Effect of Siwutang on Diminished Ovarian Reserve in Rats Based on 16S rRNA Sequencing of Intestinal Flora / 中国实验方剂学杂志

ObjectiveTo study the effect of Siwutang （SWT） on intestinal flora in rats with diminished ovarian reserve （DOR） induced by Tripterygium wilfordii polyglycoside （TWP） based on 16S rRNA sequencing. MethodTwenty 8-week-old female SD rats were randomly assigned into four groups： blank group， model group， SWT high-dose group， and SWT low-dose group. Except the blank group， the other three groups were underwent intragastric administration of TWP tablets at a dose of 50 mg·kg-1 for 14 days. On day 15， the high-dose group was administrated at 3 times of the human dosage （40 g/person/day）， the low-dose group at 1.5 times of the human dosage， and the model group and the blank group with the same volume of normal saline for 18 days. Then， feces were collected for 16S rRNA sequencing. One hour after administration， blood was sampled from abdominal aorta after anesthesia for the measurement of hormone levels by radioimmunoassay， and ovaries were sampled， embedded， sliced， and stained with haematoxylin-eosin （HE） for pathological observation. ResultThe model group had higher level of luteinizing hormone （LH， P<0.05） and lower level of estradiol （E2， P<0.05） than the blank group. The SWT high-dose group and low-dose group had lower LH levels （P<0.05） and higher E2 levels than the model group （P<0.05）. SWT reversed the elevation in follicle-stimulating hormone （FSH） and LH levels and the decline in E2 and progesterone （P） levels caused by TWP to some extent. There were a large number of follicles at different developmental stages in the blank group， while atretic follicles increased significantly in the model group. A large number of mature follicles， secondary follicles， and primary follicles were observed in the high-dose SWT group， and primordial follicles， secondary follicles， and increased corpus luteum in the low-dose SWT group. Compared with that in the blank group and the administration group， the abundance of Verrucomicrobia and Epsilonbacteraeota in the model group significantly reduced. Compared with the blank group， the model group had different intestinal flora in phylum， class， order， family， and genus levels. Specifically， the model group had increased proportions of Firmicutes and Bacteroidetes. After TWP modeling， the abundance of Lachnospiraceae decreased significantly while that of Ruminococcaceae UCG-005 increased significantly. SWT groups， blank group， and model group can be clearly distinguished， and SWT groups had a tendency to approach the blank group. ConclusionSWT may improve the ovarian function of rats with TWP-induced DOR by regulating intestinal flora diversity.

Protection of Zuoguiwan Against Apoptosis of Follicles in Rats Injured by 60Co-γ Rays： Based on PI3K/Akt/mTOR Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the protective effect of Zuoguiwan against 60Co-γ ray-induced premature aging of rats based on the phosphatidylinositol-3-kinases/protein kinase B/mammalian target of rapamycin （PI3K/Akt/mTOR） signaling pathway. MethodSixty sexually mature female SD rats were irradiated with 60Co-γ rays （6.0 Gy， LD40） for 24 h at one time. Then they were randomized into model group， Bujiale group （0.18 g·kg-1·d-1）， Bujiale （0.09 g·kg-1·d-1） + high-dose Zuoguiwan group （23.625 g·kg-1·d-1）， high-dose Zuoguiwan group （23.625 g·kg-1·d-1）， medium-dose Zuoguiwan group （9.45 g·kg-1·d-1）， and low-dose Zuoguiwan group （4.725 g·kg-1·d-1）. The administration （once a day） lasted 21 days. Serum indexes ［follicle-stimulating hormone （FSH）， luteinizing hormone （LH）， and estradiol （E2）］ of rats were detected by enzyme-linked immunosorbent assay （ELISA）， and morphological changes of ovarian tissues were observed based on hematoxylin and eosin （HE） staining. The apoptosis rate of granulosa cells was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL） and the protein expression of phosphorylated （p）-PI3K， p-Akt， p-mTOR， B-cell lymphoma-2 （Bcl-2）， and Bcl-2-associated X protein （Bax） in ovarian tissues by Western blot. ResultCompared with normal group， model group demonstrated increase in serum FSH （P<0.01）， decrease in E2 （P<0.05）， and reduction of follicles and luteum in early ovary （P<0.01）. Moreover， the elevation of apoptosis rate of granulosa cells （P<0.01）， down-regulation of p-PI3K， p-Akt， p-mTOR， and Bcl-2 in ovarian tissue， and increase in expression of Bax were also observed in the model group as compared with the normal group （P<0.01）. In comparison with the model group， the administration groups showed rise of the number of early ovarian follicles， decrease in the apoptosis rate of granulosa cells， increase in the expression of p-PI3K， p-Akt， p-mTOR， and Bcl-2， and down-regulation of Bax， particularly the Bujiale + high-dose Zuoguiwan group（P<0.05，P<0.01）. ConclusionZuoguiwan protects radiation-damaged ovary by activating the expression of PI3K/Akt/mTOR protein in ovarian tissue， increasing Bcl-2， and inhibiting the expression of Bax.

Protection of Zuoguiwan Against Apoptosis of Follicles in Rats Injured by 60Co-γ Rays： Based on PI3K/Akt/mTOR Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the protective effect of Zuoguiwan against 60Co-γ ray-induced premature aging of rats based on the phosphatidylinositol-3-kinases/protein kinase B/mammalian target of rapamycin （PI3K/Akt/mTOR） signaling pathway. MethodSixty sexually mature female SD rats were irradiated with 60Co-γ rays （6.0 Gy， LD40） for 24 h at one time. Then they were randomized into model group， Bujiale group （0.18 g·kg-1·d-1）， Bujiale （0.09 g·kg-1·d-1） + high-dose Zuoguiwan group （23.625 g·kg-1·d-1）， high-dose Zuoguiwan group （23.625 g·kg-1·d-1）， medium-dose Zuoguiwan group （9.45 g·kg-1·d-1）， and low-dose Zuoguiwan group （4.725 g·kg-1·d-1）. The administration （once a day） lasted 21 days. Serum indexes ［follicle-stimulating hormone （FSH）， luteinizing hormone （LH）， and estradiol （E2）］ of rats were detected by enzyme-linked immunosorbent assay （ELISA）， and morphological changes of ovarian tissues were observed based on hematoxylin and eosin （HE） staining. The apoptosis rate of granulosa cells was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL） and the protein expression of phosphorylated （p）-PI3K， p-Akt， p-mTOR， B-cell lymphoma-2 （Bcl-2）， and Bcl-2-associated X protein （Bax） in ovarian tissues by Western blot. ResultCompared with normal group， model group demonstrated increase in serum FSH （P<0.01）， decrease in E2 （P<0.05）， and reduction of follicles and luteum in early ovary （P<0.01）. Moreover， the elevation of apoptosis rate of granulosa cells （P<0.01）， down-regulation of p-PI3K， p-Akt， p-mTOR， and Bcl-2 in ovarian tissue， and increase in expression of Bax were also observed in the model group as compared with the normal group （P<0.01）. In comparison with the model group， the administration groups showed rise of the number of early ovarian follicles， decrease in the apoptosis rate of granulosa cells， increase in the expression of p-PI3K， p-Akt， p-mTOR， and Bcl-2， and down-regulation of Bax， particularly the Bujiale + high-dose Zuoguiwan group（P<0.05，P<0.01）. ConclusionZuoguiwan protects radiation-damaged ovary by activating the expression of PI3K/Akt/mTOR protein in ovarian tissue， increasing Bcl-2， and inhibiting the expression of Bax.

Clinical observation on premature ovarian failure by warming acupuncture at Zusanli (ST 36) and Guanyuan (CV 4) combined with ginger moxibustion at points / 中国针灸

OBJECTIVE@#To observe the clinical effect difference between warm acupuncture at Zusanli (ST 36) and Guanyuan (CV 4) combined with ginger moxibustion at points and western medication for premature ovarian failure (POF).@*METHODS@#Fifty patients with POF were randomly assigned into an acupuncture-moxibustion group and a western medication group, 25 cases in each group. Warm acupuncture (30 min a time) at Zusanli (ST 36) and Guanyuan (CV 4) combined with ginger moxibustion (1.5 h a time) at points were used in the acupuncture-moxibustion group, once a day, 5 times a week. Three courses were given, one month as a course. Climen tablet was prescribed orally from the 5th day of menstruation in the western medication group, one tablet a time, once a day. The medication was given for 3 months, 7 days between the two 21-day medication. Before and after treatment, electrochemiluminescence was used to measure follicle stimulating hormone (FSH) and luteinizing hormone (LH); euzymelinked immunosorbent assay was applied to test estradiol (E); transvaginal color doppler sonography and abdomen ultrasound were applied to determine peak systolic velocity (PSV) of ovarian blood flow, resistance index (RI), pulsatility index (PI), ovarian volume and the number of sinus follicles (AFC). The clinical effect was compared between the two groups.@*RESULTS@#After treatment, FSH and FSH/LH were lower than those before treatment in the two groups (all 0.05).@*CONCLUSION@#Warm acupuncture at Zusanli (ST 36) and Guanyuan (CV 4) combined with ginger moxibustion at points are effective for POF patients, especially in improving FSH/LH, PSV and AFC.

A case of unusual premature ovarian failure with 46,X,dup(Xq) / 대한산부인과학회지

Premature ovarian failure (POF) is defined as the complete cessation of menses less than 40 years of age. The criteria are more than four months of amenorrhea, with serum follicle stimulating hormone value of >40 mIU/mL and the frequency of POF is about 1% of all women. Although the etiologies of POF remain unknown, suggested factors are genetic, autoimmune, chemotherapy, radiotherapy and environmental toxins. The cytogenetic abnormalities predominantly concern the X chromosome, including Turner syndrome, Fragile X syndrome and deletion, translocation, or duplication of X chromosome. We report a very rare case of premature ovarian failure with the following karyotype: 46,X,dup(Xq), and report it with a brief review of literature.

+764 G/A Mutation of Inhibin-alpha Gene in Korean Patients with Idiopathic Premature Ovarian Failure / 대한산부인과학회지

OBJECTIVE: To evaluate the incidence of the +769 G/A mutation of inhibin-alpha gene in Korean patients with idiopathic premature ovarian failure. DESIGN: DNA analysis of the mutation. METHODS: One hundred patients with idiopathic premature ovarian failure (POF) were recruited. Patient with known causes of premature ovarian failure were excluded: cytogenetic abnormalities, prior chemotherapy, prior bilateral oophorectomy and autoimmune disease, etc. DNA was extracted from peripheral blood and the +769 G/A variant of Inhibin-alpha gene (INH-alpha) was analyzed by polymerase chain reaction (PCR) and restriction fragment length polymorphism using restriction enzyme BbvI. RESULTS: We didn't find any case of +769 G/A variant in Inhibin-alpha gene in 100 Korean patients with idiopathic POF. CONCLUSION: This result suggests that in contrary to the recent data reported on the Asian population, the +769 G/A variant in inhibin-alpha gene may not exist in Korean patients with idiopathic POF.

Molecular variants of the FSH receptor exon 10 (Thr307Ala; A919G) in premature ovarian failure (POF) women by PCR-SSCP / 대한산부인과학회잡지

OBJECTIVE: This study was performed to determine whether the FSH receptor mutation is present in infertile Korean patients with 46,XX premature ovarian failure (POF) women. METHODS: The variant of FSH receptor exon 10 in thirteen 46, XX idiopathic POF and 4 healthy fertile (control) women were studied. Missense mutation in Exon 10 was detected in POF patients and healthy fertile women by polymerase chain reaction-single stranded conformation polymorphism (PCR-SSCP). RESULTS: The variant types of FSH receptor exon 10 (Thr307Ala; A919G) were found in healthy fertile (control) and POF women. CONCLUSIONS: This mutation may not be specific in POF patients and further study is needed in fertile (control) and POF women.

THE INVESTIGATION OF T CELL SUBSETS AND SUPEROXIDE DISMUTASE IN PATIENTS COITH PREMATURE OVARIAN FAILURE / 西安交通大学学报(医学版)

The change of T cell subsets of peripheralblood were tested in 12 patients with prematureovarian failure (POF), among them 10 patients superoxide dismutase (SOD) activity in erythrocytewas also tested. The results showed that the num-ber of T_3, T_4 cells T_4/T_8 ratio and SOD activitywas significantly lower in patients with POF thanin normal controls, which indicated that the func-tion of proliferation and differentiation of T lym-phocyte was inhibited, the immunoregulation wasinbalanced and the ability of resistance to lipid per-oxidation was lower in patients with POF. There-fore, the results suggests that T cell subsets andSOD activity testing in patients with POF have val-ue in the investigation of the cause of the deseaseas well as in the diagnose and treatment.