Gonadotropin elevation is ootoxic to ovulatory oocytes and inhibits oocyte maturation, and activin decoy receptor ActRIIB:Fc therapeutically restores maturation.

BACKGROUND: Elevated FSH often occurs in women of advanced maternal age (AMA, age ≥ 35) and in infertility patients undergoing controlled ovarian stimulation (COS). There is controversy on whether high endogenous FSH contributes to infertility and whether high exogenous FSH adversely impacts patient pregnancy rates. METHODS: The senescence-accelerated mouse-prone-8 (SAMP8) model of female reproductive aging was employed to assess the separate impacts of age and high FSH activity on the percentages (%) of viable and mature ovulated oocytes recovered after gonadotropin treatment. Young and midlife mice were treated with the FSH analog equine chorionic gonadotropin (eCG) to model both endogenous FSH elevation and exogenous FSH elevation. Previously we showed the activin inhibitor ActRIIB:Fc increases oocyte quality by preventing chromosome and spindle misalignments. Therefore, ActRIIB:Fc treatment was performed in an effort to increase % oocyte viability and % oocyte maturation. RESULTS: The high FSH activity of eCG is ootoxic to ovulatory oocytes, with greater decreases in % viable oocytes in midlife than young mice. High FSH activity of eCG potently inhibits oocyte maturation, decreasing the % of mature oocytes to similar degrees in young and midlife mice. ActRIIB:Fc treatment does not prevent eCG ootoxicity, but it restores most oocyte maturation impeded by eCG. CONCLUSIONS: FSH ootoxicity to ovulatory oocytes and FSH maturation inhibition pose a paradox given the well-known pro-growth and pro-maturation activities of FSH in the earlier stages of oocyte growth. We propose the FOOT Hypothesis ("FSH OoToxicity Hypothesis), that FSH ootoxicity to ovulatory oocytes comprises a new driver of infertility and low pregnancy success rates in DOR women attempting spontaneous pregnancy and in COS/IUI patients, especially AMA women. We speculate that endogenous FSH elevation also contributes to reduced fecundity in these DOR and COS/IUI patients. Restoration of oocyte maturation by ActRIB:Fc suggests that activin suppresses oocyte maturation in vivo. This contrasts with prior studies showing activin A promotes oocyte maturation in vitro. Improved oocyte maturation with agents that decrease endogenous activin activity with high specificity may have therapeutic benefit for COS/IVF patients, COS/IUI patients, and DOR patients attempting spontaneous pregnancies.

Therapeutic benefits of central LH receptor agonism in the APP/PS1 AD model involve trophic and immune regulation and are reproductive status dependent.

The mechanisms that underly reproductive hormone effects on cognition, neuronal plasticity, and AD risk, particularly in relation to gonadotropin LH receptor (LHCGR) signaling, remain poorly understood. To address this gap in knowledge and clarify the impact of circulating steroid hormones on the therapeutic effects of CNS LHCGR activation, we delivered the LHCGR agonist human chorionic gonadotropin (hCG) intracerebroventricularly (ICV) and evaluated functional, structural, plasticity-related signaling cascades, Aß pathology, and transcriptome differences in reproductively intact and ovariectomized (OVX) APP/PS1 AD female mice. Here we demonstrate that CNS hCG delivery restored function to wild-type levels only in OVX APP/PS1 mice. Spine density was increased in all hCG treated groups independently of reproductive status. Notably, increases in BDNF signaling and cognition, were selectively upregulated only in the OVX hCG-treated group. RNA sequencing analyses identified a significant increase in peripheral myeloid and pro-inflammatory genes within the hippocampi of the OVX group that were completely reversed by hCG treatment, identifying a potential mechanism underlying the selective therapeutic effect of LHCGR activation. Interestingly, in intact mice, hCG administration mimicked the effects of gonadectomy. Together, our findings indicate that CNS LHCGR agonism in the post-menopausal context is beneficial through trophic and immune mechanisms. Our findings also underscore the presence of a steroid-LHCGR mechanistic interaction that is unexplored yet potentially meaningful to fully understand "post-menopausal" brain function and CNS hormone treatment response.

Enhancing endometrial receptivity: the roles of human chorionic gonadotropin in autophagy and apoptosis regulation in endometrial stromal cells.

Inadequate endometrial receptivity often results in embryo implantation failure and miscarriage. Human chorionic gonadotropin (hCG) is a key signaling molecule secreted during early embryonic development, which regulates embryonic maternal interface signaling and promotes embryo implantation. This study aimed to examine the impact of hCG on endometrial receptivity and its underlying mechanisms. An exploratory study was designed, and endometrial samples were obtained from women diagnosed with simple tubal infertility or male factor infertile (n = 12) and recurrent implantation failure (RIF, n = 10). Using reverse transcription-quantitative PCR and western blotting, luteinizing hormone (LH)/hCG receptor (LHCGR) levels and autophagy were detected in the endometrial tissues. Subsequently, primary endometrial stromal cells (ESCs) were isolated from these control groups and treated with hCG to examine the presence of LHCGR and markers of endometrial receptivity (HOXA10, ITGB3, FOXO1, LIF, and L-selectin ligand) and autophagy-related factors (Beclin1, LC3, and P62). The findings revealed that the expressions of receptivity factors, LHCGR, and LC3 were reduced in the endometrial tissues of women with RIF compared with the control group, whereas the expression of P62 was elevated. The administration of hCG to ESCs specifically activated LHCGR, stimulating an increase in the endometrial production of HOXA10, ITGB3, FOXO1, LIF and L-selectin ligands. Furthermore, when ESCs were exposed to 0.1 IU/mL hCG for 72 h, the autophagy factors Beclin1 and LC3 increased within the cells and P62 decreased. Moreover, the apoptotic factor Bax increased and Bcl-2 declined. However, when small interfering RNA was used to knock down LHCGR, hCG was less capable of controlling endometrial receptivity and autophagy molecules in ESCs. In addition, hCG stimulation enhanced the phosphorylation of ERK1/2 and mTOR proteins. These results suggest that women with RIF exhibit lower levels of LHCGR and compromised autophagy function in their endometrial tissues. Thus, hCG/LHCGR could potentially improve endometrial receptivity by modulating autophagy and apoptosis.

Vitamin D3 reduces the symptoms of ovarian hyperstimulation syndrome in mice and inhibits the release of granulosa cell angiogenic factor through pentraxin 3.

It has been reported that the effective inhibition of vascular endothelial growth factor (VEGF) can prevent the progression of ovarian hyperstimulation syndrome (OHSS). The present study aimed to investigate the mechanism underlying the effect of vitamin D3 (VD3) on OHSS in mouse models and granulosa cells. The effects of VD3 administration (16 and 24 IU) on ovarian permeability were determined using Evans blue. In addition, ovarian pathology, corpus luteum count, inflammatory responses, and hormone and VEGFA levels were assessed using pathological sections and ELISA. Molecular docking predicted that pentraxin 3 (PTX3) could be a potential target of VD3, and therefore, the effects of human chorionic gonadotropin (hCG) and VD3 as well as PTX3 overexpression on the production and secretion of VEGFA in granulosa cells were also investigated using western blotting and immunofluorescence. Twenty-four IU VD3 significantly reversed the increase in ovarian weight and permeability in mice with OHSS. Additionally, VD3 diminished congestion and the number of corpus luteum in the ovaries and reduced the secretion levels of inflammatory factors and those of estrogen and progesterone. Notably, VD3 downregulated VEGFA and CD31 in ovarian tissues, while the expression levels of PTX3 varied among different groups. Furthermore, VD3 restored the hCG-induced enhanced VEGFA and PTX3 expression levels in granulosa cells, whereas PTX3 overexpression abrogated the VD3-mediated inhibition of VEGFA production and secretion. The present study demonstrated that VD3 could inhibit the release of VEGFA through PTX3, thus supporting the beneficial effects of VD3 administration on ameliorating OHSS symptoms.

Reproductive outcomes of dual trigger therapy with GnRH agonist and hCG versus hCG trigger in women with diminished ovarian reserve: a retrospective study.

BACKGROUND: Diminished ovarian reserve (DOR) is one of the obstacles affecting the reproductive outcomes of patients receiving assisted reproductive therapy. The purpose of this study was to investigate whether dual trigger, including gonadotropin-releasing hormone agonist (GnRHa) and human chorionic gonadotropin (hCG), can improve pregnancy outcomes in patients with DOR undergoing in vitro fertilization (IVF) cycles using mild stimulation protocols. METHODS: A total of 734 patients with DOR were included in this retrospective study. Patients were divided into a recombinant hCG trigger group and a dual trigger group (hCG combined with GnRHa) according to the different trigger drugs used. The main outcome measures included the number of oocytes retrieved, the fertilization rate, the number of transferable embryos, the implantation rate, the clinical pregnancy rate, the miscarriage rate, the live birth rate (LBR), and the cumulative live birth rate (CLBR). Generalized linear model and logistic regression analyses were performed for confounding factors. RESULTS: There were 337 cycles with a single hCG trigger and 397 cycles with dual trigger. The dual trigger group demonstrated significantly higher numbers of retrieved oocytes [3.60 vs. 2.39, adjusted ß = 0.538 (0.221-0.855)], fertilized oocytes [2.55 vs. 1.94, adjusted ß = 0.277 (0.031-0.523)] and transferable embryos [1.22 vs. 0.95, adjusted ß = 0.162 (-0.005-0.329)] than did the hCG trigger group, whereas no significant difference in the fertilization rate was observed between the two groups. Moreover, the embryo transfer cancellation rate (35.5% vs. 43.9%) was obviously lower in the dual trigger group. Among the fresh embryo transfer cycles, the implantation rate, clinical pregnancy rate, miscarriage rate and live birth rate were similar between the two groups. After controlling for potential confounding variables, the trigger method was identified as an independent factor affecting the number of oocytes retrieved but had no significant impact on the CLBR. CONCLUSIONS: Dual triggering of final oocyte maturation with hCG combined with GnRHa can significantly increase the number of oocytes retrieved in patients with DOR but has no improvement effect on the implantation rate, clinical pregnancy rate or LBR of fresh cycles or on the CLBR.

Elevated chorionic gonadotropic hormone in transgenic mice induces parthenogenetic activation and ovarian teratomas.

Both male and female reproductive functions are impacted by altered gonadotrophin secretion and action, which may also influence the development of endocrine tumours. To ascertain if chronic hypersecretion of human chorionic gonadotropin (hCG) contributes to the development of gonadal tumours, double transgenic (TG) mice that overexpress hCGα- and ß-subunits were analysed. By the age of two months, ovarian tumours with characteristics of teratomas developed with 100% penetrance. Teratomas were also seen in wild-type ovaries orthotopically transplanted into TG mice, demonstrating an endocrine/paracrine mechanism for the hCG-induced ovarian tumorigenesis. Both in vitro and in vivo experiments showed oocyte parthenogenetic activation in TG females. In addition, ovaries showed reduced ovulatory gene expression, inhibited ERK1/2 phosphorylation, and impaired cumulus cell expansion. Hence, persistently high endocrine hCG activity causes parthenogenetic activation and development of ovarian teratomas, along with altered follicle development and impaired ERK1/2 signalling, offering a novel mechanism associated with the molecular pathogenesis of ovarian teratomas.

The effect of human menopausal gonadotropin and equine chorionic gonadotropin on the reproductive performance of treated ewes with short-term progesterone injections and sponge during the non-breeding season.

This study investigated the effect of human menopausal gonadotropin (hMG) on reproductive efficiency of synchronized ewes with the sponge and progesterone (P4) injection-based protocols. In study 1, anoestrous ewes (n = 120) were used. Sixty ewes were treated with sponge (S) for 12 days. The injection of eCG (SeCG group, n = 30) or hMG (ShMG, n = 30) was given at the time of sponge removal. Thirty ewes received IM injection of P4, three times every 48 h and the injection of hMG was given 24 h after the third P4 injection (3PhMG group, n = 30), and 30 ewes were used as control group. Pregnancy was diagnosed on day 50 after the release of ram. In study 2, 60 ewes were randomly divided into two equal groups. In the treated group with antibiotics (n = 30), before inserting, the sponges were impregnated with the antibiotic penicillin G sodium (5,000,000 IU) and in the control group (n = 30), there was no added antibiotics. Before inserting and after removing sponges, a vaginal cytology sample was taken with a sterile cotton swab. The number of neutrophils in each sample was counted and analysed. The rate of oestrus and total pregnancy was greater in SeCG (96.7, 93.3%), ShMG (82.8, 93.1%) and 3PhMG (67.9, 89.3%) groups compared with the control group (13.8, 41.4%) (p < .05). No significant difference was found in single, twin and total lambing and pregnancy rates after injection of eCG and hMG during the non-breeding season (p > .05). A higher percentage of control ewes had the vaginal smear with neutrophils more than 50% (96.7% vs. 76.7%; p < .05). In conclusion, a single dose of hMG can induce fertile oestrus in synchronized ewes with P4 administered by either injection or intravaginally. Purulent discharge and percentage of neutrophils were significantly reduced in the synchronized ewes by the impregnated sponges with the antibiotic penicillin.

Refrigerated storage and cryopreservation of hormonally induced sperm in the threatened frog, Litoria aurea.

As sperm cryopreservation and other assisted reproductive technologies (ARTs) advance in common amphibian species, focus on applying non-lethal sperm collection methods to the conservation and genetic management of threatened species is imperative. The goal of this study was to examine the application of logistically practical ART protocols in a threatened frog (Litoria aurea). First, we tested the efficacy of various concentrations of human chorionic gonadotropin (hCG) (20, 40 IU/g bodyweight) and Gonadotropin releasing hormone antagonist (0.25 µg/g and 0.5 µg/g body weight GnRH-a) on the induction of spermatozoa. Using the samples obtained from the previous trials, we tested the effect of cold storage and cryopreservation protocols on long-term refrigerated storage and post-thaw sperm recovery. Our major findings include: (1) high quality sperm were induced with 20 and 40 IU/g bodyweight of (hCG); (2) proportions of live, motile sperm post-thaw, were recovered at higher levels than previously reported for L. aurea (>50%) when preserved with 15% v/v DMSO and 1% w/v sucrose; and (3) spermic urine stored at 5 °C retained motility for up to 14 days. Our findings demonstrate that the protocols developed in this study allowed for successful induction and recovery of high-quality spermatozoa from a threatened Australian anuran, L. aurea, providing a prime example of how ARTs can contribute to the conservation of rare and threatened species.

Effects of equine chorionic gonadotropin dosage and its splitting in different days on reproductive performance of Nellore cows synchronized for timed-artificial insemination.

This study evaluated the effects of dose of equine chorionic gonadotropin (eCG) and its splitting in different days of the synchronization protocol on reproductive performance of primiparous and multiparous Nellore cows. In the present study, 2,536 Nellore cows (1,634 primiparous and 902 multiparous) were assigned to receive in a 2 × 2 factorial design 1) an intravaginal progesterone (P4) device and 2.0 mg of estradiol benzoate (EB) on day -11, 12.5 mg (i.m.) of dinoprost tromethamine (PGF), 300 IU (i.m.) of eCG, 0.6 mg (i.m.) of estradiol cypionate (ECP), and P4 device withdrawal on day -2, followed by TAI on day 0 (n = 632 cows, being 409 primiparous and 223 multiparous; 300-2), 2) 300 IU (i.m) of eCG administered on days -4 and -2 (150 IU of eCG/day; n = 637 cows, being 412 primiparous and 225 multiparous; 300-4-2), 3) 400 IU (i.m.) of eCG administered on day -2 (n = 633 cows, being 406 primiparous and 227 multiparous; 400-2), and 4) 400 IU (i.m) of eCG administered on days -4 and -2 (200 IU of eCG/day; n = 634 cows, being 407 primiparous and 227 multiparous; 400-4-2). Individual cow BCS was assessed on days -11, 0 (timed-AI), and 31 of the study. Body condition score of the animals was classified into LOW or HIGH using the threshold of 2.75 (≤2.75 = LOW; >2.75 = HIGH). For primiparous cows, an eCG splitting effect was observed on follicle size, as cows receiving eCG on days -4 and -2 of the synchronization protocol had a larger follicle than cows administered eCG only on day -2. For day 31 P/AI, primiparous cows receiving 400-4-2, regardless of BCS, had a greater P/AI than cows from other treatments. Administering 400-4-2 to LOW BCS cows also resulted in greater P/AI than all other treatments assigned to LOW BCS cows. For multiparous cows, no treatment effect was observed for follicle size, estrus expression, and day 31 P/AI (P ≥ 0.21). In summary, increasing the dose and splitting the dose of eCG positively impacted the pregnancy rates of primiparous cows under a BCS ≤2.75, but no effects were detected on multiparous cows.

Benzo[a]pyrene disrupts LH/hCG-dependent mouse Leydig cell steroidogenesis through receptor/Gαs protein targeting.

Steroidogenesis of gonadal cells is tightly regulated by gonadotropins. However, certain polycyclic aromatic hydrocarbons, including Benzo[a]pyrene (BaP), induce reproductive toxicity. Several existing studies have considered higher than environmentally relevant concentrations of BaP on male and female steroidogenesis following long-term exposure. Also, the impact of short-term exposure to BaP on gonadotropin-stimulated cells is understudied. Therefore, we evaluated the effect of 1 nM and 1 µM BaP on luteinizing hormone/choriogonadotropin (LH/hCG)-mediated signalling in two steroidogenic cell models, i.e. the mouse tumor Leydig cell line mLTC1, and the human primary granulosa lutein cells (hGLC) post 8- and 24-h exposure. Cell signalling studies were performed by homogeneous time-resolved fluorescence (HTRF) assay, bioluminescence energy transfer (BRET) and Western blotting, while immunostainings and immunoassays were used for intracellular protein expression and steroidogenesis analyses, respectively. BaP decreased cAMP production in gonadotropin-stimulated mLTC1 interfering with Gαs activation. Therefore, decrease in gonadotropin-mediated CREB phosphorylation in mLTC1 treated with 1 µM BaP was observed, while StAR protein levels in gonadotropin-stimulated mLTC1 cells were unaffected by BaP. Further, BaP decreased LH- and hCG-mediated progesterone production in mLTC1. Contrastingly, BaP failed to mediate any change in cAMP, genes and proteins of steroidogenic machinery and steroidogenesis of gonadotropin-treated hGLC. Our results indicate that short-term exposure to BaP significantly impairs steroidogenic signalling in mLTC1 interfering with Gαs. These findings could have a significant impact on our understanding of the mechanism of reproductive toxicity by endocrine disruptors.

Equine chorionic gonadotropin treatment and timed artificial insemination for dairy cow production under heat stress.

This study investigated the effects of timed artificial insemination (TAI) and equine chorionic gonadotropin (eCG) administration on lactating dairy cows under heat-stress conditions (average temperature-humidity index: 80). Timed artificial insemination was performed on the cows with (n = 57) or without (control, n = 41) supplementation with 500 IU of eCG at the day of PGF2α treatment using the CIDR-Ovsynch protocol. GnRH was administered, and a progesterone device (CIDR) was inserted on Day -10 of the treatment protocol. The CIDR was removed on Day -3, and the cows were treated with PGF2α. Two days later, a 2nd GnRH injection was administered. Subsequently, AI was performed on Day 0 (16-20 h after the 2nd GnRH injection), and pregnancy was diagnosed on Days 32 and 60. Plasma progesterone (P4) concentrations were measured after AI. Results showed that the eCG group had a higher pregnancy per AI (P/AI) than the control group (43.9 vs. 12.2%, P = 0.002), which was also accompanied by elevated P4 levels. Four cows in the eCG group had multiple calves, representing 7.0 and 16.0% of the group and pregnant cows, respectively. In conclusion, 500 IU of eCG combined with CIDR-Ovsynch in lactating dairy cows under severe heat stress conditions successfully improved fertility. However, the protocol may have a slight risk of multiple births.

Frozen-thawed embryo transfer: does the addition of low-dose choriogonadotropin alfa to progesterone in the luteal phase of artificial cycles improve the endometrium and increase the chances of pregnancy?

OBJECTIVE: Primary: To evaluate the effect of low doses of recombinant hCG (choriogonadotropin alfa) in the luteal phase of frozen-thawed embryo transfers (FET) of artificial cycles on the chances of pregnancy in patients aged ≤38years. Secondary: To assess the chances of pregnancy in the FET groups of artificial cycles using micronized vaginal progesterone (VP) versus injectable intramuscular progesterone (IMP) and the chances of pregnancy in type-1 embryo transfers (two top embryos). METHODS: This retrospective cohort study included 122 cycles of FET and compared two groups of patients aged 38 years or younger, one given hCG in the luteal phase and one not administered hCG. RESULTS: The clinical pregnancy rates (CPR) in the control and hCG groups were 45% and 45.16%, respectively (p=0.9999). The live birth rates (LBR) were 33.33% and 32.25%, respectively, (p=0.99909). The CPR in the VP group (83 patients) was 46.89% versus 41.02% in the IMP group, (p=0.5459). The LBR was 33.73% in the VP group and 30.76% in the IMP group (39 patients), (p=0.7559). CONCLUSIONS: The CPR and LBR of patients undergoing FET in groups prescribed and not prescribed low doses of recombinant hCG were similar. No significant difference was found between patients given VP or IMP.

Effect of GnRH agonist trigger with or without low-dose hCG on reproductive outcomes for PCOS women with freeze-all strategy: a propensity score matching study.

PURPOSE: This study aimed to compare the effect of gonadotropin-releasing hormone agonist (GnRHa) trigger alone versus dual trigger comprising GnRHa and low-dose human chorionic gonadotropin (hCG) on reproductive outcomes in patients with polycystic ovary syndrome (PCOS) who received the freeze-all strategy. METHODS: A total of 615 cycles were included in this retrospective cohort study. Propensity score matching (PSM) was performed to control potential confounding factors between GnRHa-trigger group (0.2 mg GnRHa) and dual-trigger group (0.2 mg GnRHa plus 1000/2000 IU hCG) in a 1:1 ratio. Multivariate logistic regression was applied to estimate the association between trigger methods and reproductive outcomes. RESULTS: After PSM, patients with dual trigger (n = 176) had more oocytes retrieved, mature oocytes, and 2PN embryos compared to that with GnRHa trigger alone. However, the oocytes maturation rate, normal fertilization rate, and frozen embryos between the two groups were not statistically different. The incidence of ovarian hyperstimulation syndrome (OHSS) (14.8% vs. 2.8%, P < 0.001) and moderate/severe OHSS (11.4% vs. 1.7%, P < 0.001) were significantly higher in dual-trigger group than in GnRHa-alone group. Logistic regression analysis showed the adjusted odds ratio of dual trigger was 5.971 (95% confidence interval 2.201-16.198, P < 0.001) for OHSS. The pregnancy and single neonatal outcomes were comparable between the two groups (P > 0.05). CONCLUSION: For PCOS women with freeze-all strategy, GnRHa trigger alone decreased the risk of OHSS without damaging oocyte maturation and achieved satisfactory pregnancy outcomes.

Pulsatile Gonadotropin-Releasing Hormone Therapy Is Associated With Better Spermatogenic Outcomes than Gonadotropin Therapy in Patients With Pituitary Stalk Interruption Syndrome.

OBJECTIVE: To compare the effects of combined gonadotropin and pulsatile gonadotropin-releasing hormone (GnRH) therapy on spermatogenesis in patients with pituitary stalk interruption syndrome (PSIS). METHODS: Male patients with PSIS (N = 119) were retrospectively studied. Patients received pulsatile GnRH therapy (N = 59) were divided into response and poor-response groups based on luteinizing hormone (LH) levels after 1-month treatment with a cutoff value of 1 or 2 IU/L. Participants with gonadotropin therapy were divided into human menopausal gonadotropin (hMG)/human chorionic gonadotropin (hCG) group (N = 60), and patients with pulsatile GnRH therapy were classified into GnRH group (N = 28) with treatment duration ≥6 months. RESULTS: The overall success rates of spermatogenesis for hMG/hCG and GnRH therapy were 51.67% (31/60) vs 33.90% (20/59), respectively. GnRH group required a shorter period to induce spermatogenesis (8 vs 15 months, P = .019). hMG/hCG group had higher median total testosterone than GnRH group [2.16, interquartile range(IQR) 1.06-4.89 vs 1.31, IQR 0.21-2.26 ng/mL, P = .004]. GnRH therapy had a beneficial effect on spermatogenesis compared to hMG/hCG therapy (hazard ratio 1.97, 95% confidence interval 1.08-3.57, P = .026). In patients with pulsatile GnRH therapy, compared with the poor-response group, the response group had a higher successful spermatogenesis rate (5.00% vs 48.72%, P = .002) and higher median basal total testosterone (0.00, IQR 0.00-0.03 vs 0.04, IQR 0.00-0.16 ng/mL, P = .026) with LH = 1 IU/L as the cutoff value after 1-month pulsatile GnRH therapy. CONCLUSIONS: Pulsatile GnRH therapy was superior to hMG/hCG therapy for spermatogenesis in patients with PSIS. Earlier spermatogenesis and higher concentrations of sperm could be obtained in the GnRH group if patients received therapy over 6 months.

Synergistic Effect of Human Chorionic Gonadotropin and Granulocyte Colony Stimulating Factor in the Mobilization of HSPCs Improves Overall Survival After PBSCT in a Preclinical Murine Model. Are We Far Enough for Therapy?

Strategies to improve hematopoietic stem and progenitor cell (HSPC) mobilization from the bone marrow can have a pivotal role in addressing iatrogenic bone-marrow insufficiency from chemo(radio)therapy and overcoming peripheral blood stem cell transplantation (PBSCT) limitations such as insufficient mobilization. Granulocyte-colony stimulating factor (G-CSF) represents the standard mobilization strategy for HSPC and has done so for more than three decades since its FDA approval. Its association with non-G-CSF agents is often employed for difficult HSPC mobilization. However, obtaining a synergistic effect between the two classes is limited by different timing and mechanisms of action. Based on our previous in vitro results, we tested the mobilization potential of human chorionic gonadotropin (HCG), alone and in combination with G-CSF in vivo in a murine study. Our results show an improved mobilization capability of the combination, which seems to act synergistically in stimulating hematopoiesis. With the current understanding of the dynamics of HSPCs and their origins in more primitive cells related to the germline, new strategies to employ the mobilization of hematopoietic progenitors using chorionic gonadotropins could soon become clinical practice.

Shear stress in the intervillous space promotes syncytial formation of iPS cells-derived trophoblasts†.

The intervillous space of human placenta is filled with maternal blood, and villous trophoblasts are constantly exposed to the shear stress generated by maternal blood pressure and flow throughout the entire gestation period. However, the effects of shear stress on villous trophoblasts and their biological significance remain unknown. Here, using our recently established naïve human pluripotent stem cells-derived cytotrophoblast stem cells (nCTs) and a device that can apply arbitrary shear stress to cells, we investigated the impact of shear stress on early-stage trophoblasts. After 72 h of exposure to 10 dyn/cm2 shear stress, nCTs became fused and multinuclear, and mRNA expression of the syncytiotrophoblast (ST) markers, such as glial cell missing 1, endogenous retrovirus group W member 1 envelope, chorionic gonadotropin subunit beta 3, syndecan 1, pregnancy specific beta-1-glycoprotein 3, placental growth factor, and solute carrier family 2 member 1 were significantly upregulated compared to static conditions. Immunohistochemistry showed that shear stress increased fusion index, human chorionic gonadotropin secretion, and human placental lactogen secretion. Increased microvilli formation on the surface of nCTs under flow conditions was detected using scanning electron microscopy. Intracellular cyclic adenosine monophosphate significantly increased under flow conditions. Moreover, transcriptome analysis of nCTs subjected to shear stress revealed that shear stress upregulated ST-specific genes and downregulated CT-specific genes. Collectively, these findings indicate that shear stress promotes the differentiation of nCTs into ST.

Downregulation of PGRMC1 accelerates differentiation and fusion of a human trophoblast cell line.

Mononuclear cytotrophoblasts (CTs) differentiate and fuse to form multinuclear syncytiotrophoblasts (STs), which produce human chorionic gonadotropin (hCG) and progesterone to maintain pregnancy. Impaired differentiation and fusion of CTs to form STs are associated with hypertensive disorders of pregnancy and fetal growth restriction. Progesterone receptor membrane component 1 (PGRMC1) is a multifunctional single transmembrane heme-binding protein. We previously demonstrated that downregulation of PGRMC1 promotes endometrial stromal cell differentiation (decidualization). Here, we explored the role of PGRMC1 in trophoblast differentiation and fusion. PGRMC1 expression was lower in STs than in CTs of first-trimester placental tissues. PGRMC1 expression in BeWo cells (a trophoblast-derived choriocarcinoma cell line) decreased upon dibutyryl-cAMP (db-cAMP)-induced differentiation. Both inhibition and knockdown of PGRMC1 stimulated hCG production in the presence of db-cAMP. Furthermore, a quantitative cell fusion assay we developed revealed that inhibition and knockdown of PGRMC1 enhanced db-cAMP-stimulated cell fusion. Peroxisome proliferator-activated receptor γ (PPARγ) agonists decreased PGRMC1 expression and stimulated the cell fusion in BeWo cells. These findings suggest that downregulation of PGRMC1 expression in part through activation of PPARγ during trophoblast differentiation promotes hCG production and cell fusion for formation and maintenance of placental villi during pregnancy.

Luteal tissue characteristics of Morada Nova ewes with hCG application 7.5 days after the end of estrus synchronization protocol in the breeding season.

Results with the use of hCG after synchronization protocol are still inconsistent, which may vary according to breed, season, day of application and dose of the drug used. The aim of the present study was to evaluate the functionality of luteal tissue and ovarian perfusion after hCG treatment during early luteal phase. Estrus-synchronized ewes were randomly assigned to receive i.m. injection of 300 IU of hCG (G-hCG; n = 40) or 1 mL of saline (G-Control; n = 32) on Day 7.5 after progesterone withdrawal. Ultrasonographic evaluations of the ovaries and ovarian and iliac arteries were performed on Days 7.5, 10.5, 13.5, and 21.5. The accessory corpus luteum (aCL) formation rate was 52.5% for G-hCG. There was interaction (p > 0.05) for treatment (G-hCG and G-Control), days (7.5, 10.5, 13.5 and 21.5) and PD (Pregnant and Non-pregnant) for the variables of biometric characteristics of the corpus luteum B-Mode and Color Doppler on days 7.5, 10.5, 13.5 and 21.5. There was no difference (p > 0.05) for pregnancy rates and mean fetuses per ewe between the treatment groups. It is concluded that the application of hCG 7.5 days after the hormonal protocol in Morada Nova ewes in a breeding season is efficient in inducing aCL formation and increasing luteal tissue biometry. However, there was no effect on pregnancy rate.

Comparison of Steroidogenic and Ovulation-Inducing Effects of Orthosteric and Allosteric Agonists of Luteinizing Hormone/Chorionic Gonadotropin Receptor in Immature Female Rats.

Gonadotropins, including human chorionic gonadotropin (hCG), are used to induce ovulation, but they have a number of side effects, including ovarian hyperstimulation syndrome (OHSS). A possible alternative is allosteric luteinizing hormone (LH)/hCG receptor agonists, including the compound TP4/2 we developed, which remains active when administered orally. The aim was to study the effectiveness of TP4/2 (orally, 40 mg/kg) as an ovulation inducer in FSH-stimulated immature female rats, compared with hCG (s.c., 15 IU/rat). TP4/2 stimulated progesterone production and corpus luteum formation; time-dependently increased the ovarian expression of steroidogenic genes (Star, Cyp11a1, Cyp17a1) and genes involved in ovulation regulation (Adamts-1, Cox-2, Egr-1, Mt-1); and increased the content of metalloproteinase ADAMTS-1 in the ovaries. These effects were similar to those of hCG, although in some cases they were less pronounced. TP4/2, in contrast to hCG, maintained normal LH levels and increased the ovarian expression of the LH/hCG receptor gene, indicating preservation of ovarian sensitivity to LH, and did not cause a sustained increase in expression of vascular endothelial growth factor-A involved in OHSS. Thus, TP4/2 is an effective ovulation inducer that, unlike hCG, has a lower risk of OHSS and ovarian LH resistance due to its moderate stimulating effect on steroidogenesis.

EGR1 Promotes Ovarian Hyperstimulation Syndrome Through Upregulation of SOX9 Expression.

Angiogenesis is strongly associated with ovarian hyperstimulation syndrome (OHSS) progression. Early growth response protein 1 (EGR1) plays an important role in angiogenesis. This study aimed to investigate the function and mechanism of EGR1 involved in OHSS progression. RNA-sequencing was used to identify differentially expressed genes. In vitro OHSS cell model was induced by treating KGN cells with human chorionic gonadotropin (hCG). In vivo OHSS model was established in mice. The expression levels of EGR1, SOX1, and VEGF were determined by Quantitative Real-Time polymerase chain reaction (qRT-PCR), Western blot, immunofluorescence staining, and immunochemistry assay. The content of VEGF in the culture medium of human granulosa-like tumor cell line (KGN) cells was accessed by the ELISA assay. The regulatory effect of EGR1 on SRY-box transcription factor 9 (SOX9) was addressed by luciferase reporter assay and chromatin immunoprecipitation. The ERG1 and SOX9 levels were significantly upregulated in granulosa cells from OHSS patients and there was a positive association between EGR1 and SOX9 expression. In the ovarian tissues of OHSS mice, the levels of EGR1 and SOX9 were also remarkedly increased. Treatment with hCG elevated the levels of vascular endothelial growth factor (VEGF), EGR1, and SOX9 in KGN cells. Silencing of EGR1 reversed the promoting effect of hCG on VEGF and SOX9 expression in KGN cells. EGR1 transcriptionally regulated SOX9 expression through binding to its promoter. In addition, administration of dopamine decreased hCG-induced VEGF in KGN cells and ameliorated the progression of OHSS in mice, which were companied with decreased EGR1 and SOX9 expression. EGR1 has a promoting effect on OHSS progression and dopamine protects against OHSS through suppression of EGR1/SOX9 cascade. Our findings may provide new targets for the treatment of OHSS.