Kisspeptin regulates the proliferation and apoptosis of ovary granulosa cells in polycystic ovary syndrome by modulating the PI3K/AKT/ERK signalling pathway.

BACKGROUND: The development of polycystic ovary syndrome (PCOS) is closely correlated with apoptosis and oxidative stress in ovarian granulosa cells. Kisspeptin plays an important role in reproductive organ function. This study aimed to explore the role of kisspeptin in PCOS and oxidative stress-triggered apoptosis of ovarian granular cells. METHODS: A PCOS rat model was established by injecting dehydroepiandrosterone (DHEA) and feeding the rats a high-fat diet. The RNA and protein levels of kisspeptin were analysed by quantitative PCR, western blotting, and histological staining. Tissue damage was evaluated using haematoxylin and eosin (H&E) staining. The viability and proliferation of human granulosa cell KGN were measured using the cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assays. Cell cycle and apoptosis were analysed by flow cytometry. Oxidative stress was analysed by measuring reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), and catalase (CAT) levels. RESULTS: Kisspeptin was downregulated in the ovarian granulosa cells of PCOS rats compared to those of control rats. Kisspeptin overexpression enhanced KGN cell proliferation and inhibited apoptosis. ROS generation was suppressed by kisspeptin, along with decreased levels of MDA and increased levels of the antioxidants GSH, SOD, and CAT. Kisspeptin activates PI3K/AKT and ERK signalling, and inactivation of ERK1/2 suppresses the protective role of kisspeptin in ovarian granulosa cells. CONCLUSION: Kisspeptin improves proliferation and alleviates apoptosis and oxidative stress in ovarian granulosa cells by activating PI3K/AKT and ERK signalling.

[Long non-coding RNA in spermatogenesis regulation and male infertility: Progress in research].

Long non-coding RNA (lncRNA) is an RNA molecule transcribed by RNA polymerase II, longer than 200 nt, and not translated into proteins. During gonadal development and spermatogenesis, lncRNAs are involved in epigenetic mechanisms, including DNA methylation, chromatin remodeling, and histone tail modification, which play important regulatory roles at the transcriptional or post-transcriptional level. Epigenomics including lncRNA is considered to be the second dimension of DNA sequence that can be adapted to environmental factors to specifically regulate gene expressions in some cells. Based on the functional action mechanism of lncRNAs, we reviewed the advances in the studies of lncRNAs in the direction of spermatogenesis and male infertility and analyzed the potential of lncRNAs as a biomarker of male infertility. The potential application of lncRNA in the treatment of male infertility diseases can be further explored based on the lncRNA target, RNA interference, competitive binding closed target and structural disruption of lncRNAs.

[Regulation of the ubiquitin-proteasome system in spermiogenesis].

The ubiquitin-proteasome system (UPS) is an adenosine triphosphate (ATP)-dependent enzymatic machinery that targets substrate proteins for degradation by the 26S proteasome by tagging them with an isopeptide chain composed of covalently linked molecules of ubiquitin, a small chaperone protein. UPS is the main pathway of protein degradation in eukaryotic cells, and plays an important role in spermatogenesis. The dysfunction of various ubiquitin systems results in impaired sperm development with abnormal morphology and function, which is highly associated with male infertility. This review focuses on the roles of UPS in histone-to-protamine exchange, acrosome formation, sperm mitochondrial degradation and regulation of sperm function and quality.

[L-carnitine improves sperm acrosin activity in male infertility patients].

OBJECTIVE: To evaluate the effect of L-carnitine (LC) on low sperm acrosin activity in infertile man. METHODS: A total of 240 male infertility patients with low sperm acrosin activity were randomly assigned to an LC group (n = 180) and a control group (n = 60) to be treated with LC (1g, tid) and vitamin E (VE) capsules (100 mg, tid) respectively, both for 3 months. Based on the results of routine semen analysis, the patients in the experimental group were further divided into oligozoospermia, asthenozoospermia and normozoospermia subgroups. Semen parameters and sperm acrosin activity were examined before and after treatment. RESULTS: Totally, 220 of the patients completed the treatment and follow-up, 163 in the LC medication and 57 in the VE control group. Compared with the baseline, the percentage of progressively motile sperm (PMS) was significantly increased in the LC group after 3 months of treatment (ï¼»32.58 ± 1.13ï¼½% vs ï¼»36.35 ± 1.26ï¼½%, P < 0.05), and so was sperm acrosin activity (ï¼»37.05±0.66ï¼½ vs ï¼»58.61±1.93ï¼½ µIU/106 sperm, P < 0.01). Sperm concentration, PMS and sperm acrosin activity were also improved in the VE control group after treatment, but with no statistically significant difference (P > 0.05). In comparison with pretreatment, remarkable increases were observed after LC medication in sperm concentration in the oligozoospermia subgroup (ï¼»11.27 ± 0.73ï¼½ vs ï¼»21.82 ± 4.21ï¼½ ×106/ml, P < 0.01) and PMS in the asthenozoospermia patients (ï¼»20.61 ± 0.85ï¼½% vs ï¼»29.81 ± 1.88ï¼½%, P < 0.01). And sperm acrosin activity was even higher after treatment in the asthenozoospermia than in the oligozoospermia and normozoospermia subgroups (ï¼»60.85 ± 3.04ï¼½ vs ï¼»56.32 ± 2.86ï¼½ and ï¼»57.09 ± 6.31ï¼½ µIU/106 sperm, P < 0.05). CONCLUSIONS: L-carnitine can effectively elevate sperm acrosin activity in male infertility patients, particularly in those with asthenozoospermia.

Exposure to Brefeldin A promotes initiation of meiosis in murine female germ cells.

In mammals, ontogenesis starts from a fusion of spermatozoon and oocyte, which are produced by reductive nuclear division of a diploid germ cell in a specialised but complex biological process known as meiosis. However, little is known about the mechanism of meiotic initiation in germ cells, although many factors may be responsible for meiosis both in male and female gonads. In this study, 11.5 days post coitum (dpc) female fetal mouse genital ridges were cultured in vitro with exposure to Brefeldin A (BFA) for 6h, and the changes in meiosis were detected. Synaptonemal-complex analysis implied that BFA played a positive role in meiosis initiation and this hypothesis was confirmed by quantitative PCR of meiosis-specific genes: stimulated by retinoic acid gene 8 (Stra8) and deleted in a zoospermia-like (DAZL). At the same time, mRNA expression of retinoic acid synthetase (Raldh2) and retinoic acid (RA) receptors increased in female gonads with in vitro exposure to BFA. Transplanting genital ridges treated with BFA into the kidney capsule of immunodeficient mice demonstrated that the development capacity of female germ cells was normal, while formation of primordial follicles was seen to be a result of accelerated meiosis after exposure to BFA. In conclusion, the study indicated that BFA stimulated meiosis initiation partly by RA signalling and then promoted the development of follicles.

Exposure to diethylhexyl phthalate (DEHP) results in a heritable modification of imprint genes DNA methylation in mouse oocytes.

Diethylhexyl phthalate (DEHP) is an estrogen-like compound widely used as a plasticizer in commercial products and is present in medical devices, and common household items. It is considered an endocrine disruptor since studies on experimental animals clearly show that exposure to DEHP can alter epigenetics of germ cells. This study was designed to assess the effects of DEHP on DNA methylation of imprinting genes in germ cells from fetal and adult mouse. Pregnant mice were treated with DEHP at doses of 0 and 40 µg DEHP/kg body weight/day from 0.5 to 18.5 day post coitum. The data revealed DEHP exposure significantly reduced the percentage of methylated CpG sites in Igf2r and Peg3 differentially methylated regions (DMRs) in primordial germ cells from female and male fetal mouse, particularly, in the oocytes of 21 dpp mice (F1), which were produced by the pregnant micetreated with DEHP. More surprisingly, the modification of the DNA methylation of imprinted genes in F1 mouse oocytes was heritable to F2 offspring which exhibit lower percentages of methylated CpG sites in imprinted genes DMRs. In conclusion, DEHP exposure can affect the DNA methylation of imprinting genes not only in fetal mouse germ cells and growing oocytes, but also in offspring's oocytes.

Primordial follicle assembly was regulated by Notch signaling pathway in the mice.

Notch signaling pathway, a highly conserved cell signaling system, exists in most multicellular organisms. The objective of this study was to examine Notch signaling pathway in germ cell cyst breakdown and primordial follicle formation. The receptor and ligand genes of Notch pathway (Notch1, Notch2, Jagged1, Jagged2 and Hes1) were extremely down-regulated after newborn mouse ovaries were cultured then exposed to DAPT or L-685,458 in vitro (P < 0.01). Since DAPT or L-685,548 inhibits Notch signaling pathway, the expression of protein LHX8 and NOBOX was significantly reduced during the formation of the primordial follicles. Down-regulated mRNA expression of specific genes including Lhx8, Figla, Sohlh2 and Nobox, were also observed. The percentages of female germ cells in germ cell cysts and primordial follicles were counted after culture of newborn ovaries for 3 days in vitro. The result showed female germ cells in cysts was remarkably up-regulated while as the oocytes in primordial follicles was significantly down-regulated (P < 0.05). In conclusion, Notch signaling pathway may regulate the formation of primordial follicle in mice.

Inhibition of p38 MAPK/NF-κB p65 signaling pathway activity by rare ginsenosides ameliorates cyclophosphamide-induced premature ovarian failure and KGN cell injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax ginseng C. A. Mey., one of the most used herbs in the world, shows effective treatment in reproductive injury. Recent studies have proven that the processed product, red ginseng, which is more active than ginseng itself. Therefore, it is speculated that its main functional component, rare ginsenosides (heat-transformed saponin, HTS), may be effective in treating premature ovarian failure (POF), but its efficacy has not yet been experimentally confirmed. AIM OF THE STUDY: To evaluate whether HTS could attenuate cyclophosphamide-induced inflammation and oxidative damage in POF model rats and the human granulosa-like KGN cell line and protect granulosa cell proliferation. MATERIAL AND METHODS: HTS were isolated from ginsenosides and high performance liquid chromatography (HPLC) analysis was used to analyze the HTS components. Cyclophosphamide (CP) was used to establish a POF rat model and KGN cell injury model. Reactive oxygen species (ROS) and antioxidant enzyme production was determined using specific assays, while inflammatory cytokine secretion was measured by enzyme-linked immunosorbent assay (ELISA). The proliferative function of granulosa cells was assessed using high-content screening and immunohistochemistry to determine the Ki67 protein level. Protein expression in ovarian tissues and KGN cells was analyzed by Western blotting, quantitative real-time PCR (qRT-PCR) was used to determine the transcriptional changes in ovarian tissues and KGN cells. RESULTS: In CP-treated POF model rats, HTS significantly decreased malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) levels, increased glutathione oxidase (GSH) levels, and upregulated Ki67 expression in ovarian granulosa cells. In addition, HTS significantly increased cell survival and Ki67 expression levels in CP-treated cells, and superoxide dismutase (SOD) levels were significantly increased. HTS significantly downregulated IL-6, TNF-α, and interleukin-1ß (IL-1ß) mRNA expression and significantly inhibited nuclear factor kappa-B p65 (NF-κB p65) and p38 mitogen activated protein kinase (p38 MAPK) phosphorylation in POF model rats and KGN cells. Moreover, NF-κB p65 and p38 MAPK levels were significantly increased in ovarian granulosa cells. p65 and p38 protein and gene expression was significantly downregulated. CONCLUSION: HTS ameliorated CP-induced POF and human granulosa cell injury, possibly by inhibiting inflammation and oxidative damage mediated by the p38 MAPK/NF-κB p65 signaling pathway.

Advances in understanding the reproductive toxicity of endocrine-disrupting chemicals in women.

Recently, there has been a noticeable increase in disorders of the female reproductive system, accompanied by a rise in adverse pregnancy outcomes. This trend is increasingly being linked to environmental pollution, particularly through the lens of Endocrine Disrupting Chemicals (EDCs). These external agents disrupt natural processes of hormones, including synthesis, metabolism, secretion, transport, binding, as well as elimination. These disruptions can significantly impair human reproductive functions. A wealth of animal studies and epidemiological research indicates that exposure to toxic environmental factors can interfere with the endocrine system's normal functioning, resulting in negative reproductive outcomes. However, the mechanisms of these adverse effects are largely unknown. This work reviews the reproductive toxicity of five major environmental EDCs-Bisphenol A (BPA), Phthalates (PAEs), Triclocarban Triclosan and Disinfection Byproducts (DBPs)-to lay a foundational theoretical basis for further toxicological study of EDCs. Additionally, it aims to spark advancements in the prevention and treatment of female reproductive toxicity caused by these chemicals.

Pulsatile gonadotropin-releasing hormone therapy induces spermatogenesis in pituitary stalk interruption syndrome: A case report and review of the literature.

BACKGROUND: Pituitary stalk interruption syndrome (PSIS) is a rare anatomical defect of the pituitary gland falling under the spectrum of holoprosencephaly phenotypes. It is characterized by a deficiency in anterior pituitary hormones, such as growth hormone, gonadotropins, and thyroid hormones. Due to the syndrome's rarity and nonspecific manifestations, there is a lack of standardized treatment strategies. Consequently, early diagnosis through imaging and on-time intervention are crucial for improving patients' outcomes. CASE SUMMARY: A 30-year-old man presented with absent secondary sexual characteristics and azoospermia. Laboratory evaluation revealed a deficiency in gonadotropins, while thyroid function was mostly within normal ranges. Magnetic resonance imaging of the pituitary gland showed pituitary stalk agenesis, hypoplasia of the anterior pituitary, and ectopic posterior pituitary, leading to the diagnosis of PSIS. Initially, the patient underwent 6 mo of gonadotropin therapy without significant changes in hormone levels and secondary sexual characteristics. Pulsatile gonadotropin-releasing hormone therapy was then administered, resulting in the detection of sperm in the semen analysis within 3 mo. After 6 mo, routine semen tests showed normal semen quality. The couple faced challenges in conceiving due to abstinence and underwent three cycles of artificial insemination, which was unsuccessful. They also attempted in vitro fertilization, but unfortunately, the woman experienced a miscarriage 10 wk after the embryo transfer. CONCLUSION: Early detection, accurate diagnosis, and timely treatment are crucial in improving the quality of life and fertility of PSIS patients.

[Semen quality and sperm ultrastructure in infertile men with varicocele].

OBJECTIVE: To examine and analyze semen quality and sperm ultrastructural characteristics of infertile patients with varicocele. METHODS: This study included 118 infertile patients with varicocele (the VC group) and 76 normal semen donors (the control group). We obtained routine semen parameters, seminal plasma biochemical markers and the levels of reproductive hormones in the subjects, and observed the changes in sperm structure under the scanning electron microscope and transmission electron microscope. RESULTS: Compared with the normal control, the VC patients showed significantly decreased sperm concentration, sperm progressive motility, sperm viability (P < 0.05), but no remarkable difference in semen volume and non-progressive motility (P > 0.05). The concentrations of zinc and alpha-glycoside enzyme in the seminal plasma were markedly reduced in the VC group in comparison with the controls (P < 0.05), but there was no significant difference in the level of fructose (P > 0.05), nor in such seminal plasma biochemical markers as FSH, LH, T and E2 between the two groups (P > 0.05). The percentage of morphologically normal sperm was dramatically lower in the VC than in the control group ([56.76 +/- 15.32]% vs [12.34 +/- 6.58]%, P < 0.05), and the sperm deformities were mostly in the head and neck, mainly tapering pin head accompanied by complex abnormal differentiation. CONCLUSION: This study demonstrated that VC may lead to oligo-astheno-terato zoospermia, and hence male infertility, which may be attributed to the changes of seminal plasma microenvironment and sperm ultrastructure.

The relationship between serum FSH level and ovarian response during controlled ovarian stimulation.

OBJECTIVES: To evaluate whether serum follicle stimulating hormone (FSH) level during the early controlled ovarian stimulation can be used as a predictor of the ovarian response in the in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles. MATERIAL AND METHODS: The participants of this retrospective study were chosen from Reproductive Medicine Center, Weifang People's Hospital between January 2015 and December 2020.The participants of this study met the age of 20~43 years old, anti-Müllerian hormone (AMH) ≥ 1.2 ng/mL, antral follicle count (AFC) ≥ 5, and the data was complete and no cancellation cycle. Each participant was given GnRH agonist protocol and given a fixed dose of recombinant FSH in the first four days during the controlled ovarian stimulation (COS). According to the number of oocytes retrieved, the participants were divided into two different ovarian response groups. Serum FSH level after the fourth recombinant follicle stimulating hormone (rFSH) injection were compared during the different ovarian responders. RESULTS: The number of participants who met both the inclusion criteria and exclusion criteria was 235. Serum sFSH levels (mean: 11.76 ± 3.10 IU/L) in the inappropriate responders was significantly higher than serum sFSH levels (mean: 10.79 ± 2.52 IU/L) in the superior responders(p = 0.029). There was a weak correlation between serum sFSH levels and the number of oocytes retrieved (r = -0.134, p = 0.041). Serum sFSH levels had significant clinical valuable (p = 0.0346) in predicting the number of oocytes retrieved. CONCLUSIONS: Serum sFSH levels may be a potential marker to predict the ovarian response during the early COS in the IVF/ICSI cycles, which can guide the adjustment of the exogenous rFSH dose.

Protective effect of L­carnitine against oxidative stress injury in human ovarian granulosa cells.

Granulosa cells (GCs) are important for supporting and nourishing oocytes during follicular development and maturation. Oxidative stress (OS) injury of GCs can lead to decreased responsiveness of follicles to follicular stimulating hormone (FSH), which will accelerate ovarian senescence and adversely affect oocyte and embryo quality. Since L-carnitine has been previously reported to exert strong antioxidant activity, the present study aimed to explore the possible effects of L-carnitine on OS injury and FSH receptor (FSHR) expression in ovarian GCs, results of which may be of significance for GCs protection. In the present study, OS was induced in vitro in KGN cells by treatment with H2O2. KGN cells were cultured and divided into the following four groups: Blank, OS, and 40 and 80 µmol/l L-carnitine pre-treatment groups. In the OS group, cells showed nuclear pyknosis, mitochondria swelled irregularly whilst featuring fractured cristae. In addition, cell viability, ROS levels, superoxide dismutase levels, glutathione levels, malondialdehyde levels, the mitochondrial membrane potential and FSHR expression, as determined by Cell Counting Kit-8 (CCK-8), 2,7-dichloro-dihydrofluorescein diacetate, spectrophotometry, ELISA, spectrophotometry, JC-1 and western blot analyses, respectively, were all significantly different in the OS group compared with those in the control group. However, malonaldehyde levels, reactive oxygen species levels and the apoptosis rate according to flow cytometry were all significantly increased compared with those in the control. Compared with those in the OS group, the morphology of cells and mitochondria in the L-carnitine pre-treatment groups were improved, whilst cell viability and the expression of FSHR were significantly increased but oxidative stress injury was decreased. The present results suggest that L-carnitine can protect the cells from OS damage induced by H2O2, enhance antioxidant activity whilst suppressing the apoptosis of GCs, in addition to preserving FSHR expression in GCs under OS. Therefore, the present study revealed that the introduction of L-carnitine in clinical medicine or dietary supplement may protect GCs, improve follicular quality and female reproductive function.

Effectiveness of letrozole in pituitary downregulated normogonadotrophic young women with an initial poor response.

It has been reported that 10 to 15% of young normogonadotrophic women show suboptimal response to standard long protocols. Letrozole (LE), an aromatase inhibitor, was shown to improve ovarian sensitivity to follicle stimulating hormone (FSH) and follicular response to gonadotrophin treatment in poor ovarian response patients. We reasoned that it might be possible to utilize LE in young normogonadotrophic patients with unexpected hypo-response in standard gonadotropin-releasing hormone agonist long protocol. A total of 652 patients defined as normogonadotrophic patients with unexpected hypo-response were divided into 2 groups, the +LE group and the +Gn group. +LE group: A fixed daily dose of 2.5 mg of LE was added on day 8 of stimulation. +Gn group: A fixed daily dose of 75 U of human menopausal gonadotrophin was added on day 8 of stimulation. The primary outcome measures were the number of oocytes obtained, fertilization rate, days of stimulation, and total FSH dosage. The secondary outcome measures were the implantation rate and ongoing pregnancy rate. There were no significant differences in the clinical and hormonal characteristics between the 2 groups. A shorter duration of stimulation and a lower dosage of recombinant FSH consumption on the day of human chorionic gonadotropin administration were all observed in the +LE group. Patients who received LE therapy showed a higher number of oocytes obtained and significantly higher fertilization rates. The implantation rate and ongoing pregnancy rate were comparable in both groups. LE significantly improves the number of oocytes obtained in patients with suboptimal response to standard gonadotropin-releasing hormone agonist long protocol.

Effect of Y Chromosome Microdeletions on the Pregnancy Outcome of Assisted Reproduction Technology: a Meta-analysis.

This systematic analysis aimed to summarize the effects of Y chromosome microdeletions (YCMs) on pregnancy outcomes of assisted reproductive technology (ART). This retrospective controlled meta-analysis evaluated the effect of YCMs on pregnancy outcomes of ART. Full-text retrieval was conducted in the PubMed, CBM, Web of Science, CNKI, VIP, and WANFANG databases. The pregnancy outcomes included fertilization rate, good embryo rate, clinical pregnancy rate, early miscarriage rate, miscarriage rate, live birth rate, and baby boy rate. The quality of these studies was evaluated using the Newcastle-Ottawa scale. Statistical software Review Manager 5.3 and STATA 14.0 were used. Twelve high-quality studies were included in the analysis. Compared with that in the normal group, the fertilization rate in the YCMs group decreased significantly (odds ratio [OR] = 0.75, 95% confidence interval [CI] [0.63, 0.88], P = 0.0006). However, there was no significant difference (P > 0.05) between groups in the good embryo rate (OR = 0.88, 95% CI [0.72, 1.07]), clinical pregnancy rate (OR = 0.94, 95% CI [0.78, 1.11]), early miscarriage rate (OR = 1.70, 95% CI [0.93, 3.10]), miscarriage rate (OR = 1.3, 95% CI [0.93, 1.91]), live birth rate (OR = 0.90, 95% CI [0.74, 1.08]), and baby boy rate (OR = 1.15, 95% CI [0.85, 1.56]). YCMs are associated with a reduced fertilization rate of ART, but they do not decrease the good embryo rate, clinical pregnancy rate, early miscarriage rate, miscarriage rate, live birth rate, or baby boy rate.

Blockade of epithelial sodium channels improves sperm motility in asthenospermia patients.

Epithelial Na(+) channels (ENaCs) are ion channels that play important roles in physiology as well as pathophysiology. Inhibiting ENaCs using amiloride and its derivatives has been suggested in treatment of cardiovascular disease and hypertension. By immunoblotting, we demonstrated the presence of ENaC-alpha protein in the flagellar midpiece of both rat and human sperm. Immunohistochemistry analyses in rat testis localized ENaC-alpha expressed in the Leydig cell, Sertoli cell, Ap spermatogonium, spermatocyte, spermatid and residual body. Importantly, using computer-assisted sperm motility analysis, we first observed that EIPA [5-(N-ethyl-N-isopropyl)-amiloride hydrochloride] inhibition of ENaCs, possibly including ENaC-alpha and ENaC-delta, significantly improved the sperm motility in healthy donors by 14.23% (mean +/- SEM, 68.75 +/- 9.76% vs. 78.53 +/- 6.20%, p < 0.001) and in asthenospermia patients by 115.89% from 9.50 +/- 6.11% to 20.51 +/- 12.13% (mean +/- SEM, p < 0.001). The improved sperm motility by EIPA has important clinical implications in the treatment of asthenospermia and certainly warrants further investigation.

[SHENG MAI ZHUSHEYE improves the viability and movement parameters of human sperm in vitro].

OBJECTIVE: To observe the effect of SHENG MAI ZHUSHEYE on the movement parameters and viability of human sperm in vitro. METHODS: We collected sperm samples from 33 normal fertile men, divided each into two, and cultured them in vitro with SHENG MAI ZHUSHEYE + Hams-F10 and Hams-F10 alone, respectively. Then we measured the straight line velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP) and the amplitude of lateral head displacement (ALH) of the sperm by computer-aided semen analysis at 0.5, 1, 2, 4, 8 and 12 h. And the sperm viability was detected. RESULTS: VCL was significantly higher at 8 h (P < 0.05) and very significantly higher at 12 h (P < 0.01) in the SHENG MAI ZHUSHEYE + Hams-F10 group than in the Hams-F10 group. VSL, VAP and ALH were significantly increased in the former group at 4, 8 and 12 h as compared with the latter (P < 0.05). The sperm viability was significantly decreased in the Hams-F10 group at 12 h (P < 0.05). CONCLUSION: SHENG MAI ZHUSHEYE can improve sperm movement parameters and increase sperm viability in vitro.

Expression of angiopoietin-1/-2 in the process of mouse embryo implantation.

This study examined the expression and distribution of angiopoietin-1/-2 (Ang-1/-2) in the endometrium of early pregnant mice. The expression of Ang-1/-2 was detected by immunohisto-chemical staining and in situ hybridization respectively. Computerized image analysis system was used to measure the average optical intensity of Ang-1/-2 in endometria at different time points after gestation. Mice were randomly divided into 5 groups: control group, D2 group (2 days after pregnancy), D4 group (4 days after pregnancy), D6 group (6 days after pregnancy) and D8 group (8 days after pregnancy), each containing 15 mice. The results showed that the expression of Ang-1 and Ang-2 was very different among 4 groups (P<0.01). Immunohistochemical staining revealed that Ang-1 was localized in the cytoplasma of stromal cells 2 days after pregnancy (day 2), and in luminal epithelial cells on day 4. The protein of Ang-2 was mainly expressed in the cytoplasma of glandular epithelia and stromal cells. With gestation time, the positive reactions of Ang-1/-2 were stronger in the endometria of the pregnant mice (P<0.01). In situ hybridization showed Ang-1 mRNA in stromal cells on day 2. Hybridization signal was localized in both stromal cells and vessel epithelial cells on day 4; Ang-2 mRNA was expressed in stromal cells and glandular epithelia on day 2; high mRNA levels appeared in stromal cells, glandular epithelia and vascular endothelia on day 4; an increasing in mRNA expression of Ang-1/-2 was observed on day 6 and day 8 (P<0.01). It is suggested that Ang-1/-2 may play an important role in the cross-talk between blastocyst and maternal endometrium during the process of embryo implantation.

[Effects of co-administration of growth hormone(GH) and aspirin to women during in vitro fertilization and embryo transfer (IVF-ET) cycles].

OBJECTIVE: To study the effects of the co-administration of growth hormone (GH) and aspirin to women with suboptimal response to GnRHa/FSH hyperstimulation protocol during in vitro fertilization and embryo transfer (IVF-ET) cycles. METHODS: Forty cases of poor ovarian response in previous IVF-ET cycles were randomly divided into 2 groups: the studied group of GH and aspirin (n = 20), and the control group without GH or aspirin (n = 20). RESULTS: The co-administration of GH and aspirin significantly increased the rates of retrieved oocytes (P < 0.01), promoted the maturation of oocytes (P < 0.01) and improve the fertilization rates (P < 0.05). However, there were no statistically differences between the two groups in the number of replaced embryos (P > 0.05) and the pregnancy rate (P > 0.05). CONCLUSION: The co-administration of GH and aspirin to poor ovarian responders is effective to increase the rates of retrieved oocytes, promote the maturation of oocytes and improve the fertilization rate in IVF-ET.

Differentiation of early germ cells from human skin-derived stem cells without exogenous gene integration.

Infertility has long been a difficult issue for many couples. The successful differentiation of germ cells and live progeny from pluripotent stem cells brings new hope to the couples suffering with infertility. Here we successfully isolated human fetus skin-derived stem cells (hfSDSCs) from fetus skin tissue and demonstrated that hfSDSCs can be differentiated into early human germ cell-like cells (hGCLCs). These cells express human germ cell markers DAZL and VASA. Moreover, these pluripotent stem cell-derived hGCLCs are free of exogenous gene integration. When hfSDSCs were differentiated in porcine follicle fluid (PFF) conditioned media, which has been shown to promote the differentiation of mouse and porcine SDSCs into oocyte-like cells (OLCs), we observed some vesicular structures formed from hfSDSCs. Moreover, when hfSDSCs were cultured with specific conditioned media, we observed punctate and elongated SCP3 staining foci, indicating the initiation of meiosis. Ploidy analysis and fluorescent in situ hybridization (FISH) analysis indicated that a small percentage of putative 1N populations formed from hfSDSCs when compared with positive controls. In conclusion, our data here, for the first time, demonstrated that hfSDSCs possess the differentiation potential into germ lines, and they may differentiate both male and female hGCLCs in vitro under appropriate conditions.