Electro-acupuncture attenuates the mice premature ovarian failure via mediating PI3K/AKT/mTOR pathway.

AIMS: Electro-acupuncture (EA) is frequently recommended as a complementary therapy for premature ovarian failure (POF) in the clinical. However, little information exists about its potential treatment mechanisms. The study was designed to observe the effect of EA to ovarian function and fertility in POF mice model, and investigated its potential mechanisms on PI3K/AKT/mTOR signaling pathway. MATERIALS AND METHODS: Forty-five female C57/BL6 mice were divided into the Control, the Model and the EA group. The ovaries morphology of mice was observed by hematoxylin and eosin (HE) staining, and all follicles were counted under microscope. The protein expression of PI3K, phospho-PI3K, AKT, phospho-AKT, mTOR, phospho-mTOR, S6, phospho-S6, 4E-BP1 and phospho-4E-BP1 were detected by western blotting. The data was presented as the ratio of phosphorylation protein to total protein. Serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2) and anti-Mullerian hormone (AMH) levels were measured by enzyme-linked immunosorbent assay (ELISA). The fertility was observed by giving treated mice 8 weeks for breeding. KEY FINDINGS: We found that primordial follicle counts were increased in EA group compared to Model group. The phosphorylation of PI3K, AKT, mTOR, 4E-BP1 and S6K in EA group significantly reduced compared to Model group. Serum FSH and LH levels in EA group were decreased compared to Model group, while, serum E2 and AMH levels in EA group were increased compared with Model group. The litter size in EA group was improved compared to Model group. SIGNIFICANCE: The effects of EA on the PI3K/AKT/mTOR signaling pathway may represent one of the mechanisms involved in attenuating the mice POF.

Hormone-like activities of Kuntai capsule in the uteri of ovariectomized rats and immature rabbits.

OBJECTIVE: To investigate the hormone-like activities of Kuntai capsule (KTC) in the uteri of ovariectomized rats and immature rabbits. METHODS: Following bilateral ovariectomy, rats were randomly divided into six groups including sham group, control group, estradiol valerate group, KTC 0.24, 0.6, and 1.5 g/kg groups. The rats were treated with 0.5% CMC-Na, estradiol valerate and KTC (0.24, 0.6, and 1.5 g/kg), respectively for 28 consecutive days. Then the estrous cycle, uterine changes and pathological changes were examined. Serum levels of estradiol (E2), and progesterone (P4) were measured by enzyme-linked immunosorbent assay (ELISA). Protein levels of estradiol receptor (ER), progesterone receptor (PR), vascular endothelial growth factor (VEGF), proliferating cell nuclear antigen (PCNA) and nuclear-associated antigen-67 (Ki-67) in uterine tissues were detected by western blot. Immature rabbits were estrogen-primed prior to intragastric administration with KTC for 6 d consecutively. Then, the uteri underwent hematoxylin-eosin staining to observe endometrial transformation. RESULTS: Compared with the control group, 0.6 and 1.5 g/kg KTC markedly decreased the uterine organ coefficient and endometrial thickness (P < 0.05). The serum level of P4 was increased in the KTC 0.6 g/kg group (P < 0.05). There were no significant variations in the serum level of E2 in the KTC groups compared with the control group. ER¦Â, but not ER¦Á, was markedly upregulated after KTC administration (P < 0.05). Furthermore, 1.5 g/kg KTC significantly decreased the protein level of PRA (P < 0.05) and 0.6 g/kg KTC increased the protein level of PRB in the uteri (P < 0.05). VEGF was highly expressed after treatment with 0.24 and 0.6 g/kg KTC, and Ki-67 was markedly reduced in ovariectomized rats treated with 1.5 g/kg KTC. No difference was found in the expression of PCNA. KTC 0.24 and 0.6 g/kg promoted endometrial transformation in immature rabbit uteri. CONCLUSION: KTC does not demonstrate obvious estrogen-like effect on uteri after ovariectomy, but it does exhibit weak progestogen-like effect, by which mechanism of action is yet to be further investigated.

Podophyllotoxin Extracted from Juniperus sabina Fruit Inhibits Rat Sperm Maturation and Fertility by Promoting Epididymal Epithelial Cell Apoptosis.

This study aimed to investigate the antifertility effect of Juniperus sabina fruit on male rats and its possible mechanism, and hence it might be developed as a potential nonhormonal male contraceptive. Male rats were intragastrically fed for consecutive 8-week and 4-week recovery with the fruit of J. Sabina, and sperm maturation, serum testosterone level, and histopathology were analyzed. Epididymal epithelial cell culture was prepared for detection of podophyllotoxin activities. Furthermore, cell proliferation, transmission electron microscopy, Annexin V/Propidium iodide, TUNEL, RT-PCR, ELISA, and western blotting were examined. The results showed that rat sperm motility and fertility were remarkably declined after feeding the fruit. Moreover, the fruit targeted the epididymis rather than the testis. After 4-week recovery, more than half of the male rats resumed normal fertility. It was found that podophyllotoxin significantly inhibited epididymal epithelial cell proliferation, promoted cell apoptosis, and increased the mRNA and protein levels of TNF-α and the expression levels of cytochrome c, caspase-8, caspase-9, and caspase-3. Our findings suggest that the fruit of J. sabina could inhibit male rat sperm maturation and fertility. The potential mechanism might be related to podophyllotoxin, inducing epididymal epithelial cell apoptosis through TNF-α and caspase signaling pathway.

Kuntai Capsule Inhibited Endometriosis via Inducing Apoptosis in a Rat Model.

We evaluated the effectiveness of Kuntai Capsule (KTC) for treating endometriosis using rat model and investigated its preliminary mechanism of action involved. SD rats were implanted with endometrial tissues and treated with KTC for three weeks. Then, laparotomy was performed to examine volume changes of the autografts. The serum levels of TNF-α, IL-6, COX-2, E2, and P4 were measured through ELISA. TUNEL was performed to analyze the apoptosis on ectopic endometrium. Protein levels of caspases 8, 9, and 3 and cytochrome c in the ectopic and eutopic endometrium were measured by western blotting. Results showed that KTC significantly decreased the volumes of ectopic endometrium. The level of TNF-α increased and E2 decreased in the KTC treatment groups. TUNEL and western blot assay showed that KTC could induce apoptosis of endometriotic tissues, accompanied with the increased protein expression of caspases 8 and 9, activated caspase-3, and cytochrome c in a dose-dependent manner. However, these protein expression profiles were not affected in eutopic endometrium. Our findings suggest that KTC could inhibit the growth of ectopic endometrial tissue through upregulating the level of TNF-α and its downstream signaling, including caspases and cytochrome c.

Preliminary study of quercetin affecting the hypothalamic-pituitary-gonadal axis on rat endometriosis model.

In this study, the endometriosis rats model was randomly divided into 6 groups: model control group, ovariectomized group, Gestrinone group, and quercetin high/medium/low dose group. Rats were killed after 3 weeks of administration. The expression levels of serum FSH and LH were detected by ELISA. The localizations and quantities of ERα, ERß, and PR were detected by immunohistochemistry and western blot. The results showed that the mechanism of quercetin inhibiting the growth of ectopic endometrium on rat endometriosis model may be through the decreasing of serum FSH and LH levels and then reducing local estrogen content to make the ectopic endometrium atrophy. Quercetin can decrease the expression of ERα, ERß, and PR in hypothalamus, pituitary, and endometrium, thereby inhibiting estrogen and progesterone binding to their receptors to play the role of antiestrogen and progesterone.

[Effects of Caulis Sargentodoxae Granule on expressions of vascular endothelial growth factor and its receptor-2 in rats with endometriosis].

OBJECTIVE: To investigate the effects of Caulis Sargentodoxae Granule (CSG), a compound traditional Chinese herbal medicine for treating endometriosis, on expressions of vascular endothelial growth factor (VEGF) and its receptor-2 fetal liver kinase-1 (Flk-1) in rats with endometriosis. METHODS: A rat model of endometriosis was established by autotransplant of endometria. Forty-eight endometriosis rats were randomly divided into 6 groups: untreated group, castrate group, mifepristone group (5 mg/kg) and low- (20 g/kg), medium- (40 g/kg) and high-dose (80 g/kg) CSG group. After 21-day consecutive treatment, the expressions of VEGF and its receptor Flk-1 in ectopic endometria were detected by immunohistochemical method, and the volume of the ectopic endometria was also measured by using electronic digital calipers. RESULTS: Compared with the untreated group, the endometrial volumes in the treated groups were significantly decreased (P<0.05) except for the low-dose CSG group. Histopathologic observation showed that the epithelial layer of ectopic endometrium was less and the thickness of epithelial layer was thinner than those in the untreated group (P<0.01), but the low-dose CSG group had few changes. The VEGF and its receptor Flk-1 expressed abundantly in the cytoplasm of the glandular epithelium and vascular endothelial cell in all the groups. Compared with the untreated group, the expressions of VEGF and Flk-1 of ectopic endometrium in the CSG-treated groups, the mifepristone group and the castrate group were significantly decreased (P<0.05, P<0.01). CONCLUSION: CSG can treat the endometriosis in rats, and its action may be associated with decreasing the expressions of VEGF and its receptor Flk-1 in ectopic endometrium.