| BackgroundThe physiological loss of female primitive germ cells begins after birth,and ends in menopause when germ cells are exhausted and ovarian function is permanently lost.With the progression of the disease of Premature Ovarian Insufficiency(POI),the patients’ ovarian function compared with normal women will be reduced or exhausted in advance.Prevention and early treatment of POI may be more meaningful than treatment after the disease occurrence,but the optimal time-point of prevention and early treatment is not clear.Our previous research found that electroacupuncture pretreatment before and during modeling may have a certain protective effect on the ovaries of POI rats by regulating the expression of apoptosis-related factors Bcl-2 and Bax proteins.However,the effect of electroacupuncture pretreatment at different time on ovarian function of POI rats is different and the mechanism is unclear.The team’s previous clinical study found that electroacupuncture can improve serum FSH,E2,and LH levels in patients with ovarian insufficiency at 12 weeks of treatment,and the effect remained with no side effects 12 weeks after discontinuation of treatment.However,the difference between electroacupuncture and drugs on reproductive hormones in patients with ovarian insufficiency is not clear.This study is divided into two parts:experimental study and clinical study.Through experimental research from the perspective of biological mechanisms and molecular basis,we intend to reveal different protective effect and mechanism of acupuncture pretreatment on protecting the ovarian function in POI rat at different time-point.Based on the determination of the optimal intervention time,the protective effect and mechanism of electroacupuncture combined with GnRH-a pretreatment on ovarian function of POI rats will be further discussed.Then a prospective clinical cohort trial will be conducted to investigate the effect of electroacupuncture compared with hormone replacement therapy(HRT)on reproductive hormones in patients with POI,and to provide ideas for the optimization of acupuncture and moxibustion prevention and treatment of POI.ObjectiveExperimental studyExperiment 1 The different effect of electroacupuncture pretreatment on protecting ovarian function of POI rats.Experiment 2 The molecular mechanism of electroacupuncture pretreatment to protect ovarian function of POI rats based on PI3K/Akt/mTOR pathway.Experiment 3 Comparison the effects and possible mechanism of electroacupuncture,gonadotropin-releasing hormone agonist(GnRH-a)and electroacupuncture combined with GnRH-a on protecting ovarian function in POI rats.Clinical study The efficacy and safety of electroacupuncture and drugs in improving serum reproductive hormone levels in patients with premature ovarian insufficiency,a prospective observative study.MethodsExperimental studyExperiment 1 Sixty SD female rats with regular estrous cycles were randomly divided into blank control group,model group,electroacupuncture group I(electroacupuncture before modeling),electroacupuncture group Ⅱ(electroacupuncture during modeling)and electroacupuncture group III(electroacupuncture after modeling),with 12 rats in each group.Model group,electroacupuncture group Ⅰ,Ⅱ and Ⅲ were injected with 4vinylcyclohexene diepoxide(VCD)at 80mg/kg body weight(VCD dissolved volume ratio of sesame oil 1:17)for 15 days.Electroacupuncture "Zhong Liao","TianShu" and"Uterus" acupoints were needled 5 times a week,40min each time,20 times in continuous 4 weeks in group Ⅰ,group Ⅱ and group Ⅲ,respectively,4 weeks before modeling,2 weeks before modeling and 2 weeks after modeling,and 4 weeks after modeling.The blank control group was fed normally.The model group was modeled normally without electroacupuncture intervention.All the rats were sacrificed after successful modeling in the model group.The rate of weekly change of body weight,the disorder of estrous cycle,ovarian diameter and ovarian index of rats in each group were detected.The number of follicles at each level was calculated by observing the morphology of follicles under light microscope.Cell apoptosis was detected by TUNEL method,and serum AMH,FSH and E2 were detected by enzymic immunosorbent assay(ELISA).Experiment 2 Sixty SD female rats with regular estrous cycle were randomly divided into blank control group,model group,electroacupuncture group Ⅰ,Ⅱ,Ⅲ,PI3k inhibitor(LY294002)combined with electroacupuncture group,with 10 rats in each group.The model group,electroacupuncture group Ⅰ,Ⅱ and Ⅲ,and inhibitor combined electroacupuncture group were injected intraperitoneally with VCD continuously for 15 days,and the specific electroacupuncture method was the same as experiment 1.The inhibitor combined with electroacupuncture group was intraperitoneally injected with2.5mg/(kg*d)LY294002 solution one hour before each electroacupuncture,followed by 4 weeks of electroacupuncture pretreatment intervention.The pretreatment intervention method of electroacupuncture Ⅰ,Ⅱ and Ⅲ groups were the same as experiment 1,and the blank control group was fed normally.The model group was modeled normally without electroacupuncture intervention.All the rats were sacrificed after successful modeling in the model group.Ovarian index was detected,and Western Blot was used to detect the expression of downstream proteins of PI3k/Akt/mTOR signaling pathway and the expression of pathway-related mRNA was detected by fluorescence quantitative PCR.Experiment 3 Sixty SD female rats with regular estrous cycle were randomly divided into blank control group,model group,electroacupuncture group,GnRH-a group and electroacupuncture combined GnRH-a group,with 12 rats in each group.Model group,electroacupuncture group,GnRH-a group,electroacupuncture combined with GnRH-a group,VCD was continuously injected intraperitoneally for 15 days,the specific electroacupuncture method was the same as experiment 1.Electroacupuncture group and electroacupuncture combined with GnRH-a group were pretreated with el ectroacupuncture 4 weeks before modeling,and GnRH-a group and electroacupuncture combined with GnRH-a group were subcutaneously injected with GnRH-a 0.4ug per time 2 weeks before modeling,once in total.The blank control group was fed normally.The model group was modeled normally without electroacupuncture intervention.All the rats were sacrificed after successful modeling in the model group.The rate of weekly change of body weight,the disorder of estrous cycle,ovarian diameter and ovarian index of rats in each group were detected.The number of follicles at each level was calculated by observing the morphology of follicles under light microscope.Cell apoptosis was detected by TUNEL method,and serum AMH,FSH and E2 were detected by enzymic immunosorbent assay(ELISA).Western Blot was used to detect the expression of downstream proteins in the PI3k/Akt/mTOR signaling pathway and fluorescence quantitative PCR was used to detect the mRNA expression of the pathway.Clinical researchFrom October 2019 to January 2022,Guang’anmen Hospital conducted a prospective cohort study,in which 32 patients were collected and randomly selected electroacupuncture or drug regimen according to the patients’ wishes(whether to combine HRT or not was determined according to the patients’ conditions).17 patients in the electroacupuncture group received only electroacupuncture intervention,2-3 times a week for 3 consecutive months,while 15 patients in the drug group received 12 weeks of TCM or 12 weeks of TCM combined with HRT.The main outcome index was to compare the mean value of serum FSH changes from baseline between the two groups after 12 weeks of treatment,and to preliminarily explore the efficacy and safety of electroacupuncture compared with drugs to improve serum hormones in patients with POI.ResultsExperiment 1:Effect of electroacupuncture pretreatment on the protection of ovarian function in rats with POI at different time.①The proportion of estrous cycle disorder was 100%(12/12),36.4%(4/11),60%(6/10)and 80%(8/10)in model group,electroacupuncture group Ⅰ(electroacupuncture before modeling),electroacupuncture group Ⅱ(electroacupuncture during modeling)and electroacupuncture group Ⅲ(electroacupuncture after modeling),respectively.②Ovarian index:Compared with blank control group,the ovarian index of model group was significantly decreased(P=0.007<0.01);Compared with the model group,the ovarian index of electroacupuncture group Ⅱ increased,with statistical significance(P=0.002<0.01).The ovarian index of electroacupuncture group Ⅰ was higher than that of electroacupuncture group Ⅲ(P=0.030<0.05).The ovarian index of electroacupuncture group Ⅱ was higher than that of electroacupuncture group Ⅲ(P=0.042<0.05),and the difference was statistically significant.③Ovarian morphology and follicular count:Follicular morphology:Compared with blank control group,preantral follicles of ovarian tissue in model group were significantly reduced,atresia follicles were significantly increased,secondary follicles were damaged and arranged loosely,a large number of inflammatory cells could be seen locally,and interstitial glands were significantly increased;Compared with the model group,the degree of ovarian damage in electroacupuncture group Ⅰ was lighter,the number of preantral follicles,primary follicles and secondary follicles was more than that in the model group,the granulosa cell layer around follicles was relatively intact,and the follicular fluid was enriched and intact.Piles of preantral follicles were observed in the ovarian cortex of electroacupuncture group Ⅱ,and the number of atresia follicles was significantly reduced.A large number of atresia follicles and a small amount of inflammatory cell infiltration were seen in group Ⅲ.Compared with the blank control group,the proportion of preantral follicles in the model group was significantly reduced and the proportion of atresia follicles was significantly increased(all P<0.01).Compared with the model group,the proportion of preantral follicles in electroacupuncture groups Ⅰ,Ⅱ and Ⅲ was increased and the proportion of atresia follicles was significantly decreased(all P<0.05),especially in electroacupuncture group Ⅱ,the proportion of preantral follicles was significantly increased and the proportion of atresia follicles was significantly decreased(P=0.009<0.01).There was statistical difference in the proportion of follicle count among the three groups.The ratio of preantral follicles in electroacupuncture group Ⅰ was higher than that in groupⅡ(P=0.030<0.05),and the ratio of atretic follicles was significantly lower(P=0.014<0.05).Compared with electroacupuncture group Ⅲ,the proportion of preantral follicles in group Ⅱ was higher(P=0.021<0.05),and the proportion of atresia follicles was significantly lower(P=0.016<0.05),and the difference was statistically significant.③ Ovarian TUNEL staining:Compared with blank control group,AOD in model group was significantly higher(P<0.01).Compared with the model group,AOD in electroacupuncture group Ⅰ and Ⅲ decreased(P<0.05),and significantly decreased in electroacupuncture group Ⅲ(P<0.01).Pairwise comparison between electroacupuncture groups showed no significant statistical difference.(P=0.711>0.05).④Serum hormone levels:Compared with blank control group,serum AMH in model group was significantly decreased,while serum FSH and LH levels were significantly increased(AMH P=0.027<0.05,FSH P=0.040<0.05,LH P=0.030<0.05).Compared with model group,serum AMH and E2 in electroacupuncture group Ⅱ were significantly increased(AMH P=0.034<0.05,E2 P=0.048<0.05.),and serum FSH,LH and PROG levels were significantly decreased(FSH P=0.036<0.05,LH P=0.033<0.05).PROG P=0.048<0.05).The serum LH level in electroacupuncture group Ⅰ and Ⅲ decreased(P<0.05),especially the serum LH level in electroacupuncture group Ⅲ was significantly lower than that in model group(P=0.009<0.01).There were statistical differences in serum hormone levels among the three groups.Compared with group Ⅱ,serum AMH in group Ⅱ was higher(P=0.043<0.05),serum FSH and serum PROG were lower(FSH P=0.033<0.05,PROG P=0.033<0.05).Compared with electroacupuncture group Ⅲ,serum FSH in group Ⅱwas lower(P=0.029<0.05).The serum PROG in electroacupuncture group Ⅰ was lower than that in group Ⅲ(PROG P=0.029<0.05),and the difference was statistically significant.Experiment 2:The molecular mechanism of electroacupuncture pretreatment to protect ovarian function of POI rats based on PI3K/Akt/mTOR pathway.Part Ⅰ:①Upstream and downstream protein expression of PI3k/Akt/mTOR pathway in ovarian tissues:Compared with blank control group,the proportion of Bcl-2/Bax,pAkt/Akt and p-mTOR/mTOR in model group were decreased(P=0.048<0.05),decreased(P=0.048<0.05)and increased(P=0.033<0.05).Compared with the model group,the protein expression level of Bcl-2/Bax in electroacupuncture group Ⅰ was increased(all P<0.05),and the protein expression level of p-Akt/Akt was increased in electroacupuncture group Ⅰ,Ⅱ and Ⅲ(all P<0.05),especially in electroacupuncture group Ⅲ(P<0.01).There was statistical difference in the proportion of gray value ratio of each protein among the three groups.Two comparison between electroacupuncture group,the electroacupuncture group I compared with group Ⅲ,the ratio of the Bcl2/Bax was higher in group Ⅰ(P=0.041<0.05),the ratio of the p-Akt/Akt was significantly higher higher in group Ⅲ(P<0.01).The p-Akt/Akt ratio of electroacupuncture group Ⅰwas higher than that of electroacupuncture group Ⅱ(P=0.045<0.05).The p-Akt/Akt ratio of electroacupuncture group Ⅲ was higher than that of electroacupuncture groupⅡ(P<0.01).②Fluorescent quantitative PCR was used to detect mRNA expression levels in ovarian tissue:Compared with blank control group,the relative expression levels of Akt and mTOR mRNA in model group were decreased(all P<0.05);Compared with the model group,the mRNA relative expression levels of PI3k,Akt and mTOR were increased in the electroacupuncture group Ⅰ and Ⅲ(all P<0.05),especially the mRNA relative expression levels of Akt were significantly increased in the electroacupuncture group Ⅲ(all P<0.01).The mRNA level of Bcl-2 in electroacupuncture group Ⅱ and group Ⅲ was significantly increased(P<0.01).The mRNA relative expression levels of the three groups were statistically different.Compared with electroacupuncture group Ⅲ,PI3k,Akt and Bcl-2 were significantly higher in group Ⅰ(P=0.041<0.05).Compared with electroacupuncture group Ⅱ,PI3k and Akt were significantly higher in group Ⅲ(all P<0.05),especially Akt was significantly higher in group Ⅲ(P<0.01).There was no significant difference in mRNA expression between group Ⅰ and group Ⅱ(all P>0.05).Part Ⅱ:①Ovarian index and uterine index:Compared with blank control group,uterine index and ovarian index in model group were significantly lower,with statistical differences(uterine P=0.044<0.05,ovarian P=0.036<0.05);Compared with model group,the ovarian index of electroacupuncture group was significantly increased,and the uterine index of inhibitor was significantly increased,with statistical differences(P<0.05).Compared with electroacupuncture group,the uterine index and ovarian index of inhibitor were significantly increased,with statistical differences(P<0.05 for both groups).② The expression of upstream and downstream proteins of PI3k/Akt/mTOR pathway in ovarian tissues:compared with blank control group,Bcl2/Bax(P=0.030<0.05)and p-Akt/Akt(P=0.048<0.05)in model group were significantly decreased;Compared with model group,the ratio of Bcl-2/Bax and pAkt/Akt protein in electroacupuncture group was significantly increased(P=0.048<0.05),especially p-mTOR/mTOR(P=0.002<0.01).Compared with the electroacupuncture group,the protein ratio of Bcl-2/Bax increased and the p-Akt/Akt ratio decreased in the inhibitor combined electroacupuncture group(all P<0.05),especially the p-mTOR/mTOR ratio decreased significantly(P=0.003<0.01).③Fluorescent quantitative PCR was used to detect mRNA expression levels in ovarian tissue:Compared with blank control group,the relative expression levels of Akt,mTOR and IGF-1 mRNA in model group were decreased by fluorescence quantitative PCR(P<0.05).Compared with the model group,the relative expression levels of Bcl-2 and ER-β mRNA in electroacupuncture group were increased(all P<0.05),and the relative expression levels of IGF-1 mRNA in inhibitor combined electroacupuncture group were increased(P<0.05).Compared with electroacupuncture group,the mRNA relative expression levels of Bcl-2,ER-β and IGF-1 in inhibitor combined with electroacupuncture group were increased(P<0.05).Experiment 3:The effects and possible mechanism of electroacupuncture,gonadotropin-releasing hormone agonist(GnRH-a)and electroacupuncture combined with GnRH-a on ovarian function of POI rats.①Weekly change rate of body weight in each group:Compared with blank control group,the change range of body weight in model group was significantly smaller at the first and third month of modeling(all P<0.05);Compared with model group,body weight of electroacupuncture group,GnRH-a group and electroacupuncture combined GnRH-a group was slightly larger than that of model group in 1-2 months(all P<0.05),body weight of electroacupuncture group was maintained and increased significantly in 3 months of modeling(P=0.045<0.05).Compared with the electroacupuncture group,the weight gain of the GnRH-a group and electroacupuncture combined GnRH-a group was similar to that of the electroacupuncture group at the 1 st and 2nd month,but the increase rate of GnRH-a group and electroacupuncture combined GnRH-a group was significantly lower than that of the electroacupuncture group at the 3rd month(both P<0.05).②Proportion of estrous cycle disorder:Estrous cycle disorder was different in each group,including 100%(12/12)in the model group,36.4%(4/11)in the electroacupuncture group,25%(3/12)in the GnRH-a group and 3.3%(4/12)in the electroacupuncture combined with GnRH-a group.③Ovarian index:The ovary of the blank control group was red and full,while that of the model group was dark and deep.The ovary of the GnRH-a group and electroacupuncture combined with GnRHa group were pale,and the slightly ruddy ovary of the group of electroacupuncture combined with GnRH-a was better than that of the group of GnRH-a.The ovarian index of model group was lower than that of blank control group(P=0.007<0.01).Compared with the model group,the ovarian index of electroacupuncture group was increased(P=0.037<0.05),with statistical significance.Compared with electroacupuncture group,the ovarian index of GnRH-a and electroacupuncture combined with GnRH-a group was lower(P=0.028<0.05).④Follicular morphology and follicular count:Compared with blank control group,preantral follicles of ovarian tissue in model group were significantly reduced,atresia follicles were significantly increased,secondary follicles were damaged and arranged loosely,a large number of inflammatory cells could be seen locally,and interstitial glands were significantly increased;Compared with the model group,the ovarian damage of electroacupuncture group and electroacupuncture combined with GnRH-a group was lighter,the number of preantral follicles and secondary follicles was slightly more than that of the model group,the granulosa cell layer around follicles was relatively intact,and the follicular fluid was enriched and intact in electroacupuncture group and electroacupuncture combined with GnRH-a group.In the GnRH-a group,scattered preantral follicles were seen in the ovarian cortex,and the number of atresia follicles was significantly reduced.Compared with blank control group,the proportion of preantral follicles in model group was significantly decreased and the proportion of atretic follicles was significantly increased(all P<0.01).Compared with model group,the proportion of preantral follicles in electroacupuncture group,GnRH-a group and electroacupuncture combined with GnRH-a group increased,while the proportion of atresia follicles decreased significantly(all P<0.01).There were differences among the pretreatment groups.Compared with GnRH-a group,the proportion of primary follicles in electroacupuncture combined with GnRH-a group was significantly lower(P<0.01).⑤TUNEL apoptosis detection:Compared with blank control group,AOD in model group was significantly higher(P<0.01);Compared with model group,AOD in electroacupuncture group was decreased(P=0.041<0.05),and significantly decreased in electroacupuncture combined with GnRH-a group(P<0.01).The pair-topair comparison of AOD in all pretreatment groups showed statistical difference,and compared with GnRH-a group,electroacupuncture combined with GnRH-a group significantly decreased(P<0.01).⑥Serum hormone levels in each group:compared with blank control group,serum AMH and PROG in model group were significantly decreased,while serum FSH and LH levels were significantly increased(all P<0.05);Compared with model group,serum AMH was significantly increased and serum LH level was significantly decreased in GnRH-a group(P<0.05).Serum AMH in electroacupuncture combined with GnRH-a group was significantly increased(P<0.01),while serum FSH and LH levels were significantly decreased(P<0.05).Compared with GnRH-a group,serum PROG in electroacupuncture group was higher(P<0.05).Compared with electroacupuncture group,electroacupuncture combined with GnRH-a group showed significantly higher serum AMH(P<0.01)and lower serum PROG and FSH levels(P<0.05).Compared with GnRH-a group,the serum AMH of electroacupuncture combined with GnRH-a was significantly higher(P<0.01),and the level of FSH was lower(P=0.032<0.05).⑦Expression of upstream and downstream proteins of PI3k/Akt/mTOR pathway in ovarian tissue:Compared with blank control group,the protein ratios of p-Akt/Akt,p-mTOR/mTOR and Bcl-2/Bax in model group were decreased(all P<0.05);Compared with model group,the protein ratios of p-Akt/Akt and p-mTOR/mTOR in electroacupuncture group,GnRH-a group and electroacupuncture combined GnRH-a group were increased(all P<0.05),especially the p-Akt/Akt ratio in electroacupuncture combined GnRH-a group was significantly increased(P=0.002<0.01).Pairwise comparison of pretreatment showed statistical difference among the three groups.Compared with electroacupuncture group,p-mTOR/mTOR ratio in GnRH-a group was slightly increased(P=0.039<0.05);Compared with GnRH-a group,the protein ratio of Bcl-2/Bax in electroacupuncture group and electroacupuncture combined with GnRH-a group was slightly increased(all P<0.05),while the p-Akt/Akt ratio was significantly increased in electroacupuncture combined with GnRH-a group(P=0.006<0.01).⑧Fluorescent quantitative PCR was used to detect mRNA expression levels in ovarian tissue:Compared with blank control group,the relative expression levels of Akt and mTOR mRNA in model group were decreased(all P<0.05);Compared with the model group,the mRNA relative expressions of PI3k,Akt and mTOR were increased in the electroacupuncture group and electroacupuncture combined with GnRH-a group(all P<0.05),especially the mRNA relative expression of Akt was significantly increased in electroacupuncture combined with GnRH-a group(P<0.01).The mRNA level of Bcl-2 was significantly increased in electroacupuncture combined with GnRH-a group(P=0.004<0.01).Pairwise comparison of pretreatment showed statistical difference among the three groups.Compared with electroacupuncture group,the relative expression of Bcl-2 mRNA was slightly increased(P=0.037<0.05),and Akt was significantly increased(P=0.006<0.01)in electroacupuncture combined with GnRH-a group.Compared with GnRH-a group,mRNA relative expression levels of PI3k and Bcl-2 were slightly increased(both P<0.05),while Akt was significantly increased(P<0.01)in electroacupuncture combined with GnRH-a group.Clinical researchA preliminary comparison of efficacy and safety of electroacupuncture contrast drugs in improving serum hormones in patients with POI.①The main outcome measure was the difference of serum FSH at week 12 from baseline.After 12 weeks of treatment,there was no significant difference in the difference of serum FSH reduction between the electroacupuncture group and the drug group(HRT combined with Traditional Chinese Medicine)(Electroacupuncture group:-13.16(95%CI,-14.49 to-11.83),the drug group:-7.71(95%CI,-10.88 to4.54);Difference between groups:-5.45(95%CI,-12.79 to 1.89),P=0.134>0.05);②After 12 weeks of treatment,the difference of serum E2 level increase from baseline was better in the electroacupuncture group than in the drug group(48.64(95%CI,24.18 to 73.10)and-57.67(95%CI,-95.76 to-19.58);Difference between groups:106.31(95%CI,13.81 to 287.87),P=0.015<0.05).③At 12 weeks after the end of treatment,the difference in serum FSH reduction from baseline was better in the drug group than in the electroacupuncture group(-8.84(95%CI,-11.09 to-6.59)and-15.93(95%CI,18.09 to-13.77);Difference between groups:7.09(95%CI,0.51 to 13.04),P=0.035<0.05).④The line chart of the effects of 12-week electroacupuncture on serum FSH and LH showed that 12-week electroacupuncture had a more significant effect on reducing FSH than LH.⑤Compared with the drug group,after 12 weeks of treatment,the menstrual cycle was slightly increased,the length of menstrual period was slightly shortened,and the amount of menstrual period was slightly increased compared with before treatment.There was no statistical difference between the groups.⑥Pregnancy status:In the electroacupuncture group,2 women were successfully pregnant during the treatment period,1 woman(1/14)was conceived by IVF-ET,and 1 woman(1/14)was conceived naturally;In the drug group,1 person(1/12)conceived naturally during the treatment period.All three pregnancies were successful.⑦Safety evaluation:There were 2 cases of abdominal subcutaneous hematoma in the trial process,and the hematoma was completely relieved within 2 weeks.In the drug group,2 patients showed mild dizziness and nausea due to taking progesterone pills.No other adverse reactions were recorded.ConclusionExperimental study1.The ovarian function of POI rats was protected by electroacupuncture pretreatment at different time-point,and the protective effect of electroacupuncture pretreatment before and during modeling was more obvious than that after modeling.2.The different effect of electroacupuncture at different time-point on protecting ovarian function in POI rats was related to the up-regulation of the expression of Akt and mTOR downstream molecules of PI3K/Akt/mTOR pathway in ovarian tissues by electroacupuncture,and then up-regulation of Bcl-2 and down-regulation of Bax to different degrees.3.Compared with electroacupuncture pretreatment or subcutaneous GnRH-a pretreatment before modeling,electroacupuncture combined with GnRH-a pretreatment before modeling could better protect the ovarian function of POI rats.And the mechanism may be related to the up-regulation of the expression of Akt and mTOR downstream molecules of PI3K/Akt/mTOR pathway in ovarian tissues.Clinical researchThe efficacy of 12-week electroacupuncture was comparable to that of TCM combined with HRT in improving reproductive hormone levels in patients with POI,and the efficacy lasted until 12 weeks after treatment.Electroacupuncture had certain advantages in improving serum sex hormone levels in patients with POI. |
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