Luteinizing Hormone Receptor Research Articles

Physiological role of dietary energy in the sexual maturity: clues of body size, gonad development, and serum biochemical parameters of Chinese indigenous chicken

Sexual maturity is a crucial factor in the formation and development of poultry reproductive capacity. The nutritional status has been confirmed to play an important role in the regulation of sexual maturity. To investigate the effect of dietary energy levels on sexual maturity in chicken, diets with 3 energy levels (group L: 2,573 kcal/kg, group C: 2,836 kcal/kg, group H: 3,122 kcal/kg) were implemented to feed Guangyuan Gray chickens. During this trial, body weight, body size, organ development, sexual maturity, reproductive performance and blood biochemical parameters were monitored. The earlier sexual maturity was observed in group H, as well as a heavier first egg weight, larger interpubic distance and higher total cholesterol (T-CHO) content at sexual maturity. The dietary energy levels had no significant effect on body weight at first egg and egg production at 300 d of age. Although dietary energy levels had a significant effect on body weight, comb length, tibia length and girth, abdominal fat weight, oviduct weight and length, T-CHO, triglyceride (TG) content and estradiol (E2) level during the rearing period. No significant difference of gonadotropin releasing hormone (GnRH) and luteinizing hormone (LH) level among 3 groups was observed during the trial. The dietary energy levels had effects on mRNA expression of GnRH, estrogen receptor 1 (ESR1), estrogen receptor 2 (ESR2) in hypothalamus, gonadotropin inhibitory hormone receptor (GnIHR) in pituitary and luteinizing hormone receptor (LHR), ESR2 in ovary. The GnIHR/GnRHR ratio in pituitary was higher before sexual maturity and decreased at sexual maturity. The results of correlations analysis found that all the body size, carcass traits, serum biochemical parameters negatively correlated with age at first egg except for interpubic distance and serum blood glucose content. Collectively, dietary energy levels had effects on sexual maturity of chicken, which may be achieved by affecting body weight, gonad development, endocrine and the mRNA expression of genes related to hypothalamus-pituitary-gonad axis. These results further set our understanding of how dietary energy regulates sexual maturity.

Patulin Stimulates Progenitor Leydig Cell Proliferation but Delays Its Differentiation in Male Rats during Prepuberty.

Patulin is a mycotoxin with potential reproductive toxicity. We explored the impact of patulin on Leydig cell (LC) development in male rats. Male Sprague Dawley rats (21 days postpartum) were gavaged patulin at doses of 0.5, 1, and 2 mg/kg/day for 7 days. Patulin markedly lowered serum testosterone at ≥0.5 mg/kg and progesterone at 1 and 2 mg/kg, while increasing LH levels at 2 mg/kg. Patulin increased the CYP11A1+ (cholesterol side-chain cleavage, a progenitor LC biomarker) cell number and their proliferation at 1 and 2 mg/kg. Additionally, patulin downregulated Lhcgr (luteinizing hormone receptor), Scarb1 (high-density lipoprotein receptor), and Cyp17a1 (17α-hydroxylase/17,20-lyase) at 1 and 2 mg/kg. It increased the activation of pAKT1 (protein kinase B), pERK1/2 (extracellular signal-related kinases 1 and 2), pCREB (cyclic AMP response binding protein), and CCND1 (cyclin D1), associated with cell cycle regulation, in vivo. Patulin increased EdU incorporation into R2C LC and stimulated cell cycle progression in vitro. Furthermore, patulin showed a direct inhibitory effect on 11β-HSD2 (11β-hydroxysteroid dehydrogenase 2) activity, which eliminates the adverse effects of glucocorticoids. This study provides insights into the potential mechanisms via which patulin affects progenitor LC development in young male rats.

Correlación entre el polimorfismo de los genes LHCGR y NR5A1y el riesgo de infertilidad masculina

IntroductionInfertility is one of the important phenomena in human reproduction. Genetic factors are the most important cause of male infertility. Here, we aimed to investigate the correlation between idiopathic male infertility and SNPs of the LHCGR (rs2293275) and NR5A1 (rs1057517779) genes in the Iranian-Azeri population. MethodsThis case-control study consisted of 100 males with infertility and 100 healthy males from the Iranian Azeri population. Genomic DNA isolation from whole blood samples and Tetra-primer amplification refractory mutation system-polymerase chain reaction (Tetra-ARMS-PCR) method was used for genotyping. The data analysis was performed by Chi-square (χ2) and Fisher's exact tests. ResultsGenotyping analysis for LHCGR (rs2293275) polymorphism indicated that the frequency of C in the case group was significantly higher than in the control group (P<.05). Moreover, genotyping analysis for NR5A1 (rs1057517779) polymorphism indicated that the frequencies of the A allele and heterozygote GA genotype in the case group were significantly higher than those in the control group (P<.05). ConclusionOur study demonstrated that the SNPs of LHCGR (rs2293275) and NR5A1 (rs1057517779) genes may play a critical role in male infertility in the Iranian Azeri population. However, further studies on other ethnic origins with larger sample sizes are essential for accessing more accurate results. Moreover, functional experiments might be needed to understand the role of these polymorphisms in the molecular pathways involved in male fertility.

Post-trigger luteinizing hormone concentration to positively predict oocyte yield in the antagonist protocol and its association with genetic variants of LHCGR

BackgroundThe concentration of human chorionic gonadotropin (hCG)/ luteinizing hormone (LH) after triggering is generally accepted as a predictor of the normal ovarian response to the trigger, but few studies have explored the distribution model of concentration and its impact on oocyte yield. Genetic variations in LHCGR, known as a receptor for hCG and LH, also play a role in oocyte maturation and retrieval. The objective of the study was to investigate the impact of concentrations of hCG/LH after triggering on oocyte yield and its association with genetic variants of LHCGR.MethodsA retrospective cohort study including 372 antagonist IVF cycles, in which 205 received the recombinant hCG trigger and 167 received the gonadotropin-releasing hormone agonist (GnRH-a) trigger, was conducted. The post-trigger concentrations of hCG/LH and the LHCGR N312S (rs2293275) genotype were evaluated in patients to analyse the impact of these factors on oocyte yield.ResultsThe oocyte retrieval rate (ORR) increased significantly among the low-, medium- and high-hCG-concentration groups (0.91 ± 0.25, 0.99 ± 0.23 and 1.08 ± 0.19, P < 0.001) and among the low-, medium- and high-LH-concentration groups (0.80 ± 0.29, 0.95 ± 0.21 and 1.07 ± 0.19, P < 0.001). The Pearson correlation coefficient between the post-trigger hCG concentration and ORR was 0.242 (P < 0.001), and that between the LH concentration and ORR was 0.454 (P < 0.001). After adjustment for confounding factors, high post-trigger LH concentrations remained associated with the significantly higher ORRs (adjusted R2 = 0.541, P < 0.001). Patients with the AG genotype of LHCGR N312S were more likely to have low post-trigger LH concentrations (46.10 IU/L versus 60.91 IU/L, P < 0.001) and a significantly lower ORR (0.85 versus 0.96, P = 0.042) than patients with the GG genotype after the GnRH-a trigger.ConclusionsThe post-trigger LH concentration can positively predict oocyte yield in antagonist IVF cycles, and patients with the AG genotype of LHCGR rs2293275 could have a suboptimal oocyte yield using the GnRH-a trigger.

Novel Correlation between TGF-β1/-β3 and Hormone Receptors in the Human Corneal Stroma.

This study investigated the interplay between transforming growth factor beta (TGF-β1/T1 and TGF-β3/T3), and sex hormone receptors using our 3D in vitro cornea stroma model. Primary human corneal fibroblasts (HCFs) from healthy donors were plated in transwells at 106 cells/well and cultured for four weeks. HCFs were supplemented with stable vitamin C (VitC) and stimulated with T1 or T3. 3D construct proteins were analyzed for the androgen receptor (AR), progesterone receptor (PR), estrogen receptor alpha (ERα) and beta (ERβ), luteinizing hormone receptor (LHR), follicle-stimulating hormone receptor (FSHR), gonadotropin-releasing hormone receptor (GnRHR), KiSS1-derived peptide receptor (KiSS1R/GPR54), and follicle-stimulating hormone subunit beta (FSH-B). In female constructs, T1 significantly upregulated AR, PR, ERα, FSHR, GnRHR, and KiSS1R. In male constructs, T1 significantly downregulated FSHR and FSH-B and significantly upregulated ERα, ERβ, and GnRHR. T3 caused significant upregulation in expressions PR, ERα, ERβ, LHR, FSHR, and GNRHR in female constructs, and significant downregulation of AR, ERα, and FSHR in male constructs. Semi-quantitative Western blot findings present the interplay between sex hormone receptors and TGF-β isoforms in the corneal stroma, which is influenced by sex as a biological variable (SABV). Additional studies are warranted to fully delineate their interactions and signaling mechanisms.

Synergistic Action of Virgin Coconut Oil and Clomiphene in Reversing Endocrine Dysregulation in Letrozole-Model of Polycystic Ovarian Syndrome in Rats: Role of Nrf2/HMOX-1 Pathway.

Polycystic ovarian syndrome (PCOS) is an endocrine disorder in women's reproductive age. Currently, the pathophysiology of PCOS is unclear, and the limited treatment options are unsatisfactory. Virgin coconut oil (VCO) is functional food oil associated with pharmacological effects in reproductive disorders. Therefore, we aimed to evaluate whether VCO could enhance clomiphene (CLO) therapy against PCOS in female rats. Rats were randomly divided: (1) Control, (2) PCOS model, (3) PCOS + CLO, (4) PCOS + VCO, and (5) PCOS + CLO + VCO. The PCOS was induced via daily letrozole (1 mg/kg, orally) administration for 21 days. After the PCOS induction, CLO, VCO, and CLO + VCO were administered from days 22 to 36. Serum levels of gonadotropin-releasing hormone (GnRH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone, estrogen, progesterone, and prolactin were estimated. Polymerase chain reaction gene expression for nuclear factor-erythroid-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), catalase (CAT), glutathione reductase (GSR), LH receptor (LHr), androgen receptor (AR), tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), and caspase-3 were analyzed. The letrozole-induced PCOS caused considerable increases in GnRH, LH, prolactin, estrogen, and testosterone, whereas FSH decreased significantly compared to the control. The gene expression of Nrf2, HO-1, CAT, and GSR were markedly diminished, while IL-1β, TNF-α, caspase-3, AR, and LHr prominently increased compared to control. Interestingly, the CLO and VCO separately exerted anti-inflammatory and endocrine balance effects. However, VCO-enhanced CLO effect in LH, prolactin and testosterone, Nrf2, HO-1, CAT, GSR, and AR. VCO may synergize with CLO to depress hyperandrogenism and oxidative inflammation in PCOS.

Steroidogenic Effect of Luteinizing Hormone Receptor Agonists and Metformin in Male Rats with Androgenic Deficiency Caused by Diet-Induced Obesity

Steroidogenic Effect of Luteinizing Hormone Receptor Agonists and Metformin in Male Rats with Androgenic Deficiency Caused by Diet-Induced Obesity

Influence of metformin therapy and treatment with an allosteric luteinizing hormone agonist and chorionic gonadotropin on spermatogenesis in male rats with obesity

BACKGROUND: In men, obesity is accompanied by a complex of metabolic and hormonal disorders, which leads to androgen deficiency and impaired spermatogenesis. Antidiabetic drugs, including metformin (MF), and luteinizing hormone receptor (LHR) agonists, which activate testicular steroidogenesis, can be used to correct reproductive dysfunctions. However, in diet-induced obesity (DIO), their effectiveness and mechanisms of action are poorly understood.AIM: In men, obesity is accompanied by a complex of metabolic and hormonal disorders, which leads to androgen deficiency and impaired spermatogenesis. Antidiabetic drugs, including metformin (MF), and luteinizing hormone receptor (LHR) agonists, which activate testicular steroidogenesis, can be used to correct reproductive dysfunctions. However, in dietinduced obesity (DIO), their effectiveness and mechanisms of action are poorly understood.MATERIALS AND METHODS: Obesity in male Wistar rats was induced by a 23-week diet enriched with saturated fats. MF treatment was carried out for 5 weeks at a dose of 120 mg/kg/day (orally), and the treatment with hCG and TP03 was carried out for 5 days at daily doses of 20 IU/rat (s.c.) and 15 mg/kg (i.p.), respectively. Using microscopy and histochemical analysis, the number and motility of spermatozoa (SP), the number of their defective forms and the morphology of the seminiferous tubules were assessed, and the levels of testosterone and other hormones in the blood were measured using ELISA.RESULTS: MF, hCG, and TP03 to varying degrees increased the number of SP and the proportion of their mobile forms, including those with forward movement, which were reduced in DIO rats, and also normalized the thickness of the epithelium of the seminiferous tubules and the number of spermatogonia and pachytene spermatocytes in them, but did not reduced the proportion of defective forms of SP, increased in DIO. In the case of MF, this was associated with the drug-induced normalization of body weight, glucose tolerance, and the insulin and leptin levels in DIO rats. The positive effect of hCG and TP03 on spermatogenesis was due to their stimulating effect on testosterone production.CONCLUSION: The use of long-term MF therapy and short-term courses of LHR-agonists normalizes impaired spermatogenesis in DIO, which indicates the prospects for their use to improve male fertility in obesity, and in the case of MF therapy, normalization of the metabolic and hormonal status is of great importance, while in the case of LHR-agonists the most important factor is their steroidogenic effect.

MiR-143-3p Promotes Ovarian Granulosa Cell Senescence and Inhibits Estradiol Synthesis by Targeting UBE2E3 and LHCGR.

The ovary is a highly susceptible organ to senescence, and granulosa cells (GCs) have a crucial role in oocyte development promotion and overall ovarian function maintenance. As age advances, GCs apoptosis and dysfunction escalate, leading to ovarian aging. However, the molecular mechanisms underpinning ovarian aging remain poorly understood. In this study, we observed a correlation between the age-related decline of fertility and elevated expression levels of miR-143-3p in female mice. Moreover, miR-143-3p was highly expressed in senescent ovarian GCs. The overexpression of miR-143-3p in GCs not only hindered their proliferation and induced senescence-associated secretory phenotype (SASP) but also impeded steroid hormone synthesis by targeting ubiquitin-conjugating enzyme E2 E3 (Ube2e3) and luteinizing hormone and human chorionic gonadotropin receptor (Lhcgr). These findings suggest that miR-143-3p plays a substantial role in senescence and steroid hormone synthesis in GCs, indicating its potential as a therapeutic target for interventions in the ovarian aging process.

Identification of genomic regions associated with reproductive longevity in the Rubia Gallega beef cattle breed using a censored threshold model

In beef cattle, the economic viability of farms is heavily influenced by the cow's ability to survive subsequent pregnancies. To understand the genetic basis of reproductive longevity in Rubia Gallega beef cattle breed, a ssGWAS was performed by back-solving the output of ssGBLUP under a censored threshold model. It considers the number of parities each cow reaches during its productive life as a phenotypic trait. The results of the study showed that the main source of variation of reproductive longevity is the herd. However, the posterior mean of the heritability of reproductive longevity was 0.173, indicating the potential for an appropriate genetic response to selection. Furthermore, it is shown that four genomic regions in chromosomes 2, 11 and 29 explain a large proportion of the additive genetic variance. The most important signal was detected on chromosome 2 in the vicinity of the MSTN (myostatin) gene that is associated with double muscling, and that it is segregating in the Rubia Gallega population. Some other interesting genes located within these regions encoded for several PAGs (Pregnancy-associated glycoprotein), LHCGR (luteinizing hormone/choriogonadotropin receptor), FSHR (follicle stimulating hormone receptor), PROKR1 (prokineticin receptor 1) and EHD3 (Eps15 homology domain-containing protein 3). This confirms the relevance of the reproductive performance in the reproductive longevity of cows. These findings provide valuable insights for the Rubia Gallega breeding program, as they can be used to define future selection strategies to improve reproductive longevity of the breed.

The Regulatory Effect of Eugenol on FSHR, LHCGR, and ER Expression during Follicular Development in Female Rats With Ovarian Torsion

Objectives: This study aimed to investigate the role of Eugenol in regulating the expression of FSH receptor (FSHR), human luteinizing hormone choriogonadotropin receptor (LHCGR), and estrogen receptor (ER) during follicular development in female rat ovarian torsion. Materials and Methods: In this experimental study, 48 female rats were randomly assigned to 4 groups, including G1 (i.e., sham), G2 (i.e., ovarian torsion/detorsion group), G3 (i.e., ovarian torsion/detorsion group treated with 30 mg/kg of eugenol), G4 (i.e., healthy group treated with 30 mg/kg of eugenol). After covering a treatment period of ten days, the ovarian tissue was collected for the histological analysis, the measurement of ER, FSHR, and LHCGR expression, as well as the assessment of testosterone, LH, FSH, and estrogen levels in blood serum. Results: Histological evaluation revealed the damage to ovarian tissue, the reduced oocyte, and the granulosa cell diameter in the torsion/detorsion group. However, the treatment with eugenol mitigated this damage. Eugenol administration increased the levels of estrogen, LH, and FSH, but it decreased the testosterone levels in the treated group. Moreover, the expression of ER, FSHR, and LHCGR was upregulated in the treated groups. Administration of eugenol was associated with an enhanced fertility. Conclusions: It was concluded that eugenol administration may have been effective in protecting the ovarian tissue from the damage caused by torsion/detorsion. Furthermore, eugenol was found to have the potential to modulate hormonal profiles and regulate the expression of ER, FSHR, and LHCGR, thereby contributing to an increased fertility.

Whole exome sequencing identifies a novel homozygous missense mutation of LHCGR gene in primary infertile women with empty follicle syndrome.

The genetic basis of empty follicle syndrome (EFS) is largely unknown, and the aim of this study was to investigate the genetic causes of EFS in primary infertile women. Four affected women diagnosed with anovulation were recruited, and whole exome sequencing (WES) was requested for the genetic diagnosis of the cases. One hundred healthy controls were verified by Sanger sequencing. A novel homozygous variant of the LHCGR gene (NM_000233:c.1847C>A) was revealed in one affected individual by WES. Trios analysis of the mutation revealed an autosomal recessive pattern. This LHCGR variant was absent in 100 healthy controls and predicted to be highly damaging to the function of LHCGR. The novel variant extends the mutational spectrum of the LHCGR gene associated with female sterility, which promotes the prognostic value of testing for LHCGR mutations in infertile women with EFS.

Effect of an additional GnRH 2 d after the initiation of the Resynch-25 program on pregnancy per artificial insemination in lactating dairy cows

Timed-AI protocols for the reinsemination of nonpregnant cows significantly reduce the interval between inseminations (AI) in lactating dairy cows. One of the most used programs is the Resynch-25, which starts Ovsynch on d 25 after previous AI in all cows, and only nonpregnant cows continue with the pro­gram on d 32 after previous AI. Low ovulatory response to d 25 GnRH administration can compromise the proportion of cows synchronized to the program and pregnancy per AI (P/AI) after timed-AI. One of the factors that can affect ovulatory response to this d 25 GnRH is the stage of antral follicle development at the time of treatment. For instance, non-dominant follicles can­not ovulate due to a lack of LH receptors in the granulosa cells. Thus, we hypothesized that adding an extra GnRH 2 d after d25 GnRH would increase P/AI to the Resynch-25 program. Our ob­jective was to determine the effect of an additional GnRH 2 d after initiating the Resynch-25 program on P/AI.

Myosin VI stabilizes intercellular junctions in the testis through the LHR and MAPK signalling pathway during spermatogenesis in Eriocheir sinensis

The myosin motor protein myosin VI plays an essential role in mammalian spermatogenesis, however, the effects of myosin VI on male reproduction in Crustacea remain obscure. We identified the macromolecule es-Myosin VI in Eriocheir sinensis, and studied it by multiple methods. It co-localized with F-actin and was highly expressed in the testis. We interfered es-Myosin VI using dsRNA in vivo, an apparent decrease in spermatozoa count was detected. We also found that the MAPK signalling pathway was changed, subsequently causing disruption of intercellular junctions and damage to the functional hemolymph–testis barrier. We observed that luteinizing hormone receptor es-LHR was located within seminiferous tubules, which was different from the expression in mammals. Es-LHR could bind with es-Myosin VI in testis of E. sinensis, its localization was significantly altered when es-Myosin VI was deleted. Moreover, we obtained consistent results for the MAPK signalling pathway and spermatogenesis defects between the es-LHR and es-Myosin VI knockdown groups. In summary, our research demonstrated that knockdown of es-Myosin VI disturbed the intercellular junction and HTB function via the MAPK signalling pathway by changing the localization of es-LHR in the testis of E. sinensis, which was the potential reason for its negative impact on spermatogenesis.

A delayed ovulation of progestin-primed ovarian stimulation (PPOS) by downregulating the LHCGR/PGR pathway

Progestin-primed ovarian stimulation (PPOS) is a new ovulation stimulation protocol, and its role in ovulation and regulatory mechanism is unclear. The clinical PPOS protocol was simulated in mice. The ovulated oocytes, estradiol, progesterone, and luteinizing hormone (LH) levels were analyzed at different hours after trigger. mRNA extraction and real-time PCR, hematoxylin and eosin staining, and immunofluorescence of ovaries were used to explore the involved signaling pathways. The PPOS group had a delayed ovulation at 12.5h after trigger. Its suppressed LH level reduced the expression of luteinizing hormone/choriogonadotropin receptor (LHCGR) on the preovulatory follicles before trigger and significantly decreased the following progesterone synthesis, blood progesterone level, and progesterone receptor (PGR) expression within 4-6h after trigger. Furthermore, the important ovulatory genes regulated by PGR including ADAMTS-1, VEGF-A, and EDN2 were downregulated, ultimately delaying the ovulation. PPOS suppresses the LH level before trigger and decreases the synthesis of progesterone after trigger, thus delaying the ovulation by downregulating the LHCGR-PGR pathway.

P-698 Genetics and reproductive health outcomes: An Asian perspective

Abstract Study question Does genetics play a role in the reproductive outcomes in women undergoing assisted reproductive technology (ART)? Summary answer We describe a trend towards poorer reproductive outcomes associated with Asian women and ART. Subsequently, we posit a potential biological mechanism. What is known already In the last four decades, advances in assisted reproductive technology (ART) have afforded the possibility of conceiving a child to individuals who encounter fertility complications. However, a closer examination of the clinical outcomes of ART shows a stark contrast in Asian women compared to Caucasians, with the majority of studies reporting lower reproductive success among Asian women. In this study, a systematic review was performed to elucidate the genes associated with ART clinical outcomes, with a focus on Asian ethnicities. Concomitantly, we highlight the need for personalized medicine to bridge the gaps in female reproductive health. Study design, size, duration We performed a Pubmed and Embase database search, from inception to December 2022, to identify all studies which investigated the differential reproductive outcomes in Asian women compared to their Caucasian counterparts, genetic mutations contributing to fertility in Asian women and personalized medicine. Following the PRISMA workflow, the preliminary search yielded a total of 2,145 papers to be reviewed. Participants/materials, setting, methods Following the inclusion/exclusion criteria screening, 128 studies were analyzed (ART outcomes in Asians, n=75; Genetics on women’s reproductive health, n=38; Personalized medicine in Asians, n=15). As each research paper identified one or more genes/SNPs involved in female fertility and reproductive health, we took a step further and performed a pathway analysis of gene-sets using STRING in Cytoscape v3.4.0. Network analysis and biological process associations were performed using GSEA and MSigDB. Significant gene-sets were annotated if FDR&amp;lt;0.05. Main results and the role of chance We observed that age at menarche(AAM) was found to be correlated with the timing of the first pregnancy, with Hawaiians having the lowest age(22.2yrs) and Japanese women having the highest age(25.0yrs). LIN28 mutations were highly associated with AAM and prevalent in both Chinese and American populations. FMR1 was most associated with ovarian reserve, with BMP15, ESR1, INHA, PRIM, TMEM150B to a lesser extent. Finally, FSHR polymorphisms were found to affect IVF outcomes most significantly. Network analysis highlighted a close association between 5 genes; FMR1, FSHR, ESR1, BMP15 and INHA, through biological functions of ovarian follicle development, menstrual cycle and hyperpituitarism. Network analysis revealed 4 major biological pathways (ovarian follicle development, oocyte maturation, ovulation, steroidogenesis), as well as key proteins which are involved in female fertility and reproductive health outcomes. PI3K/Akt are critical regulators of ovarian function through primordial follicle maturation and granulosa cell proliferation. MAPK signaling is involved with gonadotrophin-releasing hormone which controls FSH and LH secretion while Gq/11 signaling is activated through LH receptor. Therefore, PI3K/Akt, MAPK and Gq/11 protein signaling pathway appear to be key components to a broad range of reproductive outcomes and represent potential pathways to target in future studies investigating ovarian, follicular and steroidogenesis dysfunction. Limitations, reasons for caution A large majority of the papers reviewed relied on the self-reporting of ethnicities, which can vary considerably in accordance to how an individual defines his or her own ethnicity. Hence, we could not exclude the possibility of reporting bias in the studies. Wider implications of the findings Leveraging these findings,we propose the development of a biomarker panel which would enhance patient stratification,moving towards personalized medicine to address every woman’s unique reproductive potential.With several countries reporting low fertility rates due to shifts in socio-economic and life priorities, this has strong implications for those who wish to delay childbearing. Trial registration number Not applicable

P-667 The attenuated response to elevated serum progesterone levels on pregnancy outcomes of fresh IVF/ICSI cycles in older patients

Abstract Study question Whether there are different impacts of serum progesterone (P) levels on the day of trigger on pregnancy outcomes in patients of different age groups. Summary answer Our results suggest that the pregnancy outcome was less sensitive to progesterone elevation in older patients. What is known already It is now generally believed that the P elevation at the late follicular phase reduces the pregnancy rate by opening the implantation window prematurely and disrupt the embryo-endometrial synchrony. Previous studies further investigated the effect of developmental stages of embryo, number of oocytes retrieved, cause of infertility and different stimulation regimens on the association of P level and the likelihood of pregnancy. However, the study of maternal age on such association is scarce. A previous study found that LH receptor (LHCGR) and progesterone receptor (PGR) were up regulated in advanced age group. Study design, size, duration This was a non-interventional, retrospective cohort study of patients undergoing routine practice in a university-based fertility center between 2015 and 2021. One thousand five hundred and seventeen IVF/ICSI cycles among 1305 patients were included in the analysis. Participants/materials, setting, methods Fresh ET using autologous oocytes with serum P level measured on the day of hCG administration were included. Exclusion criteria include cycles without P measurement, using frozen oocytes, PGT cycles, freeze-all cycles, oocyte donation and no embryo transfer cycles. The patient age was divided into 3 groups: group 1 included patients at age 35 or below, group 2 included patients at age 36 to 39, group 3 included those at 40 years old or above. Main results and the role of chance We performed logistic regression analysis for each group to determine the contributing factors to the live birth outcome. In group 1, age, serum P level, number of usable embryos, embryo grading and endometrial thickness were found to be significantly associated with live birth. In group 2, age and number of usable embryos were significantly associated with live birth. However, total dose of gonadotropin was found to be the sole contributing factor to live birth in group 3. We also divided the P levels into low (≤0.50 ng/mL), median (0.51-1.00 ng/mL) and high (&amp;gt;1.00 ng/mL) and compared the live birth rate in each age groups. The live birth rate was found to be significantly reduced with increasing P levels (40.3% vs. 35.5% vs. 23.6%, p = 0.005). No significant difference in live birth rate between different P levels was found in age group 2 and 3. Limitations, reasons for caution The limitation of the study is its retrospective nature which may have led to bias in the interpretation of the data. There were also significantly higher proportion of cleavage-stage embryo to blastocyst transfer in the older group. Wider implications of the findings The negative association of premature progesterone rise at pre-ovulatory phase to the IVF fresh ET outcome was more prominent for younger patients. This may be one of the contributing factors that accounts for the great heterogeneity in the cut-off of P and its predicting power in different studies. Trial registration number Not applicable

Association of A51703G and G51656T loci polymorphism in luteinizing hormone receptor (LHR) gene with sperm traits in Holstein bulls

Today, there is an increasing desire to use Candidate genes as molecular markers to improve reproductive traits. The luteinizing hormone receptor is expressed on the Leydig cells and plays a crucial role in gametogenesis initiation and maintenance. This study aimed to study the effects of luteinizing hormone receptor (LHR) gene polymorphisms (A51703G and G51656T) on bull sperm traits. Genomic DNA was extracted from the semen, blood, and samples of 150 AI Iranian Holstein bulls according to the established protocol [10] with minor modifications. Semen traits were routinely collected at bull semen collection centers from each animal between 1992 and 2015. The bull fertility traits include semen concentration (SC 108 per ml) sperm volume (SV ml), fresh sperm motility (FSM %), and post-tallow sperm motility (PTSM %). The mutation was detected with A PCR-RFLP method. The digestion of amplicons was also carried out with BfaI and HinfI restriction enzymes in A51703G and G51656T loci, respectively. The restriction profiles of PCR products indicated two alleles in each locus (A and G for A51703G and G and T for G51656T). The frequency of the A and T alleles were 0.6647 and 0.606, respectively. Analysis of variance showed that LHR loci had a significant effect on sperm characteristics in Iranian Holstein bulls (P < 0.05). Bulls with GG genotype surpassed those with GA and AA genotypes regarding SP, FSM, and PTSM in the A51703G locus. In the G51656T locus, the association result showed that bulls with AG genotype had higher than others in all traits (P < 0.05). In conclusion, these data suggest that LHR polymorphisms are an excellent genetic marker for breeding programs in male reproductive traits in the Iranian bull population.

Identification polymorphism of LHR and FSHR genes in Indonesian Holstein dairy cattle associated with productive and reproductive traits

Abstract. Setyorini YW, Kurnianto E, Sutopo, Sutiyono. 2023. Identification polymorphism of LHR and FSHR genes in Indonesian Holstein dairy cattle associated with productive and reproductive traits. Biodiversitas 24: 2898-2905. Luteinizing and follicle-stimulating hormones and their receptors play an important role in the reproductive system’s hormonal activity and physiological function. This study aimed to identify the genotype variation of the Luteinizing Hormone Receptor (LHR) gene exon 11 and the Follicle-stimulating Hormone Receptor (FSHR) gene exon 10 and to elucidate the associating between polymorphism with the milk and reproductive traits in dairy cattle. A total of 100 samples of deoxyribonucleic acid (DNA) from Indonesian Holstein dairy cattle were used in this study. Polymerase Chain Reaction Restriction Fragment Length Polymorphism (PCR-RFLP) was performed for genotyping of the Single Nucleotide Polymorphism (SNP) of the LHR and FSHR genes with the HhaI and AluI restriction enzymes, respectively. The collected phenotype data were milk production, quality, and reproductive trait. In addition, the general linear model procedure in the SAS program was carried out to investigate the association of genotypes with productive and reproductive traits. The results showed that the LHR gene in the population of this study was monomorphic, while the FSHR gene was polymorphic with three genotypes, namely CC, CG, and GG. The sequences of the FSHR gene indicated the presence of a mutation at nucleotide number 2037 that substitutes cytosine for guanine. Cows with the CC genotype showed better parameter service per conception (P&lt;0.05) than the other genotypes but did not correlate with other reproductive parameters and milk traits. Therefore, it was concluded that SNP g.2037C&gt;G polymorphism at FSHR gene exon 10 was identified in the Indonesian Holstein Dairy cattle population and associated with service per conception parameters but not with milk trait.

Endocrine changes induced by GnRH immunization and re-stimulation of testicular function with a GnRH agonist in stallions

Resumption of testicular function after GnRH immunization varies among animals and some stallions regain fertility only after a prolonged time (Janett et al., 2009, Anim Reprod Sci 115:88e102). This study evaluated endocrine effects of GnRH immunization and subsequent re-stimulation with a GnRH agonist. We hypothesized that GnRH agonist treatment advances resumption of normal endocrine function in GnRH-vaccinated stallions. Shetland stallions were assigned to a GnRH vaccination group (Improvac; 400 µg/animal in weeks 0 and 4; n=6) or a saline control group (n=6). Blood was collected weekly for hormone analyses. Each vaccinated stallion was hemi-castrated together with an age-matched control when testosterone concentration was <0.3 ng/ml for two consecutive weeks (C1). Treatment of vaccinated stallions with the GnRH agonist buserelin (4 µg/day for 4 weeks and 8 µg/day for 6 weeks) started three weeks thereafter. Irrespective of treatment, the remaining testicle was removed when testosterone concentration increased to >0.5 ng/ml for two consecutive weeks in vaccinated stallions (C2; 9-10 weeks after start of buserelin treatment). Testicular tissues were stored at – 80°C and fixed in formalin for RT-qPCR and histology. Statistical comparisons were made by repeated measures ANOVA with group as between and time as within subject factor. Data on semen characteristics in these stallions have been published recently (Gautier et al., 2022, Theriogenology 194:27-34). Plasma testosteroneconcentration decreased after GnRH-vaccination and returned to control stallion concentrations during buserelin treatment (time p<0.001, time x group p<0.001, group p=0.001). Serum LH was undetectable after the second vaccination, increased during buserelin treatment but remained lower than in control stallions (time p=0.003, time x group p<0.001). Serum FSH concentration decreased after vaccination and increased with buserelin treatment in vaccinated but even more in control stallions after both hemi-castrations (time p<0.001, time x group p<0.001, group p=0.008). Relative mRNA abundance for LH receptor (LH-R) and aromatase (CYP19A1) were lower in testicular tissue of vaccinated than control stallions at C1 (p<0.001). At C2, LH-R mRNA abundance no longer differed between groups, whereas CYP19A1 mRNA remained lower in vaccinated than in control stallions (p<0.001). Relative FSH receptor (FSH-R) mRNA abundance was higher in vaccinated than in control stallions at C1 (p<0.001) and similar in both groups at C2. Immunolabelling for LH-R was lower in testes of GnRH-vaccinated compared to control stallions at C1 and C2 (p=0.007), whereas FSH-R protein expression at no time differed between groups. Aromatase immunostaining was reduced at C1 in vaccinated stallions and increased until C2 (time p=0.035, time x group p<0.0031, group p<0.001). In conclusion, endocrine testicular function can be advanced in GnRH-vaccinated stallions by daily low dose buserelin treatment.