Steroid Secretion Research Articles

Mechanism of Velvet Antler Peptide in Preventing Mild Cognitive Impairment Based on Non-Targeted Metabolomics

Velvet antler peptide (VAP) is one of the main active ingredients in Velvet antler (VA), and its mechanism in preventing mild cognitive impairment (MCI) needs to be further studied. In this research, rats model with MCI was established, non-targeted metabonomics method was performed. Total 50 wistar rats were divided into: control, model, piracetam (PIR), high- and low-dose VAP groups, and were administered for 18 days. Scopolamine (2 mg/kg) was injected intraperitoneally to establish the MCI rat model. The contents of superoxide dismutase (SOD) and malondialdehyde (MDA) in rat serum were detected by the enzyme-linked immunosorbent assay (ELISA). The expression of B-cell lymphoma/leukemia-2 (Bcl-2), Bcl-2-associated death promoter (Bad) and Bcl-2-associated X protein (Bax) in hippocampus of rats were detected by Western blot analysis. The brain and kidney of MCI rats were detected by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS) metabolomics technology, and the differences of metabolites and the dynamic changes of metabolic pathway were analyzed based on the results of Principal Component Analysis (PCA) and Orthogonal partial least squares discriminant analysis (OPLS-DA) The correlation analysis of significantly different metabolites was then analyzed by Cluster analysis (CA), and then the differential metabolite pathway enrichment analysis was performed. According to the statistical analysis of univariate and multivariates, the models constructed between the groups are stable and reliable, the variability and separation trend are good, and there are differences in the metabolomes. In results, VAP increased the content of serum SOD, reduced the content of MDA, increased the expression level of Bcl-2 and reduced the expression level of Bad and Bax protein in MCI model rats. The relevant potential markers in the brain and kidney of MCI rats mainly included folic acid, acetyl L-carnitine (ALC), Prostaglandin E1 (PGE1), 3′,5′-cyclic-adenosine-monophosphate (cAMP) and various amino acids. Therefore, VAP could inhibit oxidative stress response, reduce hippocampal neuron apoptosis and protect neurons in MCI rats. Its mechanism may be related to regulating folate and lipid metabolism, cyclic adenosine phosphate energy cycle, amino acid biosynthesis and renal secretion of sex hormones.

Menstruation and Sex Hormones in Covid-19: Effects in Women of Child-Bearing Age

Numerous studies have examined the effects of the COVID-19 pandemic on women’s mental health and menstrual cycles. However, most of these studies only included outpatients with COVID-19, whereas information on hospitalised women is minimal. ACE 2 is essential for the SAR-CoV2 virus to enter human cells and the human ovary. Since it stimulates steroid secretion, aids in follicle development and oocyte growth, influences ovulation, and maintains the corpus luteum, it stimulates steroid secretion and aids in follicle development and oocyte growth. Hospitalised female COVID-19 patients exhibit menstrual changes and heightened symptoms of mental health disorders. Length of isolation was the most influential factor in overall menstrual changes and mental health in female COVID-19 hospitalised patients. Infection with COVID-19 may affect the menstrual cycle in females. Further prospective research is required to confirm these findings and determine the duration of these menstrual irregularities.

CYP19A1 May Influence Lambing Traits in Goats by Regulating the Biological Function of Granulosa Cells.

Simple SummaryAromatase (CYP19A1), a member of the cytochrome family, is widely expressed in ovarian and granulosa cells and is primarily responsible for the conversion of androgens to estrogens. Increased expression of CYP19A1 in follicular granulosa cells has implications for cell proliferation, steroid hormone secretion, and the expression of related functional indicator genes. We hypothesize that CYP19A1 may indirectly influence lambing numbers in goats by regulating follicular cell growth and development, as well as ovarian ovulation.Abnormal expression of CYP19A1, a gene related to steroid hormone synthesis, causes steroid hormone disruption and leads to abnormal ovulation in granulosa cells. However, the exact mechanism of CYP19A1 regulation is unclear. In this study, we confirmed the localization of CYP19A1 in goat ovarian tissues using immunohistochemistry. Subsequently, we investigated the effects of CYP19A1 on granulosa cell proliferation, steroid hormone secretion, and expression of candidate genes for multiparous traits by overexpressing and silencing CYP19A1 in goat granulosa cells (GCs). The immunohistochemistry results showed that CYP19A1 was expressed in all types of follicular, luteal, and granulosa cells, with subcellular localization results revealing that CYP19A1 protein was mainly localized in the cytoplasm and nucleus. Overexpression of CYP19A1 significantly increased the mRNA levels of CYP19A1, FSHR, and INHBA, which are candidate genes for multiple birth traits in goats. It also promoted cell proliferation, PCNA and Cyclin E mRNA levels in granulosa cells, and secretion of estrogen and progesterone. However, it inhibited the mRNA levels of STAR, CYP11A1, and 3βSHD, which are genes related to steroid synthesis. Silencing CYP19A1 expression significantly reduced CYP19A1, FSHR, and INHBA mRNA levels in granulosa cells and inhibited granulosa cell proliferation and PCNA and Cyclin E mRNA levels. It also reduced estrogen and progesterone secretion but enhanced the mRNA levels of STAR, CYP11A1, and 3βSHD. CYP19A1 potentially influenced the lambing traits in goats by affecting granulosa cell proliferation, hormone secretion, and expression of candidate genes associated with traits for multiple births.

Transcriptomic diversification of granulosa cells during follicular development between White Leghorn and Silky Fowl hens

Egg production rate in chicken is related to the continuity of follicle development. In this study, we found that the numbers of white prehierarchical, dominant, and yellow preovulatory follicles in the high-yielding layer breed, White Leghorn (WL), were significantly higher than those in the low egg-yielding variety, Silky Fowl (SF). The proliferation and differentiation of granulosa cells (GCs) play an important role in follicle maturation. Histological observation revealed a large number of melanocytes in the outer granulosa layer of follicles in SF but not in WL. Finally, RNA-sequencing was used to analyze the gene expression profiles and pathways of the GC layer in the follicles in both WL and SF hens. Transcriptome analysis of prehierarchical GCs (phGCs) and preovulatory GCs (poGCs) between WL and SF showed that steroid hormone-, oxytocin synthesis-, tight junction-, and endocytosis-related genes were expressed at higher levels in WL phGCs than in SF phGCs, whereas the insulin signaling pathway- and vascular smooth muscle contraction-related genes were upregulated in SF phGCs. Fatty acid synthesis, calcium signaling, and Wnt signaling pathway-related genes were expressed at higher levels in WL poGCs than in SF poGCs; however, adrenergic signaling, cGMP-PKG, and melanogenesis-related genes were upregulated in SF poGCs. These results indicate that genes that promote GC proliferation and secretion of various sex hormones are more active in WL than in SF hens. The upregulated signaling pathways in SF help in providing energy to GCs and for angiogenesis and melanogenesis. In vitro experiments confirmed that both the proliferation of poGCs and synthesis of reproductive hormones were higher in WL than in SF hens.

Ovary-derived Decellularized Extracellular Matrix-based Bioink for Fabricating 3D Primary Ovarian Cells-laden Structures for Mouse Ovarian Failure Correction.

Fertility preservation is becoming a clinical duty in practice. Three-dimensional (3D) bioprinting technology is potentially realize ovarian morphological repair and reproductive endocrine function rebuild. There is no published work on 3D bioprinting ovary using a decellularized extracellular matrix (dECM)-based bioink, though dECM is the preferred matrix choice for an artificial ovary. The study aimed to explore swine ovarian dECM-based bioink to fabricate 3D primary ovarian cells (POCs)-laden structures for mouse ovarian failure correction. In this study, the ovarian dECM was converted to dECM-based bioink by dECM solution mixed with a seaweed gelatin blend solution of bioink that was characterized using scanning electron microscopy, circular dichroism, rheology, hematoxylin and eosin staining, and immunohistochemistry. The 3D scaffolds were, then, printed with or without POCs by the extrusion 3D bioprinter. The laden POCs viability was detected with the live/dead assay kit. A female castrated mouse model was established, and the mice were treated with five different methods. The results revealed that the 3D scaffold encapsulating POCs group had more positive signals in neoangiogenesis, cell proliferation and survival than the 3D scaffold group, and ensured sex hormone secretion. Meanwhile, the expression of germ cells in the 3D scaffold encapsulating POCs group was more intensely than the non-printed hydrogel encapsulating POCs group. The work shows that the 3D bioprinting ovary employing ovarian dECM-based bioink is a promising approach for ovarian failure correction.

Development of Economic Learning Innovation Using Lectora Inspire Media on Demand and Supply Materials for Class X Muhammadiyah Senior High School 2 of Kertosono

Hypogonadism in female is one of the causes of menstrual irregularities and infertility. This clinical syndrome is characterized by decreased sex steroid secretion either due to primary gonadal failure or secondarily due to central hypothalamic-pituitary causes. In our study, it is found that patients with diabetes have primary ovarian failure with low estradiol and high FSH, LH values. [Learning media, Lectora Inspire, Computer Hardware]

Expression and distribution of MTNR1A in the hypothalamus-pituitary-gonadal axis of Tibetan sheep during the estrous cycle

The melatonin 1A receptor (MTNR1A) is a membrane receptor distributed across the mammalian gonadal axis-associated membrane. Melatonin (MT) can specifically bind with MTNR1A on the cell membrane and regulates mammalian reproductive activities. However, the role of MTNR1A in regulating the reproductive physiological activities of sheep in the Tibetan Plateau remains unclear. In this study, the MT content in Tibetan sheep blood during the estrous cycle was detected by ELISA. The distribution of MTNR1A in the hypothalamus-pituitary–gonadal axis (HPGA) was analyzed by immunohistochemistry and immunofluorescence. Western blot and qRT-PCR were used to detect dynamic changes of MTNR1A mRNA and protein expression, and the protein distributions in the HPGA. The results showed that the average secretion level of MT in Tibetan sheep blood was highest occurred during diestrus and the lowest during proestrus. Additionally, the secretion of MT at night was significantly higher than during the day. The immunopositive products of MTNR1A were primarily distributed around the glial cells in the dorsal hypothalamic nucleus region, chromophobe cells, and eosinophilic cytoplasm in the pituitary gland, follicular granular layer, follicular adventitia, tubal mucosa, cilia, endometrium, interstices, and glands in the uterus. The expression trends of MTNR1A mRNA and proteins in the HPGA during the estrous cycle were the same. The relative expression levels of MTNR1A mRNA and proteins in the hypothalamus and ovaries were the highest during proestrus and the lowest during metestrus; the highest during diestrus in the pituitary and oviducts; the highest during metestrus in the uterus. Collectively, the differences in the secretion of MT in Tibetan sheep blood and the expression of MTNR1A in HPGA suggest that they may be affected by steroid hormone secretion during the estrous cycle of Tibetan sheep, which has a potential impact on the regulation of animal estrous cycle.

Effects of glucose on glycolysis and steroidogenesis as well as related gene expression in ovine granulosa cells in vitro

Our previous studies indicate that short-term dietary supplementation of ewes during the luteal phase can increase fertility due to elevated glucose and steroid hormone concentrations in follicles. However, the regulatory relationship between glucose and steroidogenesis during follicular development has not been clarified. The present study was conducted to determine the direct effects of different glucose levels on glycolysis and steroid secretion in ovarian granulosa cells (GCs) obtained from ewes, using an in vitro cell culture system. The GCs were collected from small follicles (1–3 mm) and treated with various doses of glucose (0, 2.1, 4.2, 8.4, 16.8 and 33.6 mM) in serum-free medium for 24 h. We observed that different dose of glucose increased the concentrations of lactic acid (LA) (linear and quadratic, P < 0.05), pyruvic acid (PA) (linear and quadratic, P < 0.05), oestradiol (E2) (linear and quadratic, P < 0.05) and progesterone (P4) (linear and quadratic, P < 0.05). The concentrations of PA, E2 and P4 all reached the highest levels in the 8.4 mM glucose group. Further study found that the mRNA and protein expression levels of glycolysis- related (LDHA and PKM2) and steroidogenesis- related (STAR, CYP11A1 and CYP19A1) genes increased with increasing glucose concentration up to 8.4 mM and then decreased at higher concentrations. There were positive correlations of glycolytic metabolites with steroidogenesis (P < 0.001, the range of r is: 0.746–0.929). This study showed that glucose dose had a significant effect on glycolysis synthesis and steroidogenesis of GCs, and 8.4 mM represented an optimum concentration for the secretion of steroid hormones by ovine GCs in vitro.

Forskolin Induces Endocrine Disturbance in Human JEG-3 Placental Cells.

Forskolin, used in folk medicine since ancient times, is now available as a dietary supplement, with an indication as a fat burner and appetite suppressant. However, the safety of forskolin is poorly documented especially for pregnant women. The question that we raised is what about the safety of forskolin in pregnant women? As the placenta, an endocrine organ, is the key organ of pregnancy, we evaluated the in vitro placental toxicity of forskolin. We focused first on the activation of a P2X7 degenerative receptor as a key biomarker for placental toxicity, and second on steroid and peptide hormonal secretion. We observed that forskolin activated P2X7 receptors and disturbed estradiol, progesterone, hPL and hyperglycosylated hCG secretion in human placental JEG-Tox cells. To the best of our knowledge, we highlighted, for the first time, that forskolin induced endocrine disturbance in placental cells. Forskolin does not appear to be a safe product for pregnant women and restrictions should be taken.

Molecular Cloning and Expression Analysis of Thyrotropin-Releasing Hormone, and Its Possible Role in Gonadal Differentiation in Rice Field eel Monopterus albus.

Simple SummaryThyrotropin-releasing hormone (TRH) is an important upstream regulator in the hypothalamus-pituitary-thyroid (HPT) axis in mammals. In this study, we isolated and characterized trh gene from a protogynous hermaphrodite fish rice field eel Monopterus albus. TRH had no significant effect on serum thyroid hormone levels in rice field eel. However, we found that TRH was involved in the regulation gonadal differentiation-related gene expression and serum sex steroid hormone secretion. Our results indicated that TRH may play a novel role in gonadal differentiation in rice field eel.Rice field eel (Monopterus albus), a protogynous hermaphrodite fish, is a good model for the research of sex determination and gonadal differentiation in teleosts. In this study, we cloned the full-length cDNA sequence of trh, which encoded a predicted protein with 270 amino acids. Trh mainly expressed in the brain, followed by the ovary, testis, muscle and pituitary, and had low levels in other peripheral tissues. During natural sex reversal, trh mRNA expression levels exhibited a significant increase at the late intersexual stage in the hypothalamus. In the gonad, trh mRNA expression levels showed a trend of increase followed by decrease, and only increased significantly at the middle intersexual stage. No matter static incubation or intraperitoneal (IP) injection, TRH had no significant effect on trh and thyroid-stimulating hormone β subunit (tshβ) mRNA expression levels, and serum T3, T4 and TRH release. After static incubation of ovarian fragments by TRH, the expression of gonadal soma derived factor (gsdf) was up-regulated significantly at both the doses of 10 and 100 nM. IP injection of TRH stimulated the expression of gsdf, and inhibited the expression of ovarian aromatase gene (cyp19a1a), accompanied by the increase of serum 11-KT levels. The results indicated that TRH may play a novel role in gonadal differentiation by the regulation of gonadal differentiation-related gene expression and sex steroid hormone secretion in rice field eel.

Clinical profile, etiology, outcome and new-onset diabetes: A SARI case series

Introduction: The world is currently facing the SARS-CoV-2 pandemic with evolving 2nd wave. The COVID-19 patients present most commonly with Severe Acute Respiratory Illness (SARI) in an emergency room with acute onset fever, cough, and breathlessness. However, not all SARI cases as per definition are due to COVID-19 infection, which is well proven in this case series of 113 cases of SARI. This is just the opposite of the other SARI series done in the pre-COVID-19 era. Also, no previous SARI case series data has shown significant association with Diabetes Mellitus, including new-onset diabetes thus figuring out the major Pathophysiological association of COVID-19 with glucose metabolism and has a bearing on the pathogenesis, treatment, and outcome of COVID-19 infection and perpetuity of pandemic of this magnitude. Here we raise concern for the first time about the growing association of an infectious pandemic with the lifestyle disorders which are non-communicable diseases but carry with them the potential of fertile soil for rapidly spreading epidemics. Aim and objective: To find out the etiology, clinical profile, treatment outcome, and mortality rate in different sub-groups of SARI cases in a tertiary care hospital and the incidence of new-onset Diabetes Mellitus in them and to investigate theoretically the hypothesis that maintaining normal glucose metabolism could prevent progression of a mild Flu-like illness (FLI) to a severe form of Severe Acute Respiratory Illness (SARI) and consequent complications such as Cytokine Storm Syndrome and Multi-Organ failure. Design: Retrospective, single-center case series of 113 SARI patients at a tertiary care hospital in Agra India between 1 March- 30 October 2020. Main outcome: The demographics, clinical, pathological, imaging, and treatment outcome data were collected. The SARI cases analyzed were defined as “Severe acute respiratory infections (SARIs) an acute respiratory illness of recent onset (within seven days) manifested by fever (≥ 38 °C), cough and shortness of breath or difficulty in breathing requiring hospitalization and were sub-classified according to the primary etiology producing SARI in them. The findings were compiled and compared. Conclusion: Of the 113 patients of SARI – 32.7% were associated with Diabetes, with 9.74% new-onset Diabetes and 26% previously known Diabetes. This was mainly due to SARS-CoV-2 (24 Diabetics out of 52 COVID-19 cases- 46.1%). The Average hospitalization stay of SARI cases was 10 days with a maximum in SARS-CoV-2 and a minimum stay of 5.22 days in Bacterial Pneumonia and 5.66 days in Koch’s Lungs. The death rate was maximum (4 out of 26) 15.3%. Hospitalized TB/Koch’s Lung patients who presented as SARI and 3.8% in Bacterial Pneumonia, 2.43% in SARS-CoV-2, and &lt; 1% in Sepsis. Those SARI cases who were euglycemic at the time of initial presentation recovered early and carried a good prognosis with less mortality as compared to those who were hyperglycemic on presentation. Also, those FLI cases who maintained euglycemia or did not have any other risk factor which predisposes them to stress (Diabetes, Prolonged fasting, Obesity, major organ disorder, Psychological disorder, and Cancer) did not progress to SARI as the endogenous steroid secretion and sympathetic activation did not occur, the intracellular pH levels remained in the alkaline range. 10.18% of cases developed new-onset diabetes (a total of 11 cases) out of which 10 were in COVID-19. Thus 19.2% incidence of new-onset diabetes in SARS-CoV-2 and a prevalence of 26.9% in SARS-CoV-2, making total diabetes 46.1% in SARS-CoV-2, and out of all SARI cases, 26% of patients developed pulmonary fibrosis with consequent long-term complications. In COVID-19 patients, it was seen only in diabetics SARS CoV-2 male patients, thus no death in non-diabetic females in COVID-19 in this case series.

A Proposal for Nomenclature Revision of Nonfunctioning Adrenal Incidentalomas as Adrenal Lesions of Undetermined Secretion of Adrenal Steroids (ALUSAS)

In this commentary we propose a change of the nomenclature of non-functioning adrenal incidentalomas (NFAI) to adrenal lesions of undetermined secretion of adrenal steroids (ALUSAS) since there are mounting evidence suggesting that a substantial proportion of so-called NFAIs do actually secrete steroids in excess. In fact, it is known that some NFAIs develop ACS over time. In the commentary we justify the change of term by summarizing the evidence that shows that NFAIs present a higher cardiometabolic risk than the general population, and the metabolomics studies in which an increase in cortisol metabolites is observed in patients with NFAI.

Deciphering Sex-Specific Differentiation of Human Fetal Gonads: Insight From Experimental Models.

Sex-specific gonadal differentiation is initiated by the expression of SRY in male foetuses. This promotes a signalling pathway directing testicular development, while in female foetuses the absence of SRY and expression of pro-ovarian factors promote ovarian development. Importantly, in addition to the initiation of a sex-specific signalling cascade the opposite pathway is simultaneously inhibited. The somatic cell populations within the gonads dictates this differentiation as well as the development of secondary sex characteristics via secretion of endocrine factors and steroid hormones. Opposing pathways SOX9/FGF9 (testis) and WNT4/RSPO1 (ovary) controls the development and differentiation of the bipotential mouse gonad and even though sex-specific gonadal differentiation is largely considered to be conserved between mice and humans, recent studies have identified several differences. Hence, the signalling pathways promoting early mouse gonad differentiation cannot be directly transferred to human development thus highlighting the importance of also examining this signalling in human fetal gonads. This review focus on the current understanding of regulatory mechanisms governing human gonadal sex differentiation by combining knowledge of these processes from studies in mice, information from patients with differences of sex development and insight from manipulation of selected signalling pathways in ex vivo culture models of human fetal gonads.

Mutual Shaping of Circadian Body-Wide Synchronization by the Suprachiasmatic Nucleus and Circulating Steroids.

BackgroundSteroids are lipid hormones that reach bodily tissues through the systemic circulation, and play a major role in reproduction, metabolism, and homeostasis. All of these functions and steroids themselves are under the regulation of the circadian timing system (CTS) and its cellular/molecular underpinnings. In health, cells throughout the body coordinate their daily activities to optimize responses to signals from the CTS and steroids. Misalignment of responses to these signals produces dysfunction and underlies many pathologies.Questions AddressedTo explore relationships between the CTS and circulating steroids, we examine the brain clock located in the suprachiasmatic nucleus (SCN), the daily fluctuations in plasma steroids, the mechanisms producing regularly recurring fluctuations, and the actions of steroids on their receptors within the SCN. The goal is to understand the relationship between temporal control of steroid secretion and how rhythmic changes in steroids impact the SCN, which in turn modulate behavior and physiology.Evidence SurveyedThe CTS is a multi-level organization producing recurrent feedback loops that operate on several time scales. We review the evidence showing that the CTS modulates the timing of secretions from the level of the hypothalamus to the steroidogenic gonadal and adrenal glands, and at specific sites within steroidogenic pathways. The SCN determines the timing of steroid hormones that then act on their cognate receptors within the brain clock. In addition, some compartments of the body-wide CTS are impacted by signals derived from food, stress, exercise etc. These in turn act on steroidogenesis to either align or misalign CTS oscillators. Finally this review provides a comprehensive exploration of the broad contribution of steroid receptors in the SCN and how these receptors in turn impact peripheral responses.ConclusionThe hypothesis emerging from the recognition of steroid receptors in the SCN is that mutual shaping of responses occurs between the brain clock and fluctuating plasma steroid levels.

Effects of a GnRH agonist on sex behavior in females of the southern giant pouched rat.

In general, males should be particularly attentive to cues of sexual availability and females should advertise accordingly. Vaginal patency (i.e., the openness of the vagina) is a reliable indicator of sexual maturity; if the vagina is closed, the female is unable to copulate. The southern giant pouched rat (Cricetomys ansorgei) is unusual because females can have fully fused vaginal openings (i.e., vaginal nonpatency) despite being considered 'adults' by other metrics. Moreover, some females reversibly close their vaginal openings. Thus, vaginal patency in the pouched rat is a 'flexible' reproductive state. We subcutaneously implanted a long-acting GnRH agonist (deslorelin), which over time inhibits sex steroid secretion, to better understand the endocrinology and social behavior relating to vaginal patency in this species. We hypothesized that altering GnRH would impact both patency and behavior through its effects on circulating levels of estradiol. Six months of deslorelin treatment did not alter vaginal patency. Behaviorally, deslorelin-treated females spent less time interacting with, and were more aggressive towards males (compared to controls). Notably, deslorelin did not alter female scent marking. We conclude that behavioral receptivity, but not vaginal patency, is impacted by GnRH hormonal cascades in the pouched rat.

Caught in the Middle: The Care of Transgender Youth in Texas.

In the US, approximately 0.7% of adolescents identify as transgender or gender fluid.1 Many pediatric centers now provide multidisciplinary care to assist in the transition from a child's or adolescent's assigned sex at birth (or natal sex) to their affirmed (or preferred) gender. During early puberty, therapy is available that results in the suppression of sex steroid production and secretion, followed later in adolescence by the administration of supraphysiologic doses of sex hormones appropriate for their gender identity.1-3 The duration of pubertal suppression with gonadotropin hormone releasing hormone agonists (GnRHa) varies, but can extend up to 4 years for younger patients who are not able to provide consent until age 16 for receipt of gender-affirming therapy. Puberty blockers represent an invaluable intervention for these children and adolescents, to reduce anxiety and "buy time" until final decisions can be made about gender assignment.

The transcriptome-wide N6-methyladenosine (m6A) map profiling reveals the regulatory role of m6A in the yak ovary

Background and aimYak estrus is a seasonal phenomenon, probably involving epigenetic regulation of synthesis and secretion of sex hormones as well as growth and development of follicles. N6-methyladenosine (m6A) is the most common internal modification of the eukaryotic mRNA. However, there are no detailed reports on the m6A transcriptome map of yak ovary. Therefore, this study aimed to collected the yak ovarian tissues at three different states of anestrus (YO-A), estrus (YO-F), and pregnancy (YO-P), and obtained the full transcriptome m6A map in yak by MeRIP-seq.ResultsThe HE staining revealed that the number of growing follicles and mature follicles in the ovary during the estrus period was relatively higher than those in the anestrus period and the pregnancy period. The RT-qPCR showed that the expression of METTL3, METTL14, FTO, YTHDC1 were significantly different across different periods in the ovaries, which suggests that m6A may play a regulatory role in ovarian activity. Next, we identified 20,174, 19,747 and 13,523 m6A peaks in the three ovarian samples of YO-A, YO-F and YO-P using the methylated RNA immunoprecipitation sequencing (MeRIP-seq). The m6A peaks are highly enriched in the coding sequence (CDS) region and 3′untranslated region (3′UTR) as well as the conserved sequence of “RRACH.” The GO, KEGG and GSEA analysis revealed the involvement of m6A in many physiological activities of the yak’s ovary during reproductive cycle. The association analysis found that some genes such as BNC1, HOMER1, BMP15, BMP6, GPX3, and WNT11 were related to ovarian functions.ConclusionsThe comparison of the distribution patterns of methylation peaks in the ovarian tissues across different periods further explored the m6A markers related to the regulation of ovarian ovulation and follicular development in the yak ovary. This comprehensive map provides a solid foundation for revealing the potential function of the mRNA m6A modification in the yak ovary.

In Vitro Pharmacological Characterization of CRN04894: The First Reported Oral, Selective Nonpeptide Melanocortin 2 Receptor Antagonist Evaluated in Phase 1 First‐in‐Human Clinical Trials

Adrenocorticotropic hormone (ACTH) is secreted from the pituitary gland and selectively activates the melanocortin type 2 receptor (MC2R) in the adrenal glands, where it modulates steroidal hormone (e.g., cortisol) synthesis and secretion. Excess ACTH contributes to the pathophysiology of Cushing’s disease (CD), ectopic ACTH syndrome (EAS), and congenital adrenal hyperplasia (CAH). ACTH‐secreting tumors cause CD (in pituitary) and EAS (outside pituitary) and lead to hypercortisolemia and various metabolic comorbidities. CAH is caused by inactivating mutations in adrenal steroid synthesis pathway proteins leading to low levels of cortisol that are insufficient to provide negative feedback on ACTH production. Chronic ACTH elevation can lead to life‐threatening salt wasting and significant virilization. We hypothesize that blocking the ACTH‐MC2R interaction with a selective MC2R antagonist may provide a key new therapeutic option for these patients.We initiated an iterative medicinal chemistry program to identify potent and selective nonpeptide MC2R antagonists suitable for evaluation in clinical trials. Initial assessment consisted of equilibrium radioligand binding competition assays using [125I]ACTH(1‐39) to determine Ki and EC50 shift functional assays using ACTH(1‐24) to determine ΔpEC50 values. Select compounds were then characterized using a full Schild regression analysis to determine KB values and gain insight into mechanism. There was a strong correlation between the Ki and KB values for this class of small molecule.Our efforts identified CRN04894 as a highly potent MC2R antagonist (Ki = 2.1 nM, KB = 0.34 nM). The Schild regression data for CRN04894 was consistent with a competitive, orthosteric interaction with ACTH. The individual ACTH dose‐response curves with the addition of different concentrations of CRN04894 showed the same maximum response and slopes near unity, and the Schild plot had a slope not statistically different than unity. Furthermore, CRN04894 did not affect the dissociation rate of [125I]ACTH(1‐39) initiated by a saturating concentration of ACTH(1‐39), also consistent with an orthosteric interaction. Because CRN04894 appeared to bind the orthosteric site on MC2R, it was further tested in a kinetic radioligand binding competition assay using [125I]ACTH(1‐39) to determine its binding rate constants. CRN04894 rate constants are comparable to published values for other GPCR ligands with kon = 1.2 e7 M‐1 min‐1 and koff = 0.018 min‐1, corresponding to a dissociation t1/2of 39 min. The kinetic Ki (1.5 nM) calculated from the kon and koff is in accordance with the equilibrium competition binding Ki and Schild analysis KB. In addition, CRN04894 selectively blocks the ACTH‐MC2R interaction, showing no binding to MC1R, MC3R, MC4R, and MC5R at concentrations ≥ 1 µM. To our knowledge, this class of molecule represents the first reported potent nonpeptide MC2R antagonists, and CRN04894 is currently being evaluated in a first‐in‐human phase 1 clinical trial evaluating pharmacokinetics, pharmacodynamics, and tolerability in healthy subjects.

Winter madness: Melatonin as a neuroendocrine regulator of seasonal aggression.

Individuals of virtually all vertebrate species are exposed to annual fluctuations in the deterioration and renewal of their environments. As such, organisms have evolved to restrict energetically expensive processes and activities to a specific time of the year. Thus, the precise timing of physiology and behavior is critical for individual reproductive success and subsequent fitness. Although the majority of research on seasonality has focused on seasonal reproduction, pronounced fluctuations in other non-reproductive social behaviors, including agonistic behaviors (e.g., aggression), also occur. To date, most studies that have investigated the neuroendocrine mechanisms underlying seasonal aggression have focused on the role of photoperiod (i.e., day length); prior findings have demonstrated that some seasonally breeding species housed in short "winter-like" photoperiods display increased aggression compared with those housed in long "summer-like" photoperiods, despite inhibited reproduction and low gonadal steroid levels. While fewer studies have examined how the hormonal correlates of environmental cues regulate seasonal aggression, our previous work suggests that the pineal hormone melatonin acts to increase non-breeding aggression in Siberian hamsters (Phodopus sungorus) by altering steroid hormone secretion. This review addresses the physiological and cellular mechanisms underlying seasonal plasticity in aggressive and non-aggressive social behaviors, including a key role for melatonin in facilitating a "neuroendocrine switch" to alternative physiological mechanisms of aggression across the annual cycle. Collectively, these studies highlight novel and important mechanisms by which melatonin regulates aggressive behavior in vertebrates and provide a more comprehensive understanding of the neuroendocrine bases of seasonal social behaviors broadly.

Targeted metabolomics analysis of bile acids and cell biology studies reveal the critical role of glycodeoxycholic acid in buffalo follicular atresia

The follicular fluid of mammals has a high abundance of bile acids and these have proven to be closely related to the follicular atresia. However, the origin and content of bile acids in follicular fluid and its mechanisms on follicular atresia remain largely unknown. In this work, we analyzed the origin of bile acids in buffalo follicles by using cell biology studies, and quantified the subspecies of bile acids in follicular fluid from healthy follicles (HF) and atretic follicles (AF) by targeted metabolomics. The function of differential bile acids on follicular granulosa cells was also studied. The results showed that the bile acids transporters were abundantly expressed in ovarian tissues, but the rate-limiting enzymes were not, which was consistent with the inability of cultured follicular cells to convert cholesterol into bile acids. Targeted metabolomics analysis revealed thirteen differential subspecies of bile acids between HF and AF. The free bile acids were significant down-regulated and their conjugated forms were significantly up-regulated in AF as compared to HF. Finally, cell biological validation found a specific differentially conjugated bile acid, glycodeoxycholic acid (GDCA), which could promote follicular granulosa cell apoptosis and reduce steroid hormone secretion. In summary, our studies suggest that bile acids in buffalo follicles are transported from the blood rather than being synthesized within the follicles. The conjugated bile acids such as GDCA, accumulate in buffalo follicles, and may accelerate atresia by promoting apoptosis of granulosa cells and inhibiting steroid hormone production. These results will provide new clues for studying the physiological role and mechanism of bile acids involved in buffalo follicular atresia.