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This study characterized the physicochemical properties and predicted the structure, function, and protein-protein interactions of bone morphogenetic protein 15 (BMP15) using bioinformatics tools. BMP15 was found to have a molecular weight of 45055.01 Daltons, sequence length of 392 amino acids, and an extinction coefficient of 55390 at 280 nm with a basic isoelectric point. Secondary structure analysis revealed BMP15 consists mostly of random coil (63.78%), followed by alpha helix (20.66%) and extended strand (15.56%) as well as beta turns. Amino acids with high coil structure like glycine and alanine, which are hydrophobic and flexible, represented the highest concentrations. Transmembrane helix prediction identified four helices located from inside to outside and three from outside to inside. SWISS-MODEL generated four protein structure models corresponding to sequences (Q6PX77.1. A, 5vqf.2. A, 5ntu.1. A, and 5hly.1. A) with sequence identities of (75.38%, 20.83%, 20.77% and 20.33%) respectively. Results correlate BMP15 with oocyte maturation and granulosa cell activation in follicular development. This comprehensive bioinformatics analysis of BMP15 properties, structure, and interactions provides a framework for further study of genetically inherited infertility, drug design, and new protein analysis.

Polymorphisms identified on the BMPR1B, BMP15, and GDF9 genes tend to increase multiple birth and ovulation rates in sheep. In the planned study, the productivity records of the flocks (approximately 41.000) collected from 2016 to 2022, belonging to the Public Breeding of Kangal Akkaraman Sheep in Sivas province TAGEM/58KAK2012-08 subproject of the Public Animal Breeding National Project, were used. Accordingly, the ear numbers of sheep that gave birth to at least two twins (n=96) and at least two singletons (n=96) were determined from 15 different farms relevant records. According to similar feeding characteristics, environmental variables were grouped as location, year-round feeding type, and seasonal feeding type. DNA groups were genotyped by the PCR-RFLP method for BMPR1B (FecB), BMP15 (FecXB, FecXG, FecXI, FecXH) and GDF9 (FecG) alleles. Accordingly, among the 6 SNPs examined, only the GDF9 gene-FecB SNP was determined polymorphic. Genotypic effect (FecB allele) and environmental effect variables (location, year-round feeding type, seasonal feeding type ) were also examined with a logistic model. It was determined that the relevant alleles and environmental variables did not have a statistically significant effect on the twinning phenotype. According to the results obtained, it was thought that the genes associated with multiple births in Kangal Akkaraman breed may have different variants specific to the breed. In addition, it is suggested that this character, which is affected by multiple genes, should be included in the planned breeding studies by considering the interaction of environmental variables and determining the variation of the related genes. In this respect, it is concluded that our study will guide the sequencing studies and multivariate analyses to be planned. [ABSTRACT FROM AUTHOR]

Polymorphisms identified on the BMPR1B, BMP15, and GDF9 genes tend to increase multiple birth and ovulation rates in sheep. In the planned study, the productivity records of the flocks (approximately 41.000) collected from 2016 to 2022, belonging to the Public Breeding of Kangal Akkaraman Sheep in Sivas province TAGEM/58KAK2012-08 subproject of the Public Animal Breeding National Project, were used. Accordingly, the ear numbers of sheep that gave birth to at least two twins (n=96) and at least two singletons (n=96) were determined from 15 different farms relevant records. According to similar feeding characteristics, environmental variables were grouped as location, year-round feeding type, and seasonal feeding type. DNA groups were genotyped by the PCR-RFLP method for BMPR1B (FecB), BMP15 (FecXB, FecXG, FecXI, FecXH) and GDF9 (FecG) alleles. Accordingly, among the 6 SNPs examined, only the GDF9 gene-FecB SNP was determined polymorphic. Genotypic effect (FecB allele) and environmental effect variables (location, year-round feeding type, seasonal feeding type ) were also examined with a logistic model. It was determined that the relevant alleles and environmental variables did not have a statistically significant effect on the twinning phenotype. According to the results obtained, it was thought that the genes associated with multiple births in Kangal Akkaraman breed may have different variants specific to the breed. In addition, it is suggested that this character, which is affected by multiple genes, should be included in the planned breeding studies by considering the interaction of environmental variables and determining the variation of the related genes. In this respect, it is concluded that our study will guide the sequencing studies and multivariate analyses to be planned.

Unexpected poor ovarian response (UPOR) occurs when nine or fewer oocytes are retrieved from a young patient with normal ovarian reserve. Bone morphogenetic protein15 (BMP15) and growth differentiation factor 9 (GDF9) are two oocyte-specific factors with pivotal role in folliculogenesis. The aim of this study was to assess the relation between BMP15 and GDF9 variants with UPOR. Hundred women aged ≤ 39 with AMH ≥ 1.27 IU/ml participated as UPOR and normal ovarian responders (NOR) based on their oocyte number. Each group consisted of 50 patients. After genomic DNA extraction, the entire exonic regions of BMP15 and GDF9 were amplified and examined by direct sequencing. Western blotting was performed to determine the expression levels of BMP15 and GDF9 in follicular fluid. Additionally, in silico analysis was applied to predict the effect of discovered mutations. From four novel variants of BMP15 and GDF9 genes, silent mutations (c.744 T > C) and (c.99G > A) occurred in both groups, whereas missense variants: c.967-968insA and c.296A > G were found exclusively in UPORs. The latter variants caused reduction in protein expression. Moreover, the mutant allele (T) in a GDF9 polymorphism (C447T) found to be more in NOR individuals (58% NOR vs. 37% UPOR (OR = 2.3, CI 1.32–4.11, p = 0.004). The novel missense mutations which were predicted as damaging, along with other mutations that happened in UPORs might result in ovarian resistance to stimulation. The mutant allele (T) in C447T polymorphism has a protective effect. It can be concluded that there is an association between BMP15 and GDF9 variants and follicular development and ovarian response. [ABSTRACT FROM AUTHOR]

Summary: At this time, with advances in medical science, many cancers and chronic diseases are treatable, but one of their side effects is infertility. Some women also want to delay pregnancy for personal reasons. There has been some evidence that kisspeptin activates broad signals by binding to its receptor, suggesting that the role of kisspeptin in direct control of ovarian function includes follicle growth and steroid production. In this study, the effect of kisspeptin on improving the quality and results for human ovarian follicles was investigated. A section of ovary was removed laparoscopically from women between 20 and 35 years of age (n = 12). Pieces were divided randomly into two groups, control and treatment (with 1 μM kisspeptin). Real-time PCR was performed for GDF9 , BMP15 and mTOR gene expression assessments. Western blotting was carried out to measure AKT and FOXO3a protein expression. Data were analyzed using one-way analysis of variance (ANOVA) and Tukey's test; means were considered significantly different at a P -value < 0.05. During treatment with the kisspeptin group, maturity genes are expressed. Therefore, kisspeptin is an effective substance to improve the quality of the human ovarian medium as it increases the maturity of follicles. [ABSTRACT FROM AUTHOR]

Premature ovarian insufficiency (POI) is defined as the development of hypergonadotropic hypogonadism before the age of 40 with definitive treatment being absent. In the current study, we aim to compare the efficacy of the cell sheet method with an intravenous (IV) application of adipose-derived mesenchymal stem cells (AdMSCs) to the POI with an animal model. In the current prospective study, 6-to-8-week-old Sprague Dawley rats were generated four groups: (i) a control group in which only PBS was administered; (ii) an only-POI group generated by cyclophosphamide; (iii) a POI group treated by way of IV AdMSCs; and (iv) a POI group treated by way of the cell sheet method. Twenty-eight days after an oophorectomy was performed, intracardiac blood was taken. Follicle count, immunohistochemical examination for GDF9, BMP15, and TUNEL were conducted, gene expressions of GDF9 and BMP15 were examined, and E2 was measured in the serum samples. With hematoxylin-eosin, in the third group, multi oocytes follicles were the most remarkable finding. In the fourth group, most of the follicles presented normal morphology. GDF9 involvement was similar between the first and fourth groups. BMP-15 immunoreactivity, in contrast to fourth group, was weak in all stages in the second and third groups. The current attempt represents a pioneer study in the literature in which a cell sheet method is used for the first time in a POI model. These results suggest that the cell sheet method may be a feasible and efficient method for the stem cell treatment of models with POI and could be a new treatment approach in POI. [ABSTRACT FROM AUTHOR]

Background: Kisspeptin is a neuropeptide that has an important role in the female reproductive cycle which is indicated by its role in regulating the hypothalamic-pituitary-gonadal axis (HPG axis). Aims: To analyze the correlation between serum kisspeptin levels, ovarian kisspeptin expression, and ovarian BMP15 expression in PCOS model rats. Methods: The research was true experimental research with post-test design only control group, was carried out from August to October 2022 at the Faculty of Veterinary Medicine Universitas Airlangga. 32 Rattus novergicus rats were divided into a control group and PCOS model group. Blood serum and ovaries were obtained from all groups. Blood serum was examined for kisspeptin levels by ELISA technique, kisspeptin expression, and BMP15 Ovaries were examined immunihistochemically. Results: Serum kisspeptin levels and ovarian kisspeptin expression of the PCOS model group were not significantly higher than those of the control group (p>0.05, p>0.05). The ovarian BMP15 expression of the PCOS model group was not significantly lower (p>0.05) than that of the control group. Ovarian kisspeptin expression and ovarian BMP15 expression did not have a significant correlation with serum kisspeptin levels (p>0.05). In contrast, there was a significant correlation (p [Open Vet J 2023; 13(3.000): 288-296]

Premature ovarian insufficiency (POI) is a rare cause of primary amenorrhea in adolescents. For young women with uncertain etiology of POI, genetic and autoimmune testing may be recommended to assist in treatment and management decisions. This report presents a case of POI in a 16-year-old adolescent with both poly-autoimmune disease and a heterozygous missense variant in the bone morphogenic factor 15 (BMP15) gene, both potentially involved in the pathogenesis of POI. Accurately distinguishing between autoimmune and genetic causes is crucial for effective treatment and counseling. In addition, given the significant psychological impact and the need for reproductive options counseling, a multidisciplinary approach that includes psychological support is highly recommended. [ABSTRACT FROM AUTHOR]

The general twinning rate of the Karayaka breed, which stands out with its meat quality, ranges from 0.10 to 0.20. However, it may be encountered in the flocks with higher twinning rates. The aim of this study was to investigate five different mutations in the BMPR1B (FecB) and BMP15 (FecXH, FecXI, FecXG, and FecXB) genes. The mutations were screened directly in 100 ewes of the Karayaka breed raised in the Black Sea region of Türkiye using the restriction fragment length polymorphism (PCR-RFLP) technique. The study results showed that all the ewes examined had wild-type alleles or carried no mutations. In conclusion, the monomorphic structure of the BMPR1B and BMP15 genes concerning the loci studied suggests that these genes cannot be used as candidate genes for increasing twin births in the Karayaka breed. The findings in this study are the first for the Karayaka breed, and there is a need for comprehensive studies based on phenotypic data with more samples and other candidate genes. [ABSTRACT FROM AUTHOR]

Background: As BMP15, GDF9, and kisspeptin all play critical roles in folliculogenesis and fertilization, investigating the possible relationship between obesity and these three factors could prove crucial in relation to understanding the role of obesity in infertility. Thus, the present study sought to determine the effects of obesity on the serum BMP15, GDF9, and kisspeptin concentrations in women of reproductive age. Methods: Ninety female participants were equally divided into three groups: class-1 obese (n=30), class-2 obese (n=30), and normal weight (control; n=30). The participants' serum BMP15, GDF9, and AMH concentrations were measured. Moreover, the serum kisspeptin concentrations were evaluated in the class-1 obese and control groups by means of the enzyme-linked immunosorbent assay (ELISA) method while the participants were in their menstrual period. Results: The serum BMP15 and kisspeptin concentrations were found to be much higher in the control group than in both obese groups (p= 0.001 and p=0.01, respectively). While the GDF9 concentration exhibited a statistically significant positive correlation with age, the BMP15 concentration exhibited a positive correlation with the kisspeptin and LH concentrations in the control group. In addition, a positive correlation was identified between the BMP15 concentration and both age and the glucose level and a negative correlation with the insulin level in both the obese groups. Conclusions: Obesity appears to reduce the serum BMP15 and kisspeptin concentrations in obese women of reprodučtive age. This reduction may represent a milestone in reproductive dysfunction and may be used to predict the success of infertility treatment in obese women. [ABSTRACT FROM AUTHOR]

Background: As BMP15, GDF9, and kisspeptin all play critical roles in folliculogenesis and fertilization, investigating the possible relationship between obesity and these three factors could prove crucial in relation to understanding the role of obesity in infertility. Thus, the present study sought to determine the effects of obesity on the serum BMP15, GDF9, and kisspeptin concentrations in women of reproductive age. Methods: Ninety female participants were equally divided into three groups: class-1 obese (n=30), class-2 obese (n=30), and normal weight (control; n=30). The participants' serum BMP15, GDF9, and AMH concentrations were measured. Moreover, the serum kisspeptin concentrations were evaluated in the class-1 obese and control groups by means of the enzyme-linked immunosorbent assay (ELISA) method while the participants were in their menstrual period. Results: The serum BMP15 and kisspeptin concentrations were found to be much higher in the control group than in both obese groups (p=0.001 and p=0.01, respectively). While the GDF9 concentration exhibited a statistically significant positive correlation with age, the BMP15 concentration exhibited a positive correlation with the kisspeptin and LH concentrations in the control group. In addition, a positive correlation was identified between the BMP15 concentration and both age and the glucose level and a negative correlation with the insulin level in both the obese groups. Conclusions: Obesity appears to reduce the serum BMP15 and kisspeptin concentrations in obese women of reproductive age. This reduction may represent a milestone in reproductive dysfunction and may be used to predict the success of infertility treatment in obese women.

Background: Litter size (LS) is a significant, challenging, and economical aspect of the goat industry in Indonesia. It is influenced by several different factors and genes; consequently, identifying potential genes and loci associated with litter size has become a genetic problem. Several genetic indicators have been found to be associated with litter size in goats. This has prompted the need to discuss candidate genes associated with litter size in goats in Indonesia. Methods: A systematic review was conducted using critical databases including ResearchGate, Google Scholar, PubMed, Google search engine and Science direct. There were any exclusion criteria, they were as follows: articles published in languages other than English, Conference papers, short communication papers and papers not related to animals. After reviewing the abstracts of 42 publications, the remaining 17 investigations were chosen for full paper evaluation. A further eight studies were removed after a comprehensive evaluation of the publications because they did not match our inclusion criteria. Results: These markers include growth differentiation factor 9 ( GDF9), bone morphogenetic protein 15 ( BMP15), bone morphogenetic protein receptor type IB ( BMPR1B), and kisspeptin ( KISS1). Single nucleotide polymorphisms in these genes contribute to the development of novel genetic markers that helps in the selection of goats with the most favorable genotypes for litter size. This type of genetic selection is more successful than the traditional way of selecting animals for reproductive traits, particularly litter size. Conclusions: As a result, this study summarizes the genetic impacts of polymorphisms in candidate genes associated with litter size features in Indonesian goats.

Sheep fertility genes have been studied in Turkey for many years. But so far it cannot be said that a satisfactory result has been achieved in terms of the number of detected polymorphisms on fertility genes. Exhibiting the genetic structure of local gene resources adapted to a particular region is very important for sustainability, conservation and breeding studies, and many other issues such as profitability. Therefore, the primary aim of this study was to investigate the single nucleotide polymorphisms (SNPs) on the BMP15 and GDF9 genes in two new Turkish sheep breeds, Çepni and Of. DNA sequencing was performed to identify SNPs in both genes of Çepni and Of sheep. One novel SNP (T755C) in the BMP15 gene and five known SNPs (c471C > T (G2), c477 G > A (G3), c721 G > A (G4), c978 A > G (G5), and c994 G > A (G6)) in the GDF9 gene, and a total of six SNPs were identified. Çepni and Of breeds showed a highly polymorphic structure for the genes examined. This finding shows that these two sheep breeds have great potential for future association studies. As a conclusion, the findings of this study were the first for the Çepni and Of breeds, and it should be examined for novel and other SNPs for more convincing results, especially litter size association studies, in new and other native sheep breeds in Turkey. [ABSTRACT FROM AUTHOR]

Background: Kisspeptin is a neuropeptide that has an important role in the female reproductive cycle which is indicated by its role in regulating the hypothalamic-pituitary-gonadal axis. Aims: To analyze the correlation between serum kisspeptin levels, ovarian kisspeptin expression, and ovarian Bone Morphogenic Protein-15 (BMP15) expression in polycystic ovary syndrome (PCOS) model rats. Methods: The research was accurate experimental research with a post-test design-only control group and was carried out from August to October 2022 at the Faculty of Veterinary Medicine Universitas Airlangga. 32 Rattus novergicus rats were divided into a control group and a PCOS model group. Blood serum and ovaries were obtained from all groups. In addition, blood serum was examined for kisspeptin levels by ELISA technique, and kisspeptin expression and BMP15 Ovaries were examined immunohistochemically. Results: Serum kisspeptin levels and ovarian kisspeptin expression of the PCOS model group were not significantly higher than those of the control group (p > 0.05, p > 0.05). The ovarian BMP15 expression of the PCOS model group was not significantly lower (p > 0.05) than that of the control group. Ovarian kisspeptin expression and ovarian BMP15 expression did not significantly correlate with serum kisspeptin levels (p > 0.05). In contrast, there was a significant correlation (p < 0.05) between ovarian kisspeptin expression and ovarian BMP15 expression. Conclusion: Serum kisspeptin levels and ovarian kisspeptin expression of the PCOS model group were not higher than those of the control group, and the ovarian BMP15 expression of the PCOS model group was not lower than that of the control group. There was no correlation between serum kisspeptin levels with ovarian kisspeptin expression and ovarian BMP15 expression. However, a significant correlation was found between ovarian kisspeptin expression and ovarian BMP15 expression. [ABSTRACT FROM AUTHOR]

Oocyte-secreted growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) are critical paracrine regulators of female fertility. Recent studies demonstrated that serum concentrations are associated with the number of oocytes retrieved during IVF, and therefore potential clinical use as biomarkers. However, it is unknown if the presence of endometriosis affects serum GDF9 or BMP15. An exploratory case–control study was prospectively performed on 60 women who underwent laparoscopy between April 2017 and August 2018 at two hospitals. GDF9 and BMP15 were measured by validated immunoassays in pre-operative serum samples. Data were analysed relative to laparoscopic assessment of endometriosis and staging. There were 35 women with confirmed laparoscopic diagnosis of endometriosis and 25 controls with no evidence of endometriosis at laparoscopy. GDF9 was detectable in 40% of controls and 48% of cases. There was no difference in median GDF9 concentrations between controls (20.0 pg/ml, range 20.0–2504 pg/ml) and cases (20.0 pg/ml, range 20.0–2963 pg/ml). BMP15 was detectable in 48% of controls and 58% of cases, with no difference in median concentrations between controls (26.5 pg/ml, range 24.0–1499 pg/ml) and cases (24.0 pg/ml, range 24.0–796 pg/ml). Furthermore, there were no significant differences in the proportion of detectable samples or concentrations of GDF9 or BMP15 with differing severities of endometriosis. In conclusion, serum concentrations of oocyte-secreted factors, GDF9 and BMP15 did not differ between control patients and patients with endometriosis. For clinical application in reproductive medicine, GDF9 and BMP15 serum biomarker quantitation is unlikely to be aberrant in the presence of endometriosis. [ABSTRACT FROM AUTHOR]

The Tibetan cashmere goat is a prolific goat breed in China. In sheep breeds, natural mutations have demonstrated that the transforming growth factor beta (TGF-β) super family ligands, such as growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15) and their type I receptor (bone morphogenetic protein receptor (BMPR1B), are essential for ovulation and increasing litter size. In this study, 216 female Tibetan cashmere goats were sampled, and candidate genes with fecundity traits were detected via restriction fragment length polymorphism (RFLP) and sequenced. Four polymorphic loci were found in specific amplification fragments of BMP15 and GDF9. Two SNP sites of the BMP15 gene were discovered, namely G732A and C805G. The G732A mutation did not cause the change in amino acids, and the frequencies of each genotype were 0.695 for the GG type, 0.282 for the GA type and 0.023 for the AA type. The C805G mutation caused amino acids to change from glutamine to glutamate. The genotype frequencies were 0.620 for the CC type, 0.320 for the CG type and 0.320 for the CG type. For the GG type 0.060, the G3 and G4 mutations of the GDF9 gene were all homozygous mutations. Two known SNP sites, C719T and G1189A, were detected in the Tibetan cashmere goat GDF9 gene, of which the C719T mutation caused a change of alanine to valine, with a genotype frequency of 0.944 for the CC type and 0.056 for the CT type, whereas no TT type was found. The G1189A mutation caused valine to become isoleucine, and the frequencies of each genotype were 0.579 for the GG type, 0.305 for the GA type and 0.116 for the AA type; G1, B2, B3, B4, FecXH, FecXI, FecXL, G2, G5, G6, G7, G8, FecGE, FecTT and FecB mutations were not found in Tibetan cashmere goats. The results of this study provide a data basis for future studies of BMP15, GDF9 and BMPR1B gene mutations in goats. [ABSTRACT FROM AUTHOR]

The improvement of the reproductive traits of animals is of great interest for livestock production. Due to its positive impact on the sheep industry's profitability, prolificacy is one of the most economically significant biological traits, showing variation between and within breeds of domestic sheep (Ovis aries). Different mutations in BMPR-1B, BMP15 and GDF9 genes coding for the transforming growth factor-β (TGFβ) superfamily have been shown to influence the ovulation rate and litter size. Numerous single-nucleotide polymorphisms (SNPs) in the bone morphogenetic protein 15 (BMP15) gene have been linked to ewe fecundity. Using targeted PCR amplification and Sanger sequencing, we were able to identify heterozygous SNPs in exon 2 of BMP15 in three sheep breeds reared in Romania: Tsigai, Cluj Merino and Tsurcana. The sequence analysis revealed three previously documented mutations, namely the missense mutation c.755T>C (L252P), which is predicted to change the tertiary structure of the BMP15 protein, and two silent mutations, c.747T>C (P249P) and c.1047G>A (V349V). In addition, we also identified one novel silent mutation, c.825G>A (S275S). Based on our findings and publicly available data, we indicate four putative mutational hotspots within exon 2 of BMP15 that could be considered for improving the indigenous sheep breeds through targeted gene editing and SNP genotyping strategies. [ABSTRACT FROM AUTHOR]

Ovarian dysfunction is one of the most common features of women with Turner syndrome. In these women, oocyte apoptosis is markedly accelerated from the early stage of fetal life. Reduction in the number of germ cells disturbs primordial follicle development and thereby leads to the formation of streak gonads. There are three possible causes of accelerated germ cell loss in 45,X ovaries. First, chromosomal pairing failure due to X chromosomal aneuploidy is believed to induce meiotic arrest. Indeed, it has been suggested that the dosage of the X chromosome is more critical for the survival of the oocytes than for other cells in the ovary. Second, impaired coupling between oocytes and granulosa cells may also contribute to germ cell apoptosis. Previous studies have shown that 45,X ovaries may tend to lose tight junctions which are essential for intercellular interactions. Lastly, ovarian dysfunction in women with Turner syndrome is partly attributable to the reduced dosage of several genes on the X chromosome. Specifically, BMP15, PGRMC1, and some other genes on the X chromosome have been implicated in ovarian function. Further studies on the mechanisms of ovarian dysfunction are necessary to improve the reproductive outcomes of women with Turner syndrome. [ABSTRACT FROM AUTHOR]

Bone morphogenetic protein 15 (BMP15) regulates the growth and development of follicles. In particular, the long non-coding RNA H19 plays an important role in mammalian reproduction. However, the function and regulatory mechanism of the interaction of BMP15 with H19 in yak granulosa cell (GC) proliferation, autophagy, and apoptosis are poorly understood. In our study, quantitative reverse-transcription-polymerase chain reaction analysis showed that H19 were highly expressed in yak healthy follicles. H19 was induced by BMP15 protein in yak GCs. In addition, we confirmed that overexpression of H19 promoted yak GC proliferation and autophagy and inhibited apoptosis. Bioinformatic analysis and luciferase reporter assays demonstrated that H19 directly binds to miR-26b, and SMAD1 was identified as a target of miR-26b. miR-26b overexpression inhibited GC proliferation and autophagy and promoted apoptosis through decreased SMAD1 expression, which was attenuated by H19 overexpression. RNA immunoprecipitation-quantitative polymerase chain reaction and dual-luciferase assays showed that miR-26b was sponged by H19 to preserve SMAD1 expression. Furthermore, SMAD1 mRNA expression was induced and miR-26b expression was reduced after yak GCs were treated with BMP15 protein. In conclusion, our results demonstrated that the H19/miR-26b/SMAD1 axis responds to BMP15 to regulate yack GC proliferation, autophagy, and apoptosis. [ABSTRACT FROM AUTHOR]

Objective: Growth differentiation factor‐9 (GDF9) and bone morphogenetic protein‐15 (BMP15) are critical paracrine regulators of female fertility and are predominantly expressed by oocytes. However, it is unknown if serum concentrations reflect changes in ovarian function and/or reproductive endocrine disorders. This study aimed to determine if serum GDF9/BMP15 are associated with ovarian, pituitary, oestrogenic, androgenic and metabolic characteristics and the ovarian pathologies, polycystic ovarian morphology (PCOM) and polycystic ovary syndrome (PCOS). Design: Women aged 21−45 years (n = 381) were included from a cross‐sectional study at the National University Hospital, Singapore. Patients: Participants were volunteers and patients with possible PCOS. Measurements: Anthropometric measurements, transvaginal ultrasound scans and serum sampling were performed and a questionnairecompleted. Serum GDF9 and BMP15 concentrations were matched with menstrual cycle length, ovarian protein and steroid hormone production, pituitary hormone production and metabolic assessments in women with PCOM or PCOS and those with neither (control). Results: Serum GDF9 and BMP15 were detectable in 40% and 41% of women, respectively and were positively correlated with each other (r = 0.08, p = 0.003). GDF9, but not BMP15, was positively correlated with ovarian volume (p = 0.02) and antral follicle count (AFC) (p = 0.004), but not with anti‐Müllerian hormone (p = 0.05). However, serum GDF9 and BMP15 concentrations were not significantly different between control, PCOM and PCOS women, nor associated with androgenic or metabolic PCOS features. However, the relationship between GDF9 and AFC differed between control, PCOM and PCOS women (p = 0.02). Conclusions: Serum GDF9 and BMP15 concentrations somewhat reflect ovarian but not androgenic or metabolic characteristics of PCOS, with increased GDF9 reflecting high AFC as seen in PCOM/PCOS. [ABSTRACT FROM AUTHOR]

Simple Summary: BMPRIB, GDF9, BMP15, LEPR, and B4GALNT2 genes were regarded as leading candidates for reproductive traits in sheep. Our recent studies identified 13 novel mutations associated with litter size in the Mongolia sheep breed in these 5 genes. In this study, we performed an association analysis of the 13 new mutations and 7 known ovine prolificacy-related mutations with litter size in Ujimqin, the F1 population of Dorper × Ujimqin crossbred, and the F1 population of Suffolk × Ujimqin crossbred. The results suggested that BMPR1B c.746A>G (FecB), GDF9 c.994A>G (FecGA), and BMP15 c.31_33CTTinsdel (B1) may be potentially effective genetic markers to improve the litter size in sheep. Prolificacy is a crucial characteristic of livestock, particularly for species such as sheep that have many births. The objectives of this study were as follows: (1) to investigate the genetic diversity of the 13 new and 7 known variants in the BMPRIB, GDF9, BMP15, LEPR, and B4GALNT2 genes in Ujimqin (UM), the F1 population of Dorper × Ujimqin crossbred (DPU), the F1 population of Suffolk × Ujimqin crossbred (SFKU), Sonid sheep (SN), Tan sheep (Tan), Hu sheep (Hu), and Small-tailed Han sheep (STH) sheep breeds/populations; (2) to perform an association analysis of the above 20 variants with litter size in 325 UM, 304 DPU, and 66 SFKU sheep populations; (3) to compare the frequencies of the litter-size-related alleles of these 20 variants among 8 sheep breeds/populations (the above seven sheep breeds + Mongolia sheep breed). With the use of the Sequenom MassARRAY®SNP assay technology, these 20 mutations were genotyped. The association analysis results showed that the c.746A>G (FecB) mutation in BMPR1B was significantly associated with the litter size of UM and DPU, the c.994A>G (FecGA) in GDF9 was significantly associated with the litter size of SFKU, and the c.31_33CTTinsdel (B1) in BMP15 was significantly associated with the litter size of UM. Our findings might provide valuable genetic markers for expanding sheep litter sizes. [ABSTRACT FROM AUTHOR]

Purpose: To analyze the level of growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) in follicle fluid (FF) and granulosa cells (GCs) derived from young patients with low prognosis for in vitro fertilization and embryo transfer (IVF-ET) treatment. Methods: A prospective cohort study was carried out by enrolling 52 young patients with low prognosis according to the POSEIDON classification group 3 (low prognosis group) and 51 young patients with normal ovarian reserve (control group). The concentration of the GDF9 and BMP15 proteins in FF was determined by enzyme-linked immunosorbent assay. The mRNA level of the GDF9 and BMP15 in the GCs was measured by quantitative real-time PCR. Results: The concentration of GDF9 (1026.72 ± 159.12 pg/mL vs. 1298.06 ± 185.41 pg/mL) and BMP15 (685.23 ± 143.91 pg/mL vs. 794.37 ± 81.79 pg/mL) in FF and the mRNA level of GDF9 and BMP15 in the GCs and the live birth rate per treatment cycle started (30.77% vs. 50.98%) and oocytes retrieved (4.25 ± 1.91 vs.12.04 ± 4.24) were significantly lower, whereas the canceled cycle rate was significantly higher (9.62% vs. 0) in the low prognosis group compared with the control group (P < 0.05). The expression of GDF9 and BMP15 in the ovary was positively correlated with live birth (P < 0.05). Conclusion: The expression of GDF9 and BMP15 in the ovary was decreased in young patients with low prognosis accompanied by a poorer outcome of IVF-ET treatment. Trial registration: ChiCTR1800016107 (Chinese Clinical Trial Registry), May 11, 2018. (http://www.chictr.org.cn/edit.aspx?pid=27216&htm=4). [ABSTRACT FROM AUTHOR]

Bone morphogenetic protein 15 (BMP15) is an X-linked gene encoding an oocyte secreted factor, which plays varied functions in the female fertility between mono-ovulatory and poly-ovulatory mammalian species. We previously found that knockout of BMP15 completely blocked porcine follicular development at preantral stages. However, the specific function of BMP15 on porcine oocytes in vitro maturation remains largely unknown. Here, we injected the pre-assembled crRNA + tracrRNA + Cas9 ribonucleoprotein (ctRNP) complex into the cytoplasm of germinal vesicle stage porcine oocytes to disrupt BMP15. The ctRNP composed of Cas9 nuclease and crRNA-tracrRNA complex at 1.2/1 content ratio. The tested crRNA-tracrRNA complex concentration ranging from 50 to 200 ng/μL, all presented effective editing of BMP15 in porcine oocytes, and the 125 ng/μL crRNA-tracrRNA complex presented the highest editing efficiency (39.23 ± 3.33%). Surprisingly, we found approximately 95% edited oocytes presented monoallelic mutations, and only 5% edited oocytes harbored biallelic mutations. Interestingly, the coinjected two crRNAs guided the ctRNP complex to concurrently cut within a 10 bp window of the PAM (protospacer adjacent motif), resulting in a precise deletion within BMP15 in 85.9% edited oocytes, and additional deletion happened in 14.1% edited oocytes, which resulted in large fragment deletions in BMP15. Most deletions caused frameshift and introduced premature stop codon in BMP15 , resulting in the disruption of BMP15 protein expression, which was confirmed by the Western blot analysis showing the reduced BMP15 protein expression in ctRNP injected oocytes. The disruption of BMP15 attenuated the activation of SMAD1/5/8 signaling, and impaired cumulus expansion of porcine cumulus cell-oocyte complexes (COCs). Our study proved that delivering CRISPR ctRNP into porcine oocytes by microinjection was able to edit BMP15 efficiently, providing a new strategy to investigate the functions of oocyte-specific secreted factors in oocyte in vitro maturation. [ABSTRACT FROM AUTHOR]

Reproductive traits, such as ovulation rate and litter size, are important factors influencing the sheep industry. The bone morphogenetic protein 15 (BMP15) is a major gene affecting the reproductive traits in sheep, and multiple mutations in BMP15 gene could affect the ovulation rate and litter size in many sheep breeds, showing high breed specificity. However, identification of novel variations and seeking breed-specific markers associated with litter size in other sheep breeds are still important. In this study, we sequenced the BMP15 gene of Mongolia sheep, and 12 novel variants were detected by direct sequencing and whole-genome resequencing. Among them, the g.50985975 G > A polymorphism in intron and synonymous c.755 T > C (Leu252Pro) in exon 2 of BMP15 were significantly associated with the litter sizes of Mongolia ewes (P < 0.01 and P < 0.01, respectively), as well as the g.50988478C > A and g.50987863G > A in the promoter region of BMP15 were significantly associated with the litter sizes of Ujimqin ewes (P < 0.05 and P < 0.01, respectively). The c.755 T > C mutation is predicted to change the tertiary structure of BMP15. Our findings may provide potentially useful genetic markers for increasing litter size in sheep. • Twelve novel variants were identified in ovine BMP15. • The g.50985975 G > A and c.755 T > C mutations associated with litter size in Mongolia sheep were found. • The g.50988478C > A and g.50987863G > A mutations associated with litter size in Ujimqin sheep were found. • The c.755 T > C (Leu252Pro) is predicted to affect the tertiary structure of BMP15 protein. [ABSTRACT FROM AUTHOR]

Abstract Background The suggested effects of the oocyte secreted GDF9 and BMP15 growth factors on oocyte maturation are currently based on recombinant proteins, and little is known about native GDF9 and BMP15 in humans. Methods Human immature cumulus-oocyte complexes (COCs) obtained in connection with ovarian tissue cryopreservation (OTC) underwent in vitro maturation (IVM). Oocyte-produced GDF9 and BMP15 were detected in COCs using immunofluorescence, and in fresh GV oocytes and in GV and MII oocytes after IVM by western blot. Concentrations of GDF9, BMP15 homodimers, and GDF9/BMP15 heterodimer in spent media after IVM were measured by ELISA. The relative expression of seven genes from the GDF9 and BMP15 signaling pathways (BMPR2, ALK5, ALK6, SMAD1, SMAD2, SMAD3, and SMAD5) was evaluated in fresh cumulus cells (before IVM) and in cumulus cells from GV and MII oocytes after IVM by RT-qPCR. Results We detected native pro-mature GDF9 and BMP15 in human oocytes with molecular weights (Mw) of 47 kDa and 43 kDa, respectively. Concentrations of GDF9 and BMP15 in spent media after IVM were detected in 99% and 64% of the samples, respectively. The GDF9/BMP15 heterodimer was detected in 76% of the samples. Overall, the concentration of GDF9 was approximately 10-times higher than BMP15. The concentrations of both GDF9 and BMP15 were significantly lower in spent medium from MII oocytes than in media from oocytes that remained at the GV stage. Concentrations of the GDF9/BMP15 heterodimer did not differ between GV and MII oocytes. Furthermore, BMPR2, SMAD3, and SMAD5 were significantly upregulated in cumulus cells from MII oocytes, indicating that both GDF9 and BMP15 signaling were active during oocyte meiotic resumption in vitro. Conclusion These data suggest that the driving mechanisms for oocyte nuclear maturation may involve both GDF9 and BMP15 homodimers, while the role of the GDF9/BMP15 heterodimer is questionable.

Introduction: According to previous reports, treatment of infertile women who are candidates for in vitro fertilization (IVF), about 15% of the oocytes remains immature. Therefore, finding an efficient approach to make these types of oocytes usable in the clinic, especially in women with limited oocytes, is necessary. One of these approaches is the laboratory culture of immature oocytes in such a way that the expression of genes involved in cytoplasmic maturation increases. Methods: Forty-eight infertile women candidates for IVF donated their immature oocytes at the time of oocyte retrieval. The obtained immature oocytes were randomly divided into two groups: the control group without in-Vitro culture and the intervention group in the form of 24-hour culture. The total oocytes of both groups (ninety-five immature oocytes) were pooled separately and analyzed by quantitative polymerase chain reaction (q-PCR). Results: The level of expression of MT-ATP6 and BMP15 genes in the intervention group has a significant increase compared to the control group, with a fold change of 7.867± 3.12 and 6.327 ± 0.78, respectively (P<0.001). Furthermore, according to the morphological changes, 54% of the immature oocytes in the intervention group had resumed meiosis. Conclusion: Increasing the expression of two cytoplasmic maturation genes, MT-ATP6 and BMP15 can cause the resumption of meiosis in immature oocytes. Therefore, this strategy can be promising in women with low eggs. [ABSTRACT FROM AUTHOR]

Background: Litter size (LS) is a significant, challenging, and economical aspect of the goat industry in Indonesia. It is influenced by several different factors and genes; consequently, identifying potential genes and loci associated with litter size has become a genetic problem. Several genetic indicators have been found to be associated with litter size in goats. This has prompted the need to discuss candidate genes associated with litter size in goats in Indonesia. Methods: A systematic review was conducted using critical databases including ResearchGate, Google Scholar, PubMed, Google search engine and Science direct. There were any exclusion criteria, they were as follows: articles published in languages other than English, Conference papers, short communication papers and papers not related to animals. After reviewing the abstracts of 42 publications, the remaining 17 investigations were chosen for full paper evaluation. A further eight studies were removed after a comprehensive evaluation of the publications because they did not match our inclusion criteria. Results: These markers include growth differentiation factor 9 ( GDF9), bone morphogenetic protein 15 ( BMP15), bone morphogenetic protein receptor type IB ( BMPR1B), and kisspeptin ( KISS1). Single nucleotide polymorphisms in these genes contribute to the development of novel genetic markers that helps in the selection of goats with the most favorable genotypes for litter size. This type of genetic selection is more successful than the traditional way of selecting animals for reproductive traits, particularly litter size. Conclusions: As a result, this study summarizes the genetic impacts of polymorphisms in candidate genes associated with litter size features in Indonesian goats.

Abstract Background Bone morphogenetic protein 15 (BMP15) is expressed in oocytes and plays a crucial role in the reproduction of mono-ovulating species. In humans, BMP15 gene mutations lead to imperfect protein function and premature ovarian insufficiency. Here we investigated the BMP15 gene variants in a population of Iranian women with premature ovarian insufficiency. We conducted predictive bioinformatics analysis to further study the outcomes of BMP15 gene alterations. Methods Twenty-four well-diagnosed premature ovarian insufficiency cases with normal karyotype participated in this study. The entire coding sequence and exon-intron junctions of the BMP15 gene were analyzed by direct sequencing. In-silico analysis was applied using various pipelines integrated into the Ensembl Variant Effect Predictor online tool. The clinical interpretation was performed based on the approved guidelines. Results By gene screening of BMP15, we discovered p.N103K, p.A180T, and p.M184T heterozygous variants in 3 unrelated patients. The p.N103K and p.M184T were not annotated on gnomAD, 1000 Genome and/or dbSNP. These mutations were not identified in 800 Iranians whole-exome sequencing that is recorded on Iranom database. We identified the p.N103K variant in a patient with secondary amenorrhea at the age of 17, elevated FSH and atrophic ovaries. The p.M184T was detected in a sporadic case with atrophic ovaries and very high FSH who developed secondary amenorrhea at the age of 31. Conclusions Here we newly identified p.N103K and p.M184T mutation in the BMP15 gene associated with idiopathic premature ovarian insufficiency. Both mutations have occurred in the prodomain region of protein. Despite prodomain cleavage through dimerization, it is actively involved in the mature protein function. Further studies elucidating the roles of prodomain would lead to a better understanding of the disease pathogenesis.

Growth and differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) are oocyte-secreted factors with a leading role in the control of ovarian function in female reproduction, modulating both the cell fate of the somatic granulosa cells and the quality and developmental competence of the egg. This short review aims to consolidate the molecular aspects of GDF9 and BMP15 and their integral actions in female fertility to understand particularly their effects on oocyte quality and fetal growth. The significant consequences of mutations in the GDF9 and BMP15 genes in women with dizygotic twins as well as the clinical relevance of these oocyte factors in the pathogenesis of primary ovarian insufficiency and polycystic ovary syndrome are also addressed.

Growth and differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) are oocyte-secreted factors with a leading role in the control of ovarian function in female reproduction, modulating both the cell fate of the somatic granulosa cells and the quality and developmental competence of the egg. This short review aims to consolidate the molecular aspects of GDF9 and BMP15 and their integral actions in female fertility to understand particularly their effects on oocyte quality and fetal growth. The significant consequences of mutations in the GDF9 and BMP15 genes in women with dizygotic twins as well as the clinical relevance of these oocyte factors in the pathogenesis of primary ovarian insufficiency and polycystic ovary syndrome are also addressed.

Background: In female fertility, age is a most important factor. With increasing age, there will be decrease in both oocyte quantity and quality, and ovulatory dysfunction, is more common in younger female (1). Aim of study: Evaluation of the effect of age on serum and follicular fluid BMP15 and GDF9, the oocyte secreted factors. Subjects, Materials and Methods: One hundred and seventy six women were included in this study, eighty eight women (Group 1) are selected from those undergoing intra-cytoplasmic sperm injection, also eighty eight postmenopausal women (group 2). Results: Age was not significantly correlated to serum or follicular fluid BMP15 and serum and follicular GDF9 (p > 0.05) in premenopausal group, but decreased significantly with age in postmenopausal group. Conclusions: Very low level of serum BMP15 in women could be used as indicator of empty follicles and menopause. [ABSTRACT FROM AUTHOR]

Objective: To analyze the changes of serum bone morphogenetic protein-15 (BMP15), interleukin-1β (IL-1β), serum inhibin B (INHB) and macrophage migration inhibitory factor (MIF) in patients with autoimmune premature ovarian failure (APOF) and their relationship with the expression of serum sex hormones and peripheral blood T lymphocyte subsets.Methods: 52 patients diagnosed with APOF who visited The First Affiliated Hospital of Hebei North University from January to December 2021 were included in the APOF group, and 50 healthy women with normal ovarian function who underwent physical examination in The First Affiliated Hospital of Hebei North University during the same period were included in the health group. The levels differences of serum BMP15, IL-1β, INHB and MIF between APOF group and healthy group, patients with different stages of APOF were compared, and the levels differences of serum sex hormones and peripheral blood T lymphocyte subsets were compared between APOF group and healthy group. The correlation of serum BMP15, IL-1β, INHB, MIF with serum sex hormone levels and peripheral blood T lymphocyte subsets was analyzed.Results: The levels of serum BMP15 and INHB in APOF group were significantly lower than those in healthy group, while the levels of serum IL-1β and MIF in APOF group were significantly higher than those in healthy group (all P＜0.05). The levels of serum BMP15 and INHB in patients with course of disease > 3 years were significantly lower than those in patients with course of disease >3 years, while the levels of serum IL-1β and MIF were significantly increased (all P＜0.05). Follicle stimulating hormone (FSH) in APOF group was significantly higher than that in healthy group, the level of serum estradiol (E2) was significantly lower than that in healthy group (all P＜0.05). The level of CD4+ in APOF group was significantly lower than that in healthy group (P＜0.05). Serum BMP15 and INHB were negatively correlated with FSH and CD4+, and positively correlated with E2, serum IL-1β and MIF were positively correlated with FSH and CD4+, and negatively correlated with E2 (all P＜0.05).Conclusion: With the prolongation of the course of APOF, the levels of serum BMP15 and INHB are decreased, while the levels of serum IL-1β and MIF are increased, and the above indicators are closely related to the levels of sex hormones and peripheral blood T lymphocyte subsets. [ABSTRACT FROM AUTHOR]

Background: Premature ovarian insufficiency (POI) occurs in women before the age of 40 years, accompanied by amenorrhea, hypoestrogenism, hypergonadotropinism, and infertility. The pathology of POI is complex and the molecular genetic mechanisms are poorly understood. Bone morphogenetic protein 15 (BMP15) plays a crucial role in oocyte maturation and follicular development through the activation of granulosa cells. Dysfunction of BMP15 causes ovarian dysgenesis and is related to POI. Identifying pathogenic variants contributes to revealing genetic mechanisms and making clinical diagnoses of POI. Methods: The study involved two sisters diagnosed with POI. Whole-exome sequencing (WES) was performed to identify causative genes. Sanger sequencing was used to validate the mutations in patients with POI and members of the family with no clinical signs or symptoms. The effect of the novel mutations on the BMP15 structure was analyzed by PSIPRED. By over-expressing wild-type (WT) or mutant BMP15 plasmids in vitro, a functional study of the BMP15 mutant was conducted by real-time qPCR and western blotting. Through cocultivation with HEK293T cells, the effects of secreted BMP15 WT and variants on granulosa cell proliferation and apoptosis were detected through a cell counting kit-8 assay and flow cytometric analysis. Results: We identified biallelic variants in BMP15, c.791G > A (p. R264Q) and c.1076C > T (p. P359L), in two siblings with POI. Both sisters carried the same biallelic variants, while the other female members of their family carried only one of them. Structural prediction showed that the variants have not affected the secondary structure of BMP15 but may change the conformation of water molecules around protein surfaces and thermal stability of BMP15. Real-time qPCR showed no significant difference in mRNA levels among WT and the two variants. Western blotting indicated a reduction in BMP15 expression with the c.791G > A and c.1076C > T variants compared to WT. Moreover, mutants 791G > A and 1076C > T impaired the function of secreted BMP15 in promoting granulosa cell proliferation and suppressing cell apoptosis caused by reactive oxygen species. Conclusions: This study identified novel biallelic variants, c.791G > A and c.1076C > T, of BMP15 in two siblings with POI. Both missense variants reduced the level of the BMP15 protein and impaired the function of BMP15 in promoting granulosa cell proliferation in vitro. Taken together, our findings provide a novel molecular genetic basis and potential pathogenesis of BMP15 variants in POI. [ABSTRACT FROM AUTHOR]

Objectives: Use of serum and follicular fluid concentration of oocyte secreted factors BMP15 as biomarkers of oocyte quality, embryo quality and it's relation to pregnancy rate. Methods: Eighty eight women were included in this study; they were selected from those undergoing intra-cytoplasmic sperm injection. Results: Positive pregnancy was achieved by 14 women accounting for 19.0% (total number of women that reach embryo transfer was 72). No significant difference in mean serum BMP15 between pregnant and non-pregnant women, but the level of follicular fluid BMP15 was significantly higher in pregnant women. MI oocyte count was not significantly correlated to serum or follicular fluid BMP15 (P > 0.05). MII oocyte count showed highly significant positive correlation to serum and follicular fluid BMP15 (P < 0.01). Grade 1 embryo count showed highly significant positive correlation to serum and follicular fluid BMP15 (P < 0.01), Grade 2 embryo count showed significant positive correlation to serum BMP15 (P = 0.032), but the grade 2 embryo count showed non-significant correlation to follicular BMP15 (P > 0.05). Also grade 3 embryo count showed non-significant correlation to serum and follicular BMP15 (P > 0.05). Conclusion: The current study revealed that serum and follicular BMP15 could be used as indicator for oocyte maturity, and serum BMP15 could be used as indicator of good quality embryos. [ABSTRACT FROM AUTHOR]

مقدمه: نارسایی زودرس تخمدان (POF)، به توقف فعالیت تخمدان‌ها قبل از سن 40 سالگی گفته می‌شود که با سطح پایین هورمون‌های جنسی و سطح بالای گنادوتروپین‌ها همراه است. عوامل ژنتیکی بیشترین نقش را درPOF ایفا می‌کنند. BMP15 و گیرنده ی آن BMPR1B از اعضای ابرخانواده‌ی TGFβ هستند که در بافت تخمدان بیان می‌شوند و در مسیر فولیکولوژنز دخالت دارند. هرگونه تغییر در این ژن‌ها ممکن است مسیر فولیکولوژنز را مختل کند. بنابراین هدف از این مطالعه بررسی 4 واریانت در اگزون 2 ژن BMP15 و دو واریانت در اگزون 9 ژن BMPR1B به عنوان واریانتی کاندید در زنان نابارور ایرانی مبتلا به POF می‌باشد.روش کار: با استفاده از دیتابیس Uniprot و تکنیک PCR-SSCP ارتباط بین واریانت‌های 443T>C، 468G>A، 538G>A، 598C>T ژن BMP15 و 945G>A،1020A>G ژن BMPR1B در 45 زن مبتلا به POF و 45 زن سالم صاحب فرزند، بدون هیچگونه سابقه بیماری‌های مرتبط با فولیکولوژنز و تخمک‌گذاری به عنوان کنترل بررسی شد. سپس تعدادی از محصولات PCR با تغییر الگوی حرکتی باندها، تعیین توالی شدند و نتایج حاصل از آن با نرم‌افزار CLC تحلیل شد.نتایج: نتایج ژل الکتروفورز، SSCP و گراف های تعیین توالی هیچ کدام از واریانت‌ها، تغییری را در توالی ژنوم افراد بیمار نسبت به افراد کنترل نشان ندادند.بحث و نتیجه گیری: با توجه به نقش کلیدی BMP15 در مسیر فولیکولوژنز و گزارش مطالعات قبلی، احتمالا BMP15 در پیدایش POF مؤثر باشد. عدم مشاهده‌ی SNP در بیماران این مطالعه می‌تواند به علت کم بودن حجم نمونه‌های مورد مطالعه و تفاوت های ژنتیکی افراد باشد.

Collichthys lucidus (C. lucidus) is an economically important fish species, exhibiting sexual dimorphism in its growth rate. However, there is a lack of research on its underlying sex-related mechanisms. Therefore, small RNA sequencing was performed to better comprehend these sex-related molecular mechanisms. In total, 171 differentially expressed miRNAs (DEmiRNAs) were identified between the ovaries and testes. Functional enrichment analysis revealed that the target genes of DE-miRNAs were considerably enriched in the p53 signaling, PI3K-Akt signaling, and TGF-beta signaling pathways. In addition, sex-related miRNAs were identified, and the expression of miR-430c-3p and miR-430f-3p was specifically observed in the gonads compared with other organs and their expression was markedly upregulated in the testes relative to the ovaries. Bmp15 was a target of miR-430c-3p and was greatly expressed in the ovaries compared with the testes. Importantly, miR-430c-3p and bmp15 co-expressed in the ovaries and testes. This research provides the first detailed miRNA profiles for C. lucidus concerning sex, likely laying the basis for further studies on sex differentiation in C. lucidus. [ABSTRACT FROM AUTHOR]

Background: The suggested effects of the oocyte secreted GDF9 and BMP15 growth factors on oocyte maturation are currently based on recombinant proteins, and little is known about native GDF9 and BMP15 in humans. Methods: Human immature cumulus-oocyte complexes (COCs) obtained in connection with ovarian tissue cryopreservation (OTC) underwent in vitro maturation (IVM). Oocyte-produced GDF9 and BMP15 were detected in COCs using immunofluorescence, and in fresh GV oocytes and in GV and MII oocytes after IVM by western blot. Concentrations of GDF9, BMP15 homodimers, and GDF9/BMP15 heterodimer in spent media after IVM were measured by ELISA. The relative expression of seven genes from the GDF9 and BMP15 signaling pathways (BMPR2, ALK5, ALK6, SMAD1, SMAD2, SMAD3, and SMAD5) was evaluated in fresh cumulus cells (before IVM) and in cumulus cells from GV and MII oocytes after IVM by RT-qPCR. Results: We detected native pro-mature GDF9 and BMP15 in human oocytes with molecular weights (Mw) of 47 kDa and 43 kDa, respectively. Concentrations of GDF9 and BMP15 in spent media after IVM were detected in 99% and 64% of the samples, respectively. The GDF9/BMP15 heterodimer was detected in 76% of the samples. Overall, the concentration of GDF9 was approximately 10-times higher than BMP15. The concentrations of both GDF9 and BMP15 were significantly lower in spent medium from MII oocytes than in media from oocytes that remained at the GV stage. Concentrations of the GDF9/BMP15 heterodimer did not differ between GV and MII oocytes. Furthermore, BMPR2, SMAD3, and SMAD5 were significantly upregulated in cumulus cells from MII oocytes, indicating that both GDF9 and BMP15 signaling were active during oocyte meiotic resumption in vitro. Conclusion: These data suggest that the driving mechanisms for oocyte nuclear maturation may involve both GDF9 and BMP15 homodimers, while the role of the GDF9/BMP15 heterodimer is questionable. [ABSTRACT FROM AUTHOR]

Context: The oocyte-secreted factors growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) play essential roles in follicle development and oocyte maturation, and aberrant regulation might contribute to the pathogenesis of polycystic ovary syndrome. Objective: Are there measurable differences in concentrations of GDF9, BMP15, and the GDF9/BMP15 heterodimer in small antral follicle fluids from women with and without polycystic ovaries (PCO)? Design and Setting: Follicle fluids (n = 356) were collected from 4- to 11-mm follicles in unstimulated ovaries of 87 women undergoing ovarian tissue cryopreservation for fertility preservation. Patients: Twenty-seven women with PCO were identified and 60 women without PCO-like characteristics (non-PCO women) were matched according to age and follicle size. Main outcome measures: Intrafollicular concentrations of GDF9, BMP15, GDF9/BMP15 heterodimer, anti-Mullerian hormone (AMH), inhibin-A and -B, total inhibin, activin-B and -AB, and follistatin were measured using enzyme-linked immunosorbent assays. Results: The detectability of GDF9, BMP15, and the GDF9/BMP15 heterodimer were 100%, 94.4%, and 91.5%, respectively, and concentrations were significantly negatively correlated with increasing follicle size (P < 0.0001). GDF9 was significantly higher in women with PCO (PCO: 4230 ± 189 pg/mL [mean ± SEM], n = 188; non-PCO: 3498 ± 199 pg/mL, n = 168; P < 0.03), whereas BMP15 was lower in women with PCO (PCO: 431 ± 40 pg/mL, n = 125; non-PCO: 573 ± 55 pg/mL, n = 109; P = 0.10), leading to a significantly higher GDF9:BMP15 ratio in women with PCO (P < 0.01). Significant positive associations between BMP15 and AMH, activins, and inhibins in non-PCO women switched to negative associations in women with PCO. Conclusions: Intrafollicular concentrations of GDF9 and BMP15 varied inversely in women with PCO reflecting an aberrant endocrine environment. An increased GDF9:BMP15 ratio may be a new biomarker for PCO. [ABSTRACT FROM AUTHOR]

The inhibin a (INHA), inhibin ßA (INHBA) and bone morphogenetic protein 15 (BMP-15) were investigated as candidate genes for reproductive traits in four Iranian sheep breeds (Bahmaei, Lak Ghashghaei, Lori-Bakhtiari and Karakul). Based on the ovine sequences of BMP-15, INHA and INHBA genes, three pairs of primers were designed to identify single nucleotide polymorphisms (SNPs) in exon 2 of BMP-15, INHA and INHBA in multiparous ewes by DNA sequencing. Two SNPs were detected in exon 2 of the ovine BMP15 gene at positions 367 and 430, which lead to amino acid substitutions at position 231 and 252 in the BMP15 protein sequence, respectively. Substitution of Leucine to Proline at position 252 is predicted to affect the protein function. A synonymous mutation was found in the amplified fragment of exon 2 at position 752 in ovine INHBA gene. In addition, the c752C>T mutation was only found in heterozygous condition in only one Lori-Bakhtiari ewe, while other breeds were in wild type genotype for c752C>T mutation. The INHA gene was shown to be highly polymorphic. A total of 7 SNPs including 6 nucleotide substitutions and one insertion were found in the amplified fragment of the INHA locus. The insertion mutation was found in two animals of Bahmaei and Karakul breeds. Interestingly, homozygous condition for the mutant alleles in all identified SNPs in BMP15, INHA and INHBA loci was absent in these breeds. Generally, these breeds showed different genetic structures with regard to the identified SNPs in BMP15, INHBA and INHA genes. However, further research with larger sample size and phenotype data on reproductive performance is required to investigate the definitive effect of the identified mutations in this study. [ABSTRACT FROM AUTHOR]

BackgroundAberrant signaling between germ cells and somatic cells can lead to reproductive disease and depends on diffusible signals, including transforming growth factor-beta (TGFB) -family proteins. The TGFB-family protein Gsdf (gonadal soma derived factor) controls sex determination in some fish and is a candidate for mediating germ cell/soma signaling.ResultsZebrafish expressed gsdf in somatic cells of bipotential gonads and expression continued in ovarian granulosa cells and testicular Sertoli cells. Homozygous gsdf knockout mutants delayed leaving the bipotential gonad state, but then became a male or a female. Mutant females ovulated a few oocytes, then became sterile, accumulating immature follicles. Female mutants stored excess lipid and down-regulated aromatase, gata4, insulin receptor, estrogen receptor, and genes for lipid metabolism, vitellogenin, and steroid biosynthesis. Mutant females contained less estrogen and more androgen than wild-types. Mutant males were fertile. Genomic analysis suggests that Gsdf, Bmp15, and Gdf9, originated as paralogs in vertebrate genome duplication events.ConclusionsIn zebrafish, gsdf regulates ovarian follicle maturation and expression of genes for steroid biosynthesis, obesity, diabetes, and female fertility, leading to ovarian and extra-ovarian phenotypes that mimic human polycystic ovarian syndrome (PCOS), suggesting a role for a related TGFB signaling molecule in the etiology of PCOS. Developmental Dynamics 246:925-945, 2017. © 2017 Wiley Periodicals, Inc.

Assisted reproductive technology has important clinical applications and commercial values in the horse industry. However, this approach is limited largely by the low efficiency of oocyte in vitro maturation (IVM), especially cytoplasmic maturation. To improve the efficiency of mare oocyte IVM, we evaluated the effects of co-culture with cumulus–oocyte complexes (COCs) and granulosa cells (GCs) from follicles with small (35 mm). Our results showed that oocyte nucleus maturation was not significantly improved by co-culturing with GCs. Interestingly, the cytoplasmic maturation of oocytes, defined by the distribution of cortical granules and mitochondria, as well as reactive oxygen species (ROS) levels, improved dramatically by co-culture with GCs, especially those derived from small follicles. Moreover, GCs promoted cumulus cell expansion by upregulating the expression of BMP15 in oocytes. To determine the mechanism underlying the effects of GCs, the transcriptomes of GCs from large and small follicles were compared. Expression levels of COL1A2, COL6A1, and COL6A2 were significantly higher in GCs from small follicles than in those from large follicles. These three genes were enriched in the extracellular matrix proteins-receptor interaction pathway and were involved in the regulation of collagens. Taken together, our results suggest that co-culture with GCs is beneficial to oocyte cytoplasmic maturation, and the increased expression of COL1A2, COL6A1, and COL6A2 improve the mare oocyte IVM system via the regulation of collagen.

The improvement of the reproductive traits of animals is of great interest for livestock production. Due to its positive impact on the sheep industry’s profitability, prolificacy is one of the most economically significant biological traits, showing variation between and within breeds of domestic sheep (Ovis aries). Different mutations in BMPR-1B, BMP15 and GDF9 genes coding for the transforming growth factor-β (TGFβ) superfamily have been shown to influence the ovulation rate and litter size. Numerous single-nucleotide polymorphisms (SNPs) in the bone morphogenetic protein 15 (BMP15) gene have been linked to ewe fecundity. Using targeted PCR amplification and Sanger sequencing, we were able to identify heterozygous SNPs in exon 2 of BMP15 in three sheep breeds reared in Romania: Tsigai, Cluj Merino and Tsurcana. The sequence analysis revealed three previously documented mutations, namely the missense mutation c.755T>C (L252P), which is predicted to change the tertiary structure of the BMP15 protein, and two silent mutations, c.747T>C (P249P) and c.1047G>A (V349V). In addition, we also identified one novel silent mutation, c.825G>A (S275S). Based on our findings and publicly available data, we indicate four putative mutational hotspots within exon 2 of BMP15 that could be considered for improving the indigenous sheep breeds through targeted gene editing and SNP genotyping strategies.

Prolificacy is a crucial characteristic of livestock, particularly for species such as sheep that have many births. The objectives of this study were as follows: (1) to investigate the genetic diversity of the 13 new and 7 known variants in the BMPRIB, GDF9, BMP15, LEPR, and B4GALNT2 genes in Ujimqin (UM), the F1 population of Dorper × Ujimqin crossbred (DPU), the F1 population of Suffolk × Ujimqin crossbred (SFKU), Sonid sheep (SN), Tan sheep (Tan), Hu sheep (Hu), and Small-tailed Han sheep (STH) sheep breeds/populations; (2) to perform an association analysis of the above 20 variants with litter size in 325 UM, 304 DPU, and 66 SFKU sheep populations; (3) to compare the frequencies of the litter-size-related alleles of these 20 variants among 8 sheep breeds/populations (the above seven sheep breeds + Mongolia sheep breed). With the use of the Sequenom MassARRAY®SNP assay technology, these 20 mutations were genotyped. The association analysis results showed that the c.746A>G (FecB) mutation in BMPR1B was significantly associated with the litter size of UM and DPU, the c.994A>G (FecGA) in GDF9 was significantly associated with the litter size of SFKU, and the c.31_33CTTinsdel (B1) in BMP15 was significantly associated with the litter size of UM. Our findings might provide valuable genetic markers for expanding sheep litter sizes.

Oocyte-derived bone morphogenetic protein 15 (BMP15) is one of the main local regulators of ovarian physiology, but its role in the regulation of preovulatory follicles and ovulation is not well established. Therefore, this study was conceived to determine the effect of intrafollicular injection (IFI) of BMP15 on final follicular growth, ovulation and luteinization in cattle. Initially, it was observed that relative mRNA abundance of the BMP15 receptor BMPR1B in granulosa cells was regulated by GnRH treatment, and it was negatively correlated (R2 = 0.5; P < 0.001) to progesterone concentration in follicular fluid (FF) from preovulatory follicles. The IFI of recombinant human BMP15 tended to inhibit the growth of dominant follicles, as evidenced by an average increase of only 7.7% in the follicular diameter (from 8.8 mm to 9.1 mm) at 36 h post injection compared to 36.4% increase (from 8.9 mm to 14 mm) in the control group. Injection of BMP15 into preovulatory follicles (12–14 mm), simultaneously to im GnRH treatment, inhibited ovulation compared to control group, but did not prevent luteinization and progesterone production. Most of preovulatory follicles injected with BMP15 became luteinized cysts. Collectively, these findings indicate a suppressive role of BMP15 on later follicular development and ovulation in cattle, but not on luteogenesis and progesterone secretion. • GnRH treatment decreases relative mRNA abundance of BMPR1B in granulosa cells. • BMP15 intrafollicular injection tended to inhibit follicle growth. • BMP15 intrafollicular injection inhibits GnRH-induced ovulation, but not luteinization. • The intrafollicular treatment with BMP15 induces cyst formation. [ABSTRACT FROM AUTHOR]

Sexual size dimorphism (SSD), characterized by the body size difference in different sexes, has been commonly announced in various species included mammals, birds, reptiles, and fishes. The endocrine factors in the gonads has been regarded to be involved in SSD. Two oocyte secreted factors-growth differentiation factor 9 (gdf9) and bone morphogenetic factor 15 (bmp15) has been shown to be differentially expressed in the gonad of Chinese tongue sole (Cynoglossus semilaevis), a typical marine fish demonstrating female-biased SSD. To figure out their possible roles in fish SSD, gdf9 and bmp15 of C. semilaevis were firstly cloned. The subsequently phylogenetic and structural analysis revealed that gdf9 and bmp15 were clustered with other fish species and both contained TGF-beta domain in the C-terminal. Furthermore, the temporal and spatial expression by qRT-PCR showed that gdf9 and bmp15 displayed the highest expression level in the female gonad. Moreover, the highest levels of gdf9 and bmp15 transcripts were both detected in the 1.5-year-old female gonad. The in situ hybridization and immunofluorescence experiments revealed that their mRNAs and proteins were both located in the oocyte. Based on the methylome data and bisulfite sequencing PCR, the lowest DNA methylation levels for gdf9 was observed in the female gonad, mainly distributed in the upstream and genebody regions. As for bmp15 gene, the methylation level of females in the genebody region, especially the exon 1, was higher than that of males and pseudomale, while the methylation level of females in the downstream was the lowest. Finally, knock-down of gdf9 siRNA in C. semilaevis ovarian cells resulted in the down-regulation of alk4 and tgfbr1 , and up-regulation of bmpr2 , smad8 , and bmp15. Taken together, the female-gonad-biased expression of gdf9 and bmp15 may be partly attributed to their upstream or genebody DNA methylation status. Gdf9 might be involved in reproduction and growth regulation of C. semilaevis by affecting Smad signaling pathway. Further exploration for these two ovarian factors would be helpful to better understand C. semilaevis SSD. • C. semilaevis gdf9 and bmp15 showed female-biased expression in the gonad, with mRNAs and proteins located in the oocyte. • The female-biased expression of gdf9 in the gonad may be partly attributed to its hypomethylation in the upstream region. • The gdf9 knock-down by siRNA resulted in down-regulation of alk4 and tgfbr1 , and up-regulation of bmpr2 , smad8 , and bmp15. [ABSTRACT FROM AUTHOR]

The Surti is a dual-purpose goat breed of Gujarat. The bone morphogenetic protein 15 (BMP15), a gene of transforming growth factor-beta (TGF-β) superfamily ligands, plays a role in ovulation and litter size. Mutation in the Exon-2 region of BMP15 gene with base size 575 bp increases litter size. Based on the known mutation information in goat and sheep PCR primers were designed to screen polymorphism in total 100 Surti goats, 50 Surti goats from University farm and 50 Surti goats from field units of the Southern part of Gujarat. Two polymorphic sites were found during the study, one at 500 bp and another at 400 bp. Based on this polymorphism, three genotypes were denoted as AA, AB, and AC for a wild or non-polymorphic region of 575 bp, polymorphic region at 500 bp, and polymorphic region at 400 bp, respectively. The AA, AB, and AC genotypic frequency was 0.58, 0.29, and 0.13 with average kidding rate 3.01 ± 0.09, 4.65 ± 0.32 and 2.84 ± 0.24, respectively. Polymorphic region of BMP15 gene at 500 bp (AB genotype) might be used as marker genotype for higher kidding rate at early age selection of female Surti goat. [ABSTRACT FROM AUTHOR]

Background: Bone morphogenetic protein 15 (BMP15) is expressed in oocytes and plays a crucial role in the reproduction of mono-ovulating species. In humans, BMP15 gene mutations lead to imperfect protein function and premature ovarian insufficiency. Here we investigated the BMP15 gene variants in a population of Iranian women with premature ovarian insufficiency. We conducted predictive bioinformatics analysis to further study the outcomes of BMP15 gene alterations. Methods: Twenty-four well-diagnosed premature ovarian insufficiency cases with normal karyotype participated in this study. The entire coding sequence and exon-intron junctions of the BMP15 gene were analyzed by direct sequencing. In-silico analysis was applied using various pipelines integrated into the Ensembl Variant Effect Predictor online tool. The clinical interpretation was performed based on the approved guidelines. Results: By gene screening of BMP15, we discovered p.N103K, p.A180T, and p.M184T heterozygous variants in 3 unrelated patients. The p.N103K and p.M184T were not annotated on gnomAD, 1000 Genome and/or dbSNP. These mutations were not identified in 800 Iranians whole-exome sequencing that is recorded on Iranom database. We identified the p.N103K variant in a patient with secondary amenorrhea at the age of 17, elevated FSH and atrophic ovaries. The p.M184T was detected in a sporadic case with atrophic ovaries and very high FSH who developed secondary amenorrhea at the age of 31. Conclusions: Here we newly identified p.N103K and p.M184T mutation in the BMP15 gene associated with idiopathic premature ovarian insufficiency. Both mutations have occurred in the prodomain region of protein. Despite prodomain cleavage through dimerization, it is actively involved in the mature protein function. Further studies elucidating the roles of prodomain would lead to a better understanding of the disease pathogenesis. [ABSTRACT FROM AUTHOR]

Abstract Background Atypical external genitalia are often a sign of reproductive organ pathologies and infertility with both environmental or genetic causes, including karyotypic abnormalities. Genome-wide association studies (GWAS) provide a means for identifying chromosomal regions harboring deleterious DNA-variants causing such phenotypes. We performed a GWAS to unravel the causes of incidental cases of atypically small vulvae in German Landrace gilts. Results A case-control GWAS involving Illumina porcine SNP60 BeadChip-called genotypes of 17 gilts with atypically small vulvae and 1818 control animals (fertile German Landrace sows) identified a significantly associated region on the X-chromosome (P = 8.81 × 10− 43). Inspection of whole-genome sequencing data in the critical area allowed us to pinpoint a likely causal variant in the form of a nonsense mutation of bone morphogenetic protein-15 (BMP15; Sscrofa11.1_X:g.44618787C>T, BMP15:p.R212X). The mutant allele occurs at a frequency of 6.2% in the German Landrace breeding population. Homozygous gilts exhibit underdeveloped, most likely not functional ovaries and are not fertile. Male carriers do not seem to manifest defects. Heterozygous sows produce 0.41±0.02 (P=4.5 × 10-83) piglets more than wildtype animals. However, the mutant allele’s positive effect on litter size accompanies a negative impact on lean meat growth. Conclusion Our results provide an example for the power of GWAS in identifying the genetic causes of a fuzzy phenotype and add to the list of natural deleterious BMP15 mutations that affect fertility in a dosage-dependent manner, the first time in a poly-ovulatory species. We advise eradicating the mutant allele from the German Landrace breeding population since the adverse effects on the lean meat growth outweigh the larger litter size in heterozygous sows.

The sex chromosome complement, XX or XY, determines sexual differentiation of the gonadal primordium into a testis or an ovary, which in turn directs differentiation of the germ cells into sperm and oocytes, respectively, in eutherian mammals. When the X monosomy or XY sex reversal occurs, XO and XY females exhibit subfertility and infertility in the mouse on the C57BL/6J genetic background, suggesting that functional germ cell differentiation requires the proper sex chromosome complement. Using these mouse models, we asked how the sex chromosome complement affects gene transcription in the oocytes during follicular growth. An oocyte accumulates cytoplasmic components such as mRNAs and proteins during follicular growth to support subsequent meiotic progression, fertilization, and early embryonic development without de novo transcription. However, how gene transcription is regulated during oocyte growth is not well understood. Our results revealed that XY oocytes became abnormal in chromatin configuration, mitochondria distribution, and de novo transcription compared to XX or XO oocytes near the end of growth phase. Therefore, we compared transcriptomes by RNA-sequencing among the XX, XO, and XY oocytes of 50–60 µm in diameter, which were still morphologically comparable. The results showed that the X chromosome dosage limited the X-linked and autosomal gene transcript levels in XO oocytes whereas many genes were transcribed from the Y chromosome and made the transcriptome in XY oocytes closer to that in XX oocytes. We then compared the transcript levels of 3 X-linked, 3 Y-linked and 2 autosomal genes in the XX, XO, and XY oocytes during the entire growth phase as well as at the end of growth phase using quantitative RT-PCR. The results indicated that the transcript levels of most genes increased with oocyte growth while largely maintaining the X chromosome dosage dependence. Near the end of growth phase, however, transcript levels of some X-linked genes did not increase in XY oocytes as much as XX or XO oocytes, rendering their levels much lower than those in XX oocytes. Thus, XY oocytes established a distinct transcriptome at the end of growth phase, which may be associated with abnormal chromatin configuration and mitochondria distribution. [ABSTRACT FROM AUTHOR]

Abstract Background Oocyte growth is accompanied by follicular development in mammalian ovaries. Since the discovery of two oocyte‐derived factors, growth differentiation factor 9 (GDF9), and bone morphogenetic protein 15 (BMP15), knowledge of the bidirectional communication between oocytes and granulosa cells for ovarian function and fertility has been accumulated. In addition, the growth culture system of oocytes has been improved, further promoting the studies on the communication between oocytes and granulosa cells in vitro. Methods We provide an overview of the role of granulosa cells in oocyte growth and the role of oocytes in follicular development along with our recent findings in culture experiments of bovine growing oocytes. Main findings Granulosa cells supply nutrients and metabolites through gap junctions to oocytes and secrete paracrine signals to regulate oocytes. Oocytes regulate granulosa cell proliferation and differentiation and induce antrum formation via GDF9 and BMP15. Conclusion Oocytes actively participate in various aspects of follicular development, including antrum formation via the oocyte‐derived factors GDF9 and BMP15, whose synthesis is probably regulated by granulosa cells. In vitro studies will reveal the precise communication loop between oocytes and granulosa cells that facilitates the coordinated development of oocytes and granulosa cells in the follicles.