Premature Ovarian Failure Mice Research Articles

Transplantation of umbilical cord-derived mesenchymal stem cells on a collagen scaffold improves ovarian function in a premature ovarian failure model of mice.

Premature ovarian failure (POF) is a refractory disease; one of the most important goals of treatment is to improve fertility. In the study, collagen scaffold loaded with human umbilical cord-derived mesenchymal stem cells (collagen/UC-MSCs) transplantation in POF mice preserved ovarian function, as supported by increased estrogen (E2) and anti-Mullerian hormone (AMH) levels, increased ovarian volume, and an increased number of antral follicles. Immunohistochemistry results of Ki67 indicated transplantation of collagen/UC-MSCs promoted granulosa cell proliferation, which is crucial to oocyte maturation and follicular development. Additionally, transplantation of collagen/UC-MSCs significantly promoted ovarian angiogenesis with the increased expression of CD31. In general, collagen/UC-MSCs transplantation probably is an effective therapeutic strategy of POF.

Thymopentin promotes ovarian angiogenesis in mice by activating N6-methyladenosine (m6A) RNA modification of key factors in the Notch/Tie1 pathway

Abstract: BACKGROUND: Premature ovarian failure (POF) is a typical condition of pathological ovarian ageing. Injuries and atrophies of blood vessels in the ovaries can lead to ovarian insufficiency, but the underlying mechanism remains unclear. AIMS: This study investigated the epigenetic mechanism by which thymopentin (TP-5) activated ovarian angiogenesis to achieve its effects on POF. MATERIALS AND METHODS: The qPCR, western blot and immunofluorescence staining were used to detecte the expression levels of gene. The hematoxylin and eosin stainingwas used to histopathological examination. The RNA-Seq sequencing was used to transcriptome detection. RESULTS: First, pathological examination revealed that TP-5 significantly increased ovary weight in the cyclophosphamide-induced POF mouse model. The proportion of atretic follicles was reduced, and the level of E2 in the peripheral blood was elevated. Meanwhile, immunofluorescence staining showed that TP-5 increased protein expression of CD31 and Tie1 in the ovarian tissues of mice with POF, suggesting that TP-5 could induce ovarian angiogenesis. RNA-Seq sequencing results suggested that TP-5 could induce the high level of Notch/Tie2 pathway expression that is associated with angiogenesis in ovarian tissues. qPCR and western blotting showed significantly increased expression of METTL3, a methyltransferase that catalyses the formation of N6-methyladenosine (m6A) in RNA. Meanwhile, the results of the dot blot suggested that the overall RNA methylation level was significantly higher in the ovarian tissues of mice in the TP-5 group than in the control group. Finally, RIP-PCR analysis showed that specific sites in the 3'-untranslated region (3'UTR) of the mRNA of key factors in the Notch/Tie2 pathway were hypermethylated in the ovaries of mice in the TP-5 treatment group. CONCLUSION: Therefore, this study demonstrated that TP-5 exerted its therapeutic effect on POF by stimulating angiogenesis. The mechanism of action is the promotion of the expression of RNA m6A methyltransferases by TP5, which increases the overall RNA m6A methylation level in mouse ovarian tissue. Specifically, the methylation ofspecific m6A sites in the 3'UTR of the RNA of key factors in the Notch/Tie2 pathway increased, which enhanced their stability and expression levels, leading to the induction of angiogenesis.

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

AimsElectro-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 methodsForty-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 findingsWe 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. SignificanceThe effects of EA on the PI3K/AKT/mTOR signaling pathway may represent one of the mechanisms involved in attenuating the mice POF.

Mitigation of premature ovarian failure by over-expression of lentivirus vector-mediated Wilms tumor-suppressor gene

Purpose: To investigate the effect of WT1(Wilms tumor-suppressor gene) overexpression on premature ovarian failure (POF)-mediated ovarian dysfunction. Methods: Three mice groups were used: control group (untreated mice), POF group (mice sterilized by intravenous injection of cyclophosphamide and busulfan), and POF-LV(lentiviral vector)􀀀GFP(green fluorescent protein)􀀀WT1 group (POF mice given intra-ovarian microinjection of LV􀀀GFP􀀀WT1, a WT1􀀀overexpressing lentiviral vector, one week after sterilization). Real time-PCR was employed to analyze in vitro WT1 overexpression levels. Overall ovarian function was measured by hormonal assay, H & E staining, and immuno-histochemical techniques. Results: Overexpression of WT1 in mice models of POF alleviated ovarian granulosa cell (GC) damage, increased ovary weight, and significantly increased follicular number (p < 0.05). Radioimmunoassays revealed reduction in plasma estradiol (E2) and follicle-stimulating hormone (FSH, p < 0.05). However, results from immune-histochemical assays showed reduced Bax expression levels, and increased expression of Bcl-2 in WTI-overexpression mice, relative to POF mice. Conclusion: Overexpression of WT1 may stimulate repair of ovarian tissue while improving endocrine function by inhibiting ovarian cell apoptosis signaling pathway in POF mice. Keywords: Premature ovarian failure, Wilms tumor suppressor gene, Chemotherapy, Apoptosis, Estradiol

Restoring Ovarian Function With Human Placenta-Derived Mesenchymal Stem Cells in Autoimmune-Induced Premature Ovarian Failure Mice Mediated by Treg Cells and Associated Cytokines.

Regulatory T (Treg) cells play a key role in the regulation of autoimmunity and transplantation. Human placenta-derived mesenchymal stem cell (hPMSC) transplantation has a potential to restore ovarian dysfunction associated with premature ovarian failure (POF), while the exact function of the Treg cells in the transplantation still needs to be further investigated. In this study, hPMSCs were intravenously injected into POF mice following zona pellucida glycoprotein 3 (pZP3) treatment. Ovarian function was measured by analyzing estrous cycle, folliculogenesis, and hormone secretion, also, with the detection of apoptotic granular cells (GCs) in ovarian tissues. To determine whether immune response is involved in the regulation of ovarian function change, the population of Treg cell populations and expression of associated cytokines, for example, transforming growth factor β (TGF-β) and interferon γ (IFN-γ) were measured. After hPMSCs transplantation, the injured ovarian function is significantly improved. Also, the pZP3-treatment-induced apoptotic GCs were significantly decreased as compared with the POF mice. The transplantation of hPMSCs significantly increased the population of Treg cells which was inhibited by pZP3 treatment. The decrease in TGF-β and increase in IFN-γ in serum caused by pZP3 treatment have been reversed following hPMSCs transplantation. These findings strongly suggest that the recovery of ovarian function in POF mice is mediated via the regulation of Treg cells and production of associated cytokines following hPMSCs transplantation.

Human chorionic plate-derived mesenchymal stem cells transplantation restores ovarian function in a chemotherapy-induced mouse model of premature ovarian failure

BackgroundPrevious studies have reported that transplantation of mesenchymal stem cells (MSCs) from many human tissues could ameliorate ovarian dysfunction. However, no study has revealed the therapeutic efficiency of MSCs derived from the chorionic plate (CP-MSCs) for premature ovarian failure (POF).MethodsWe investigated the restorative effects of CP-MSCs on cyclophosphamide (CTX)-induced POF. The POF mouse models were established via intraperitoneal injection of 50 mg/kg CTX into female mice for 15 consecutive days. After that, CP-MSCs were intravenously transplanted into the mice once a week for 4 weeks. The serum estradiol (E2) and follicle-stimulating hormone (FSH) levels in the mouse models were detected using enzyme-linked immunosorbent assay (ELISA) before and after treatment. Ovarian function was evaluated through counting the follicles, estrous cycles, and oocytes.ResultsCP-MSC transplantation restored the serum hormone level and ovarian function of the mice in the mouse model of POF induced by CTX. The levels of serum E2 and FSH in the POF model group was 232.33 ± 17.16 pg/mL and 4.48 ± 0.29 mIU/mL, respectively, after 6 weeks of treatment, which were similar to the values in the wild-type (WT) group. The superovulation demonstrated that ovarian function was significantly improved compared with nontreated POF model mice. The CP-MSC transplantation could restore CTX-induced ovarian dysfunction.ConclusionsOur results offer a potential application for human CP-MSCs in POF treatment.

RETRACTED ARTICLE: hPMSC transplantation restoring ovarian function in premature ovarian failure mice is associated with change of Th17/Tc17 and Th17/Treg cell ratios through the PI3K/Akt signal pathway

BackgroundHuman placenta-derived mesenchymal stem cell (hPMSC) transplantation has been demonstrated to be an effective way of recovering ovarian function in mice with autoimmune induced premature ovarian failure (POF). But the exact mechanism remains unclear. The goal of the present study is to investigate the role of immune factors (T-helper 17 (Th17), cytotoxic T (Tc17) and regulatory T (Treg) cells) in the recovery of ovarian function and whether the phosphatidylinositol 3-kinase (PI3K)/Akt signal pathway is involved in the regulation.MethodsThe inhibitor of PI3K/Akt was administered to observe its effect on ovarian function recovery and immune regulation. Serum levels of estradiol (E2), follicle stimulation hormone (FSH), luteinizing hormone (LH) and anti-Müllerian hormone (AMH)) and anti-Zona pellucida antibody (AZPAb) were measured by ELISA to evaluate ovarian function. The morphological changes of ovaries were observed by HE staining. Apoptosis of granular cells (GCs) was determined by detecting the expression of capase-3. Expression of p-Akt protein was detected by immunohistochemistry and western blot assay in ovarian tissues. The MTT assay was performed to assess GC proliferation. GC apoptosis was performed using flow cytometry analysis. Percentages of Th17, Tc17 and Treg cells were detected by flow cytometry. Expression of interleukin (IL)-17 in serum was measured by ELISA.ResultsLY294002 administration decreased serum levels of E2 and AMH, while the levels of FSH, LH and AZPAb in serum were increased compared with mice in the hPMSC transplantation group. The ovarian morphology presented as atrophy and fibrosis, with functional follicles exhausted. The expression of p-Akt in ovarian tissue was significantly decreased. Also, LY294002 administration significantly decreased proliferation and increased cell apoptosis in GCs, and for immune factors the ratios of Th17/Tc17 and Th17/Treg cells were significantly increased, as well as the serum levels of IL-17.ConclusionsOur data suggest that the PI3K/Akt signal pathway is involved in the recovery of ovarian function by changing the ratios of Th17/ Tc17 and Th17/Treg cells in POF mice following hPMSC transplantation.

Curcumin exerts a protective effect against premature ovarian failure in mice.

This study was designed to investigate the protective effect of curcumin against d-galactose (d-gal)-induced premature ovarian failure (POF) in mice. A mouse POF model was induced by subcutaneous injection of d-gal (200 mg/kg/day) daily for 42 days. Mice in the curcumin group received both d-gal treatment and intraperitoneal injection of curcumin (100 mg/kg/day) for 42 days. Ovarian function, oxidative stress and apoptosis were evaluated. The P, E2 and SOD levels were higher, and the FSH, LH and MDA levels were significantly lower in the curcumin group than those in the d-gal group. The proportion of primordial follicles was also significantly higher in the curcumin group than that in the d-gal group. In addition, curcumin treatment after d-gal administration resulted in significantly lower Sod2, Cat, 8-OhdG, 4-HNE, NTY and senescence-associated protein P16 expression levels, higher Amh expression levels and less apoptosis in granulosa cells than was observed in the d-gal group. Moreover, the p-Akt, Nrf2 and HO-1 protein expression levels were significantly higher and the apoptosis-related cleaved caspase-3 and -9 protein expression levels were markedly lower in the curcumin group than in the d-gal group. In conclusion, curcumin effectively inhibited d-gal-induced oxidative stress, apoptosis and ovarian injury via a mechanism involving the Nrf2/HO-1 and PI3K/Akt signaling pathways, suggesting that curcumin is a potential protective agent against POF.

RETRACTED ARTICLE: Effects of hPMSCs on granulosa cell apoptosis and AMH expression and their role in the restoration of ovary function in premature ovarian failure mice

BackgroundThis study was performed to determine the effects of human placenta mesenchymal stem cell (hPMSC) transplantation on granulosa cell apoptosis and anti-Müllerian hormone (AMH) and follicle-stimulating hormone receptor (FSHR) expression in autoimmune drug-induced premature ovarian failure (POF) mice. The aim of this research is to investigate the mechanisms of hPMSCs on ovarian reserve capacity.MethodsThe POF mice model was established by injection of zona pellucida 3 peptide (pZP3). hPMSC transplantation was conducted by intravenous injection into mice following pZP3 treatment. The follicle number was examined by histopathology. The serum levels of FSH, LH, E2, AMH and anti-zona pellucida antibody (AzpAb) were measured by enzyme-linked immunosorbent assay. AMH and FSHR expression in the ovary was analyzed by immunohistochemistry and western blot analysis. Granulosa cell apoptosis of the ovaries was examined by In Situ Cell Death Detection Kit. Granulosa cells were isolated and treated with SiAmh interference and hPMSC supernatant to observe the effects of AMH expression on granulosa cell apoptosis in vitro.ResultsThe results showed that hPMSC transplantation can significantly recover the estrus cycle in the POF group. Morphological staining showed that the basal follicles and sinus follicles after hPMSC transplantation were higher in POF mice than in those without treatment, and the follicle number was significantly decreased with atresia. The serum levels of FSH, LH and AzpAb in the hPMSC transplantation group were reduced considerably, but the E2 and AMH levels were significantly increased. After hPMSC transplantation, the AMH and FSHR expression in ovarian tissue was significantly higher than in the POF group as determined by immunochemistry and western blot analysis. The FSHR expression was shown in granulosa cells only, and FSHR expression increases with AMH expressed in the ovary; granulosa cell apoptosis was decreased following hPMSC transplantation. The same results were observed from the in-vitro study.ConclusionshPMSC transplantation can significantly improve the serum levels of high gonadotropin and low estrogen of POF mice, promote follicular development, inhibit excessive follicular atresia and granulosa cell apoptosis, and improve the ovarian reserve capacity. The mechanism may be achieved by increasing the expression of AMH and FSHR in ovaries.

Therapeutic potential of BMSCs for premature ovarian failure in mice

To explore the therapeutic effects of bone marrow-derived mesenchymal stem cells (BMSCs) on premature ovarian failure (POF) in mice induced by cyclophosphamide (CTX) and the possible mechanisms. Methods: Mouse BMSCs were identified through detection of cell surface markers by flow cytometry. The model of mouse POF was induced by intraperitoneal injection of CTX at a dose of 50 mg/kg, once daily for 15 days. BMSCs were transplanted into POF mice at 2×106 cells/mouse by tail veil. The ovarian tissues were collected for HE staining at 7 days after transplantation to observe the changes of ovarian structure and real-time PCR was performed to detect the folliculogenesis gene expression. Results: BMSCs showed positive expression of CD29 and CD90 while low expression for endothelial and hematopoietic cell markers CD31 and CD34. The numbers of primodial follicle, primary follicle, secondary follicle and antral follicle were significantly decreased, but the numbers of atretic follicle were significantly increased in CTX induced-POF mice (P<0.05). BMSCs transplantation effectively repaired the structure of damaged ovary. The significant reduction of atretic follicle and significant increase of antral follicle and secondary follicle were observed in ovaries of BMSCs-treated mouse(P<0.05). BMSCs-transplanted mouse ovaries showed the increased mRNA expression levels of Nano3, Nobox, and Lhx8 (P<0.05). Conclusion: BMSCs could effectively repair ovarian structure and promote follicle development in CTX-induced POF mouse.

Establishment of a Mouse Model of Premature Ovarian Failure Using Consecutive Superovulation

Background/Aims: This study investigated the effect of consecutive superovulation on the ovaries and established a premature ovarian failure (POF) model in mice. Methods: The mouse POF model was induced by 5-15 consecutive superovulation treatments with pregnant mare serum gonadotropin (PMSG), human chorionic gonadotropin (HCG) and prostaglandin F2α (PGF2α). Normal adult mice were compared with mice displaying natural ovarian aging. The following serum biochemical parameters were measured: including follicle-stimulating hormone (FSH), luteinizing hormone (LH), progesterone (P), estradiol (E2), inhibin B (INH B), malondialdehyde (MDA), total superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels. Follicles were counted using H&E staining. Levels of 8-hydroxyguanosine (8-OhdG), 4-hydroxynonenal (4-HNE), nitrotyrosine (NTY), anti-Mullerian hormone (AMH) and CDKN2A/ p16 (p16) were detected using immunohistochemical staining. Reactive oxygen species (ROS) levels were measured using dihydroethidium (DHE) staining. Cell apoptosis was detected using an in situ TUNEL fluorescence staining assay. Levels of proteins involved in ROS-related pathways and the p16 protein were detected using Western blotting. Sod1, Sod2 and Sod3 mRNA levels were detected using quantitative polymerase chain reaction (Q-PCR). Oocyte quality was evaluated using in vitro fertilization (IVF) and zygote culture. Results: Consecutive superovulation groups presented lower P, E2, SOD, GSH-Px and INH B levels, significantly higher FSH, LH, MDA and ROS levels, and significantly fewer primordial follicles compared with the control group. Consecutive superovulation groups presented significantly increased levels of Sod2, 8-OhdG, 4-HNE, NTY, significantly increased levels of the SIRT1 and FOXO1 proteins, significantly increased levels of the senescence-associated protein p16, as well as decreased AMH, Sod1 and Sod3 levels and increased granulosa cell apoptosis compared with the control group. Conclusion: Consecutive superovulation significantly decreased ovarian function and oocyte quality and increased oxidative stress and apoptosis in the ovary via a mechanism involving the p16 and SIRT1/FOXO1 signaling pathways. These findings suggest that consecutive superovulation may be used to establish a mouse model of ovarian aging.

PROTECTIVE EFFECTS OF CISTANCHES HERBA AQUEOUS EXTRACT ON CISPLATIN-INDUCED PREMATURE OVARIAN FAILURE IN MICE

Background: Chemotherapeutic treatment of premenopausal women has been linked to premature ovarian failure (POF). Cistanches Herba (CH) is a commonly used male impotence and female infertility treatment in China; however, whether CH protects ovaries from chemotherapeutic drug-induced POF remains unclear. In this study, we investigated the protective effects of CH in a mouse model of chemotherapeutic drug-induced POF.&#x0D; Materials and Methods: We administered low- and high-concentration CH to cisplatin-induced POF mice for 2 weeks and determined body and ovarian weights, as well as serum follicle-stimulating hormone (FSH) and estradiol concentrations, to evaluate ovarian function. In addition, we evaluated the protective mechanisms of CH by detecting the levels of apoptosis-related proteins and evaluating markers of mitochondrial function.&#x0D; Results: In POF mice, we observed reduced body and ovarian weights; elevated serum FSH and attenuated estradiol concentrations; apoptosis of ovarian granulosa with concomitant changes in apoptosis-related proteins (including caspase-3, poly adenosine diphosphate-ribose polymerase, Bcl-2, and Bax); and mitochondrial dysfunction, such as a reduction in mitochondrial numbers, destruction of ultrastructural morphology, decrease in ATPase activity, and decreases in mitochondrial membrane potential and mitofusin-2 (a mitochondria dynamin-like GTPase). Significantly, CH reversed, to an extent, functional and morphologic injuries and ovarian tissue apoptosis by up-regulating the level of Mfn2 and the ratio of Bcl-2/Bax. Furthermore, CH reduced cisplatin-induced mitochondrial dysfunction in ovarian tissues.&#x0D; Conclusion: The present findings showed that CH inhibited cisplatin-induced POF through interactions between Mfn2 and Bcl-2/Bax proteins and, possibly, by up-regulation of Mfn2 expression. Ultimately, CH protects ovarian tissues from cisplatin-induced apoptosis.

Efficacy of Bushenjianpi prescription on autoimmune premature ovarian failure in mice

ObjectiveTo assess the efficacy of Bushenjianpi prescription (BSJPP), a formula from Traditional Chinese Medicine, on a mouse model of autoimmune premature ovarian failure (POF) induced by mouse zona pellucida (ZP3) and to investigate the mechanisms underlying the action. MethodsAfter randomization, POF was induced in the model mice by immunization with ZP3. One week later, mice received low (8.1 mg/kg), moderate (16.2 mg/kg) and high (32.4 mg/kg) doses of BSJPP by gastrogavage once daily for 90 days. Premarin (0.03 mg/kg) served as the positive group. Serum samples were collected 1 week after the last dose and stored at −20 °C for analysis. After cervical dislocation, the uterus and ovaries were collected aseptically for evaluation by histological assessment, scanning electron microscopy, immunohistochemical staining, and Western blot and reverse transcription-polymerase chain reaction analyses. ResultsSerum E2 levels in POF model mice were decreased, whereas follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels were dramatically increased. Serum levels of E2, LH and FSH were reduced in POF model mice treated with BSJPP (moderate and high doses) and premarin. Bmp15 and connexin 43 (Cx43) were repressed in autoimmune POF model mice, whereas high expression was observed in control mice and those treated with BSJPP (moderate and high doses) and premarin. ConclusionBSJPP is effective in treating ZP3-induced POF in mice and the increase in the expression of Bmp15 and Cx43 may be implicated in the mechanism underpinning the action.

Role of SDF-1/CXCR4 and cytokines in the development of ovary injury in chemotherapy drug induced premature ovarian failure mice

ObjectiveTo explore the mechanism of chemotherapy drug induced ovarian injury in premature ovarian failure (POF) mice. MethodsC57BL/6 mice were treated with Cyclophosphamide and Busulfan by intraperitoneal injection. One week after treatment, the estrous cycles, folliculogenesis, ovarian endocrine function and ovarian histopathological changes were evaluated the ovarian function. The serum levels of cytokines, follicle stimulating hormone (FSH) and estradiol (E2) were measured by enzyme-linked immunosorbent assay (ELISA). The protein levels of SDF-1/CXCR4 and FSHR in ovary were evaluated by immunohistochemistry and Western blot analysis. The ovarian cells apoptosis was measured by TUNEL Assay. ResultsThe ovaries from POF mice show the evidence of reduced ovarian function such as irregular estrous cycles, stromal hyperplasia, decreased follicle numbers, atresia follicles and less granular cell layer as well as corpora luteum. The lower levels of E2 and higher levels of FSH in serum characterize the ovarian injury; a great number of granular apoptotic cells were observed in the POF mice; the serum concentrations of pro-inflammatory cytokines of IL-6, IL-8 and TNF-α level were increased but anti-inflammatory cytokine of IL-10 was decreased. SDF-1/CXCR4 and FSHR expressed in ovaries were detected in the cytoplasm of preantral and antral follicles; the expression of SDF-1/CXCR4 was increased and FSHR was decreased in POF mice. ConclusionOur data suggest that the inflammatory regulation, SDF-1/CXCR4 and cellular apoptosis in ovarian tissues are involved in the development of ovarian injury of POF. These data provide useful information to develop new therapeutic approach to treat POF disorders in the future.

Ginsenoside Rg1 improves fertility and reduces ovarian pathological damages in premature ovarian failure model of mice.

This study aims to investigate the effect as well as mechanism of ginsenoside Rg1 (Rg1) on premature ovarian failure (POF) induced by d-galactose (d-gal) in mice. C57BL/6 female mice were divided into four groups randomly, which were the saline group, the d-gal group, the d-gal + Rg1 group, and the Rg1 group. Body weight was recorded. Overall ovarian function including estrous cycles, sex hormone secretion, ovarian follicle development, and ovarian morphology was analyzed by H&E staining and ELISA. Effect of Rg1 on aging was determined by analyzing the activities of oxidation-associated biomarkers, pro-inflammatory cytokine secretion, expression of senescence-associated proteins, and fertility. Compared with the d-gal group, in Rg1 + d-gal group, body weight was increased significantly, estrous cycle block was released, and fertility and the morphology of ovaries were restored. And, Rg1 treatment after d-gal administration significantly reduced senescence-associated protein expression, increased the activity of total superoxide dismutase and glutathione peroxidase from bovine erythrocyte, and induced higher follicle stimulating hormone receptor protein expression. Additionally, the expression levels of malondialdehyde, interleukin-1β, tumor necrosis factor-α, and interleukin-6 were significantly decreased. Together, Rg1 improves mouse fertility and reduces ovarian pathological damage in d-gal-induced POF model possibly through enhancing anti-inflammatory and antioxidant capacities and reducing expression of senescence signal pathway proteins. Impact statement Ginsenoside Rg1 (Rg1) is a kind of natural estrogen and it has antioxidation and antiaging effects. However, whether Rg1 has effects on premature ovarian failure (POF) is still not clear. In this study, aging model induced by d-galactose was used to mimic POF. The effect and possible mechanism of Rg1 on ovary aging was investigated. We found that Rg1 treatment up-regulated the expression of follicle stimulating hormone receptor and down-regulated senescence-associated protein expression in granule cells of POF mice. Particularly, Rg1 improved fertility ability and reduced ovarian pathological damages by its antioxidative and anti-inflammation capacity. Thus, Rg1 enhances the antiaging ability of ovary and fertility ability of POF mice through enhancing the anti-inflammatory and antioxidant capacities of ovary.

Study of the reparative effects of menstrual-derived stem cells on premature ovarian failure in mice

BackgroundYoung female patients who receive chemotherapy frequently face premature ovarian failure (POF). The therapeutic potential of stem cells in these patients has been explored in stem cells derived from different sources. However, many of these types of stem cells are either difficult to obtain or obtaining them involves invasive procedures. Here, we show that menstrual-derived stem cells (MenSCs) are easy to access and exhibit mesenchymal stem cell-like properties. MenSCs are therefore a novel source of stem cells that can be used for tissue repair. The aim of this study was to explore the reparative capacity and the mechanism underlying the activities of MenSCs.MethodsPOF mouse models were established by 7 consecutive days of intraperitoneal injection of cisplatin, and then MenSCs or MenSC-derived conditioned media (CM) were infused via the tail vein. The ovaries were excised after either 7 or 21 days of treatment and the follicles were counted and categorized. Apoptosis of granulosa cells was observed by terminal deoxynucleotidyl transferase mediated dUTP nick end labelling staining. Ovarian function was evaluated by monitoring serum sex hormone levels. Furthermore, MenSC tracking, Q-PCR, and small interfering RNA transfection were used to reveal the inner mechanism of repair.ResultsMenSC transplantation could improve the ovarian microenvironment by reducing apoptosis in granulosa cells and the fibrosis of ovarian interstitium, which contributes to increase the follicular numbers and return sex hormone levels to normal values. Meanwhile, the transplanted MenSCs directively migrate to ovarian interstitium to play a role in repair rather than differentiate to oocytes directly. Additionally, MenSCs and CM derived from these cells exerted protective effects on damaged ovaries partially by secreting FGF2.ConclusionMenSCs repair ovarian injury, improve ovarian function, and stimulate regeneration, suggesting that transplantation of MenSCs may provide an effective and novel method for treating POF.

Effect and mechanism of Bushen Huoxue recipe on ovarian reserve in mice with premature ovarian failure.

The aim of the present study was to explore the effect and mechanism of Bushen Huoxue recipe (BHR) on ovarian reserve in mice with premature ovarian failure (POF). Mice were divided into 3 groups: normal group, model group and BHR group. Intraperitoneal injection of cyclophosphamide was performed to create the POF model. Primordial follicular (PDF) number, ovarian wet weight, ovarian index, and estrous cycle were analyzed to evaluate the effect of BHR on POF. Meanwhile, the mRNA and protein level of Mouse Vasa Homologue (MVH) in the bone marrow, peripheral blood and ovary were detected, to explore the underlying mechanism of the treatment efficacy of BHR on ovarian reserve. By the time of BHR treatment for 28 days, BHR increased the PDF number and shortened the estrous cycle of POF mice. BHR also decreased the mRNA level of MVH in the bone marrow, and increased mRNA and protein level of MVH in the ovary of POF mice. Our results demonstrated a treatment efficacy of BHR on POF mice, and revealed that BHR might repair the dysfunction of germline stem cells in the bone marrow, and thus to improve the ovarian reserve and enhance the ovarian function of POF mice through neo-oogenesis.

Telocytes as potential targets in a cyclophosphamide-induced animal model of premature ovarian failure.

Premature ovarian failure (POF) refers to the presence of ovarian atrophic permanent amenorrhea in women under the age of 40. The pathogenesis of POF remains to be fully elucidated. Telocytes are a group of specialized cells with a small cell volume and very long cytoplasmic prolongations with dichotomous branching. Previous studies have indicated that telocytes function to support the trachea and serve as stem cell niches. Although it has been confirmed that telocytes are present in numerous organs in mammals, it remains to be determined whether they are present in ovarian tissues and whether they are involved in the development of POF. The present study used a cyclophosphamide-induced mouse model of POF. Hematoxylin and eosin staining and an enzyme-linked immunosorbent assay revealed that cyclophosphamide induced edema and apoptosis of ovarian stromal and granulosa cells and increased atretic follicles. In addition, cyclophosphamide induced abnormal peripheral blood FSH and E2 levels in mice. Transmission electron microscopy revealed a small number of telocyte-like cell structures in the ovarian stroma of wild-type mice. In addition, flow cytometry and immunohistochemical staining results suggested that the number of cluster of differentiation (CD)34/platelet-derived growth factor receptor (PDGFR)α, CD34/PDGFRβ and CD34/vimentin double-positive cells in the ovaries of POF mice was significantly decreased compared with wild-type mice. In conclusion, mouse ovarian tissues appear to contain telocytes, and cyclophosphamide treatment significantly reduced the number of ovarian telocytes. Therefore, telocytes may serve as a potential novel marker of POF induced by cyclophosphamide.

Growth hormone treatment of premature ovarian failure in a mouse model via stimulation of the Notch-1 signaling pathway.

Premature ovarian failure (POF) is a condition affecting 1% of women in the general population, causing amenorrhea, hypergonadotropism and hypoestrogenism before the age of 40. Currently, POF cannot be reversed and, although treatments are available, there is an urgent need for improved treatment strategies. Growth hormone (GH) is a pleiotropic hormone that affects a broad spectrum of physiological functions, from carbohydrate and lipid metabolism to the immune response. GH has previously been used to treat POF in non-transgenic preclinical trials, but the biochemical mechanism underlying these effects are unclear. In the present study, a mouse model of POF was generated using cyclophosphamide. Treatment of POF mice with recombinant mouse growth hormone (rmGH) was revealed to markedly reduce POF histopathology in ovarian tissue, relieve ovarian granulosa cell injury, reduce the number of atretic follicles and significantly increase the number of mature oocytes. Furthermore, an enzyme-linked immunosorbent assay revealed that plasma estradiol levels increased and plasma follicle stimulating hormone levels decreased with time in a group of mice treated with a medium dose of rmGH (0.8 mg/kg) when compared with the POF model group (P<0.05). In addition, reverse transcription-quantitative polymerase chain reaction and immunohistochemical analysis demonstrated elevated levels of Notch-1 signaling pathway factors (Notch1, CBF1, and HES1) in wild-type mice and those treated with medium and high doses of rmGH, but not in those treated with low doses of rmGH. In conclusion, GH may promote ovarian tissue repair, estrogen release and oocyte maturation via activation of the Notch-1 signaling pathway in ovarian tissue.

Transplantation of ovarian granulosa‑like cells derived from human induced pluripotent stem cells for the treatment of murine premature ovarian failure.

Premature ovarian failure (POF) is a common cause of female infertility, for which there are currently no ideal treatments or medications. Furthermore, apoptosis of ovarian granulosa cells (OGCs) is an important mechanism underlying the decline in ovarian reserve and function. In the present study, several cellular growth factors and hormones were used to induce the differentiation of human induced pluripotent stem cells (iPSCs) into ovarian granulosa-like cells (OGLCs) in vitro. Immunohistochemical staining demonstrated that OGLCs derived from iPSCs strongly expressed granulosa cell markers, including anti-Müllerian hormone, inhibin α, inhibin β and follicle-stimulating hormone receptor, but did not express stem cell markers, including octamer-binding transcription factor 4, SRY (sex determining region Y)-box 2, Nanog and stage-specific embryonic antigen-4 12 days post-induction. In addition, a mouse model of POF was generated by cyclophosphamide treatment. Subsequently, iPSC-derived OGLCs were transplanted into the POF mice (OGLCs-iPSCs-POF group) in vivo. Results indicated that, compared with the control group (POF mice treated with phosphate-buffered saline), the growth state of OGLCs was markedly improved, and mature follicles could be detected in the ovarian tissue of the OGLCs-iPSCs-POF group. Immunohistochemical staining demonstrated that iPSC-derived OGLCs transplanted into POF mice not only exhibited substantial growth in murine ovarian tissues, but also strongly expressed OGC markers. Furthermore, enzyme-linked immunosorbent assays indicated that the levels of the hormone estradiol in peripheral blood samples were significantly enhanced following transplantation of iPSC-derived OGLCs into POF mice. Furthermore, ovarian tissue weight was significantly higher in the OGLCs-iPSCs-POF group compared with in the control group, and the number of atretic follicles in OGLCs-iPSCs-POF mice was significantly reduced, as compared with in the control mice. These results suggest that OGLCs derived from human iPSCs may not only effectively enhance OGC growth and repair damaged ovarian tissue, but may also maintain the ovarian tissue niche, promoting follicular development and maturation in a mouse model of POF.