreased after treatment,while the level of bE2 were both significantly increased after treatment(P<0.05).Comparing with the WMG after treatment,the level of bE2 of the CMG was significantly lower(P<0.05),while the level of bLH、bFSH of the CMG were not significantly differences(P>0.05).3.Comparison of AMH in two groups before and after the treatment:Comparing of the AMH between the two groups before treatment,there were not significantly differences(P>0.05).The AMH of CMG and WMG were significantly increased after treatment compared with before treatment(P<0.05).After treatment the AMH were not significantly differences between two groups not significantly differences.4.Comparison of INHB,ACTA and FS in two groups before and after the treatment:Comparing of the INHB,ACTA and FS between the two groups before treatment,there were not significantly differences(P>0.05).After treatment of the two groups,INHB,FS were significantly higher than those before treatment,ACTA was significantly lower than that before treatment,the differences were statistically.Significance(P<0.01);After treatment,INHB,FS,ACTA were no significant difference between the two groups(P>0.05).5.Comparison of the ultrasound results in the two groups:Comparing of the AFC between the two groups before treatment,there were not significantly differences(P>0.05).The AFC of CMG and WMG were significantly increased after treatment compared with before treatment(P<0.05).After treatment the AFC were not significantly differences between two groups not significantly differences.But the ovarian size of two groups had no difference between before treatment and after treatment(P>0.05).6.Comparison of TCM syndrome score in two groups:Before treatment,the TCM syndrome integral difference were not statistically significant between the two groups(P>0.05).The scores of menstrual cycle,menstrual quality,menstrual quantity,loss of libido,irritability,depression of two groups after treatment were both lower than those before treatment,and the difference was statistically significant(P<0.05);WMG after treatment,the menstrual cycle integral compared with CMG was significantly decreased,(P<0.05 or P<0.01);After treatment,comparing the dizziness and tinnitus,breast pain,pain of waist and knee score between the two groups to before treatment,CMG were significantly lower(P<0.05 or P<0.01),while the difference were not statistically significant in WMG(P>0.05);Comparing the dizziness and tinnitus,breast pain,pain of waist and knee score between the two groups after treatment,CMG were lower than WMG,the difference was statistically significant(P<0.05 or P<0.01).7.Comparison of the two groups after treatment of pregnancy rate:There were 9 cases of infertility in the two groups,3 cases of pregnancy in CMG after treatment,while 1 cases of pregnancy in WMG.Although there was no significant difference in the pregnancy rate between the two groups(P>0.05),but the number of pregnancy cases in CMG was higher than that in WMG.8.Comparison of the effects of two groups:The total effective rate index of two groups had no significant difference(P>0.05),the difference between the efficacy index distribution was not statistically significant(P>0.05).Conclusion1.Kidney deficiency and liver stagnation was a common pathogenesis in DOR,and Bushen Shugan was an effective therapeutic principle.mYJD could reduce the levels of FSH、LH of DOR patients,could also increase levels of E2,AMH,so as to improve the ovarian function of the patients,and mYJD in improving the ovarian function and curative effect was equal to sequential therapy of estrogen and progesterone.2.In improving the symptoms of patients with DOR,mYJD and sequential therapy of estrogen and progesterone could both improve menstrual status in patients with DOR,including the adjustment of the menstrual cycle,increase the amount of menstrual hypomenorrhea,both can improve the loss of libido,emotional irritability,depression and other symptoms,and similar efficacy between the two groups;But in improvement of patients with dizziness and tinnitus,pain of waist and knee score and breast pain and other symptoms,and mYJD was significantly better than sequential therapy of estrogen and progesterone.Therefore,mYJD is a more appropriate choice for the patients who have the need for pregnant or hormone use contraindication,for mYJD was better than western medicine,and side effects were more rare.3.mYJD and sequential therapy of estrogen and progesterone therapy both could regulate the function of INHB-ACTA-FS system to improve the ovarian function.The mechanism that mYJD improve the clinical symptoms,ovarian function,improve pregnancy may be as followings:mYJD could regular the overall function of patients by the way of multiple links,multi-channel,multi-system etc.And mYJD could improve the function of INHB-ACTA-FS system,and also couid regular the function of TGF-β super family members to promote the mature follicles,and improve ovarian function.Experimental study of Modified Yijing Decoction on diminished ovarian reserveObjectiveTo study the mechanism of the treatment of diminished ovarian reserve(DOR),withing the modified Yijing Docoction high dose group、middle dose group、low dose group and the control drug group of Progynova to intervent DOR model rats,by detecting the level of sex hormone inhibin B(INHB)-activated A(ACTA)-follistatin(FS)from serum and the level of Bcl-2/Bax on ovarian tissue from rats.So as to explore the efficacy and mechanism of modified Yi jing Docoction(mYJD)in treating DOR,and provide a theoretical basis for clinical of modified Yijing Docoction in treatment of DOR.Methods1.With using the prospective randomized research methods,360 SPF normal grade of female adult SD rats were randomly divided into blank control group(BG),model control group(MG),and the modified Yijing Decoction high(JHG),medium(JMG)and low dose group(JLG),progynova group(PG)(Except rats in NG had been induced by the same volume of saline,the other rats had been induced by 75mg/kg CTX).All the rats of the five groups were respectively given by gavage with the same volume of saline,saline,23.58g/kg YJD,11.79g/kg YJD,5.895g/kg YJD and 0.09mg/kg progynova once a day for 4 weeks.10 rats were killed in each group at each time of 1th,2th,3th,4th,6th,8th week after gavage.2.Observing the general conditions of all the rats.3.To detect the serum FSH,LH,E2,AMH,INHB,ACTA and FS in rats with enzyme linked immunosorbent assay(ELISA)technology,and the Bcl-2,Bax and AMH with immunohistochemistry(IHC).Results1.Successfully modeled:after modeling by CTX,the modelled rats were depressed、anorexia and unresponsive with curled back,loose and matt hair,limb weakness,shortness of breath,thin stool,clear long urine.All rats were found to have a phenomenon of visceral congestion in different degrees after being dissected.These were the physiological performance of rats with kidney deficiency and liver stagnation,spleen deficiency and blood stasis which showed the DOR model rats was successful.BG rats were generally in good condition:good spirit,hair smooth,normal food intake and water intake,activities freely,quick reaction,normal urine.The rats of JHG、JMG、JLG and PG were returned to normal condition at the 4th weeks after modeling:appearance,behavior,mental state,the color of stool and urine were normal,which had no difference from the rats in BG.2.Changes in estrous cycleAfter modeling,the estrous cycle of the rats changed from 4～5d extended to 7～16d,the anoestrum time changed from 1.5～2.5d extended to 5～7d.While the estrous cycle of BG and JLG rats recovery normally by the time of the 6th week after modeling;the estrus cycle of JDG and PG rats recovery by the time of the 4th week after modeling.3.Changes in FSHAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of FSH of MG rats serum were higher than that of BG,and the differences were statistically signi.ficant(P<0.01).Comparing to the 1th week after modeling,MG rats serum FSH levels of the 4th,6th,8th week after modeling were lower,those were significantly different(P<0.05 or P<0.01).Comparing with MG,the serum level of FSH of JHG and JMG rats began to decrease at 1th week after modeling(P<0.05),and continuously decrease to the 8th week after modeling(P<0.01),while that in JLG began to decrease at 2th week after modeling(P<0.05)and continuously decrease to the 4th week after model ing(P<0.05 or P<0.01).Comparing with BG,JHG、JMG and PG rats FSH level returned to normal level by the time of the 4th week after modeling(P>0.05);and JLG FSH level returned to normal level by the time of the 6th week after modeling(P>0.05).4.Changes in LHAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of LH of MG rats serum were higher than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum LH levels of the 4th,6th,8th week after modeling were lower,those were significantly different(P<0.05 or P<0.01).Comparing with MG,The serum LH level of rats in each group began to decrease at 2th week after modeling(P<0.05),while serum LH level of JHG and JMG continuously decrease to the 8th week after modeling(P<0.05 or P<0.01),and that in PG continuously decrease to the 6th week after modeling(P<0.05 or P<0.01),and that in JLG continuously decrease to the 4th week after modeling(P<0.05).Comparing with BG,JHG、JMG and PG rats FSH level returned to normal level by the time of the 2th week after modeling(P>0.05);and JLG FSH level returned to normal level by the time of the 3th week after modeling(P>0.05).5.Changes in E2At the 1th,2th,3th,4th,6th,8th week after modeling,the levels of E2 of MG rats serum were lower than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum E2 levels of the 6th,8th week after modeling were higher,those were significantly different((P<0 01).Comparing with MG,the serum level of E2 of JHG、JMG and PG rats began to increase at 1th week after modeling(P<0.05),and continuously decrease to the 8th week after modeling(P<0.05 or P<0.01),while that in JLG began to decrease at 4th week after modeling(),and continuously increase to the 6th week after modeling(P<0.05 or P<0.01).Comparing with BG,JHG、JMG and PG rats E2 level returned to normal level by the time of the 4th week after modeling(P>0.05);PG rats serum level of E2 was significantly higher than that of the other treatment groups since 1th week after modeling(P<0.05 or P<0.01).6.Changes in INHBAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of INHB of MG rats serum were lower than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum INHB levels of the 6th,8th week after modeling were higher,those were significantly different(P<0.05 or P<0.01).Comparing with MG,the serum level of INHB of JHG and JMG rats began to increase at 1th week after modeling(P<0.05),and continuously increase to the 8th week after modeling(P<0.05 or P<0.01),while that in JLG and PG began to decrease at 4th week after modeling(P<0.05).Comparing with BG,JHG and JMG rats INHB level returned to normal level by the time of the 8th week after modeling(P>0.05).JHG rats INHB level were higher than that of PG at the 6th and 8th week after modeling(P<0.05).7.Changes in ACTAAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of ACTA of MG rats serum were higher than that of BG,and the difference were statistically significant(P<0.01).Comparing with MG,the serum level of ACTA of JHG rats began to increase at 3th week after modeling(P<0.01).Comparing with BG,JHG and JMG rats ACTA level returned to normal level by the time of the 4th week after modeling,PG rats ACTA level returned to normal levelby the time of the 8th week after modeling(P>0.05).8.Changes in FSAt the 1th,2th,3th,4th,6th week after modeling,the levels of FS of MG rats serum were lower than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum FS levels of the 2th,3th,4th,6th,8th week after modeling were higher,those were significantly different(P<0.05 or P<0.01).Comparing with MG,the serum level of FS of JHG and JMG rats began to increase at 2th week after modeling(P<0.05),and continuously increase to the 8th week after modeling(P<0.05 or P<0.01),while that of PG increase at 2th week(P<0.05).Comparing with BG,JHG rats FS level returned to normal level by the time of the 2th week after modeling(P>0.05),and continuously normal to the 8th week after modeling(P>0.05),while JMG、JLG and PG rats FS level returned to normal level by the time of the 3th week after modeling(P>0.05).At the 2th,4th,6th week after modeling,he levels of FS of JHG was higher than that of other groups(P<0.05 or P<0.01).9.Changes in AMHAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of AMH of MG rats serum were lower than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum AMH levels of the3th,4th,6th,8th week after modeling were higher,those were significantly different(P<0,05 or P<0.01).Comparing with MG,the serum level of AMH of JHG、JMG and PG rats began to increase at 1th week after modeling(P<0.05),and continuously increase to the 8th week after modeling(P<0.05 or P<0.01).the serum level of AMH of JLG rats began to increase at 3th week after modeling(P<0.05),and continuously increase to the 8th week after modeling(P<0.05).Comparing with BG,JHG rats FS level returned to normal level by the time of the 8th week after modeling(P>0.05).At the 1th,2th,3th,4th,6th,8th week after modeling,the levels of AMH IOD of MG rats ovarian tissue were lower than that of BG,and the difference were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum AMH IOD levels of the 4th,6th,8th week after modeling were higher,it was significantly different(P<0.05 or P<0 01).Comparing with MG,the ovarian tissue level of AMH IOD of JHG and JMG rats began to increase at 1th week after modeling(P<0.05),and continuously increase to the 8th week after modeling(P<0.05 or P<0.01).the ovarian tissue level of AMH IOD of JLG and PG rats began to increase at 2th week after modeling(P<0.01),and that of PG rats continuously increase to the 8th week after modeling(P<0.05 or P<0.01),while that of JLG rats continuously increase to the 4th week after modeling(P<0.05).Comparing with.BG,PG、JHG and JMD rats ovarian tissue AMH IOD level returned to normal level by the time of the 6th week after modeling(P>0.05),and continuously normal to the 8th week after modeling(P>0.05).10.Changes in Bcl-2At the 1th,2th,3th,4th,6th,8th week after modeling,the levels of Bcl-2 IOD of MG rats ovarian tissue were lower than that of BG,and the differences were statistically significant(P<0.01).Comparing to the 1th week after modeling,MG rats serum Bcl-2 IOD levels of the 6th,8th week after modeling were higher,those were significantly different(P<0.01).Comparing with MG,the ovarian tissue level of Bcl-2 IOD of JHG rats began to increase at 2th week after modeling(P<0.01),and continuously increase to the 8th week after modeling(P<0.05 or P<0.01).The ovarian tissue level of Bcl-2 IOD of JMG and JLG rats began to increase at 3th week after modeling(P<0.01),and that of PG rats continuously increase to the 6th week after modeling(P<0.05 or P<0.01).Comparing with BG,JHG、JMD and JLG rats ovarian tissue Bcl-2 IOD level returned to normal level by the time of the 6th week after modeling(P>0.05),and continuously normal to the 8th week after modeling(P>0.05).PG rats ovarian tissue Bcl-2 IOD level returned to normal level by the time of the 8th week after modeling(P>0.05).Comparing with JHG,Bcl-2 IOD level of PG was lower by the 3th week after modeling,and continuously lower to he 8th week after modeling(P<0.05 or P<0.01),while that of JLG by the 3th week and 4th week after modeling(P<0.05 or P<0.01);Bcl-2 IOD level of JMG was higher than that of PG and JLG in the 3th week after modeling(P<0.01).11.Changes in BaxAt the 1th,2th,3th,4th,6th,8th week after modeling,the levels of Bax IOD of MG rats ovarian tissue were higher than that of BG,and the differences were statistically significant(P<0.05 or P<0.01).Comparing to the 1th week after modeling,MG rats serum Bax IOD levels of the3th,4th,6th,8th week after modeling were lower,those were significantly different(P<0.01).Comparing with MG,the ovarian tissue level of Bax IOD of JHG and PG rats began to decrease at 3th week after modeling(P<0.01),and continuously decrease to the 8th week after modeling(P<0.05 or P<0.01).The ovarian tissue level of Bax IOD of JMG rats began to decrease at 4th week after modeling(P<0.05 or P<0.01),and continuously decrease to the 8th week after modeling(P<0.05).The ovarian tissue level of Bax IOD of JLG rats began to decrease in 8th week after modeling(P<0.05).Comparing with BG,JHG and PG rats ovarian tissue Bax IOD level returned to normal level by the time of the 6th week after modeling(P>0.05),and continuously normal to the 8th week after modeling(P>0.05).JMG and JLG rats ovarian tissue Bax IOD level returned to normal level by the time of the 8th week after modeling(P>0.05).The rats ovarian tissue Bax IOD level of JLG was higher than that of JHG and PG in the 6th after modeling.Conclusion1.Copying 75mg/kg CTX DOR rat model was made by single intraperitoneal injection of female rats was successful.mYJD and progynova could both reduce the level of FSH and LH of DOR model rats,they could also increase E2,AMH levels.So as to improve the ovarian function of the rat model,and mYJD in improving the ovarian function and curative effect was equal to progynova.mYJD on improving rat ovarian function showed a dose-response relationship,it was that the high and middle dosage group of mYJD was better than the low dose group.2.The function of system of INHB-ACTA-FS and apoptotic cell of Bcl-2/Bax on ovarian tissue both appeared disorder in different degree after modeling,withing INHB、FS and Bcl-2 decreased,ACTA and Bax increased.Based on the previous research results,it suggested that the reason that CTX led to excessive activation of primordial follicles and reduced mature follicles,was that CTX made the function of INHB CTX-ACTA-FS to decline,and promote the speed of Papoptosis from oocyte and granulosa cells which mediated by TEN/PI3K and Tsc1/mTORC1 signaling pathway.However mYJD and progynova could obviously improved the levels of INHB、FS、Bcl-2,while decreased the level of ACTA and Bax.3.mYJD and progynova could obviously improved the levels of INHB、FS、Bcl-2,while decreased the level of ACTA and Bax.So mYJD could regulate the function of INHB-ACTA-FS system and inhibit papoptosis from oocyte and granulosa cells which mediated by TEN/PI3K and Tsc1/mTORC1 signaling pathway,to increase the primordial follicle,mature follicle,and improve ovarian function,which could provide a theoretical basis for clinical treatment of DOR. |
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