Steroid Synthesis Research Articles

The toxicological effects of life-cycle exposure to harmful benthic cyanobacteria Oscillatoria on zebrafish growth and reproduction: A comparative study with planktonic Microcystis

The risks of planktonic cyanobacteria blooms have been the focus of much scientific research, but studies on the ecotoxicological effects of benthic cyanobacteria are lagging. The impacts of cyanobacteria cells on fish populations might be more complex in contrast to purified cyanotoxins or cyanobacteria extracts. This study systematically compared the chronic effects of benthic Oscillatoria sp. (producing cylindrospermopsins) and planktonic Microcystis aeruginosa (producing microcystins) on the growth and reproduction of zebrafish through life-cycle exposure (5– 90 days post fertilization). The results showed that both Oscillatoria sp. and M. aeruginosa exposure caused growth inhibition and fecundity reduction in F0 generation by disrupting sex hormone levels, delayed ovarian and sperm development, and induced pathological lesions in zebrafish gonads. Furthermore, exposure to Oscillatoria sp. or M. aeruginosa in adult zebrafish increased mortality and teratogenicity in F1 embryos (without exposure), indicating a parental transmission effect of developmental toxicity. The difference was that M. aeruginosa exposure led to significant alterations in pathways, such as tissue development, redox processes, and steroid hormone synthesis. In contrast, Oscillatoria sp. exposure primarily disrupted the PPAR signaling pathway, cell adhesion molecules, and lipid transport pathways. Interestingly, the differentially expressed genes revealed that male fish were more sensitive to harmful cyanobacteria than females, whether exposed to Oscillatoria sp. or M. aeruginosa. These findings contribute to a better mechanistic understanding of the chronic toxic effects of distinct types of harmful cyanobacteria, suggesting that the ecological risk of benthic cyanobacteria requires further attention.

Glucocorticoids improve sperm performance in physiological and pathological conditions: their role in sperm fight/flight response.

Glucocorticoids play a physiologic role in the adult male reproductive functions, modulating gonadal steroid synthesis and spermatogenesis, through the glucocorticoid receptor (GR). The expression of GR has been described in several key testicular cell types, including somatic cells and early germ cell populations. Nothing is known on GR in human spermatozoa. Herein, we explored the GR expression and its possible role in normal and testicular varicocele semen samples from volunteer donors. After semen parameter evaluation by macro- and microscopic analysis, samples were centrifuged; then spermatozoa and culture media were recovered for further investigations. By western blotting and immunofluorescence analyses we evidenced for the first time in spermatozoa the presence of GR-D3 isoform which was reduced in sperm from varicocele patients. By treating sperm with the synthetic glucocorticoid dexamethasone (DEXA), we found that survival, motility, capacitation, and acrosome reaction were increased in both healthy and varicocele samples. GR involvement in mediating DEXA effects, was confirmed by using the GR inhibitor mifepristone (M2F). Worthy, we also discovered that sperm secretes different cortisol amounts depending on its physio-pathological status, suggesting a defence mechanism to escape the immune system attach in the female genital tract thus maintaining the immune-privilege as in the testis. Collectively, our data suggests a role for glucocorticoids in determining semen quality and function, as well as in participating on sperm immune defensive mechanisms. The novelty of this study may be beneficial and needs to take into account in artificial insemination/drug discovery aimed to enhancing sperm quality.

NPC1L1 Plays a Novel Role in Nonalcoholic Fatty Liver Disease.

Niemann-Pick C1-Like 1 (NPC1L1) is a key protein in the transport of cholesterol, which exists in the brush marginal membrane of the intestinal epithelial cells and the timid duct membrane of the liver. It affects cholesterol absorption and plasma low-density lipoprotein levels. Cholesterol is both an important component of the cell membrane and a precursor of bile acid and steroid hormone synthesis. Abnormal cholesterol metabolism is closely related to nonalcoholic steatohepatitis (NASH). NASH can progress to fibrosis and cirrhosis, with serious consequences. NPC1L1 is involved in the regulation of cholesterol and lipid metabolism and plays an important role in maintaining the balance of cholesterol metabolism in the body. It also plays an important role in some metabolic diseases such as nonalcoholic fatty liver disease, obesity, and hypercholesterolemia. Therefore, it is necessary to elucidate the molecular pathological mechanism of NPC1L1 in the regulation of cholesterol metabolism and the occurrence and development of NASH, which can provide a target for the development of novel drugs for the treatment of NASH and other diseases. More importantly, it helps to accelerate the development of drugs that regulate lipid metabolism at multiple levels and reduce liver steatosis, which is extremely important for the prevention and treatment of NASH and related severe metabolic diseases.

Embryonic Exposure to the Benzotriazole Ultraviolet Stabilizer 2-(2H-benzotriazol-2-yl)-4-methylphenol Decreases Fertility of Adult Zebrafish (Danio rerio).

Benzotriazole ultraviolet stabilizers (BUVSs) are emerging contaminants of concern. They are added to a variety of products, including building materials, personal care products, paints, and plastics, to prevent degradation caused by ultraviolet (UV) light. Despite widespread occurrence in aquatic environments, little is known regarding the effects of BUVSs on aquatic organisms. The aim of the present study was to characterize the effects of exposure to 2-(2H-benzotriazol-2-yl)-4-methylphenol (UV-P) on the reproductive success of zebrafish (Danio rerio) following embryonic exposure. Embryos were exposed, by use of microinjection, to UV-P at <1.5 (control), 2.77, and 24.25 ng/g egg, and reared until sexual maturity, when reproductive performance was assessed, following which molecular and biochemical endpoints were analyzed. Exposure to UV-P did not have a significant effect on fecundity. However, there was a significant effect on fertilization success. Using UV-P-exposed males and females, fertility was decreased by 8.75% in the low treatment group and by 15.02% in the high treatment group relative to control. In a reproduction assay with UV-P-exposed males and control females, fertility was decreased by 11.47% in the high treatment group relative to the control. Embryonic exposure to UV-P might have perturbed male sex steroid synthesis as indicated by small changes in blood plasma concentrations of 17β-estradiol and 11-ketotestosterone, and small statistically nonsignificant decreases in mRNA abundances of cyp19a1a, cyp11c1, and hsd17b3. In addition, decreased transcript abundances of genes involved in spermatogenesis, such as nanos2 and dazl, were observed. Decreases in later stages of sperm development were observed, suggesting that embryonic exposure to UV-P impaired spematogenesis, resulting in decreased sperm quantity. The present study is the first to demonstrate latent effects of BUVSs, specifically on fish reproduction. Environ Toxicol Chem 2024;43:385-397. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Rfamide-related peptide-3(RFRP-3) receptor gene is expressed in mouse ovarian granulosa cells: Potential role of RFRP-3 in steroidogenesis and apoptosis

RFRP-3 is a functional ortholog of avian GnIH and regulates reproductive activities in the gonads of animals. However, the role of RFRP-3 in the function of ovarian granulosa cells in mice remains unclear. First, we detected the expression of the RFRP-3 receptor (GPR147) in the ovarian granulosa cells of mice. Second, the effect of RFRP-3 treatment on estradiol and progesterone secretions from granulosa cells was tested by ELISA. Meanwhile, the expression of genes and proteins regulating steroid hormone synthesis was respectively examined by qPCR and western blot. Furthermore, the effect of RFRP-3 treatment on the apoptosis of granulosa cells was analyzed. The results revealed that the GPR147 protein (a RFRP-3 receptor) was expressed in the ovarian granulosa cells of mice. Low and medium doses RFRP-3 treatment significantly reduced progesterone secretion in the granulosa cells (P < 0.05), while RFRP-3 suppressed p450scc, 3β-HSD, StAR, and FSHR expression in a non-dose-dependent manner. Moreover, RFRP-3 treatment might induce the apoptosis of granulosa cells. Additionally, low doses RFRP-3 significantly reduced p-ERK1/2 protein expression (P < 0.05) in the ovarian granulosa cells. We here, for the first time, confirmed that GPR147 was expressed in the ovarian granulosa cells of mice. Our findings suggested that and RFRP-3 regulates the granulosa cell function through the ERK signaling pathway, which will lay the foundation for uncovering molecular mechanisms by which RFRP-3 regulates follicle development in future.

The Developmental Toxicity and Endocrine-Disrupting Effects of Fenpropathrin on Gobiocypris rarus during the Early Life Stage.

In the present study, the developmental toxicity and endocrine-disrupting effects of fenpropathrin on Gobiocypris rarus during the early life stage were studied using a semi-static water exposure method. The results showed that the LOEC (lowest observed effect concentration) of fenpropathrin on the incubation of rare minnow embryos was above 2.5 μg·L-1. The LOEC and NOEC (no observed effect concentration) of fenpropathrin on the developmental malformations and death indicators were 2.0 and 1.5 μg·L-1, respectively. After exposure to 1.5 μg·L-1 of fenpropathrin for 31 days, the expressions of androgen receptor genes (AR) and sex hormone-synthesis-related genes (CYP17 and CYP19a) were significantly decreased and the expressions of thyroid hormone receptor genes (TRβ) and aryl hydrocarbon receptor genes (AhR1a and AhR2) were significantly increased in juvenile Gobiocypris rarus. The expression levels of the androgen receptor gene (AR), estrogen receptor gene (ER1), and the sex hormone-synthesis-related genes (HMGR, CYP17, and CYP19a) were significantly decreased, while the estrogen receptor gene (ER2a), thyroid hormone receptor gene (TRβ), and aromatic hydrocarbon receptor genes (AhR1a and AhR2) were upregulated in juvenile Gobiocypris rarus under exposure to 2.0 μg·L-1 of fenpropathrin. Relatively low concentrations of fenpropathrin can affect the expression of sex hormone receptor genes, genes related to sex hormone synthesis, thyroid hormone receptor genes, and aromatic hydrocarbon receptor genes, thus interfering with the reproductive system, thyroid system, and metabolic level in Gobiocypris rarus. Therefore, more attention should be paid to the endocrine-disrupting effect caused by the pyrethroid insecticides in the water environment. Furthermore, studies on the internal mechanism of the endocrine-disrupting effect of pyrethroid insecticides on fish is needed in the future.

Effect of zinc intake on association between fluoride exposure and abnormal sex steroid hormones among US pubertal males: NHANES, 2013-2016.

Excessive fluoride exposure can disturb the balance of sex hormones. Zinc is essential for sex hormone synthesis and spermatogenesis. But it is not clear how zinc affects the relationship of fluoride exposure with abnormal sex steroid hormones. Here, a total of 1008 pubertal males from the National Health and Nutrition Examination Survey (NHANES) in two cycles (2013-2014, 2015-2016) were enrolled. The concentrations of water fluoride and plasma fluoride and the levels of serum testosterone, estradiol, and sex hormone binding globulin (SHBG) were measured. Two 24-h dietary recall interviews were conducted to assess the dietary zinc intake. The relationships of fluoride exposure and zinc intake with sex hormones were examined using linear regression and logistic regression models, while the generalized additive model was used to evaluate their non-linear relationship. Our findings revealed that for every two-fold increase in plasma fluoride concentration, testosterone levels decreased by 7.27% (95% CI - 11.49%, - 2.86%) and estradiol levels decreased by 8.73% (95% CI - 13.61%, - 3.57%). There was also significant non-linear association observed between zinc intake and SHBG levels. Being in the first tertile of plasma fluoride had a 60% lower risk of high SHBG (OR = 0.40, 95% CI 0.18, 0.89) compared with being in the second tertile. When compared to the first tertile, being in the second tertile of zinc intake was associated with a 63% (OR = 0.37, 95% CI 0.14, 0.98) lower risk of high SHBG. Furthermore, we observed an interactive effect between the plasma fluoride and zinc intake on estradiol and SHBG, as well as the risk of high SHBG (P-interaction < 0.10). These findings suggest that fluoride exposure and zinc intake can affect sex steroid hormone levels and the risk of high SHBG. Notably, zinc intake may alleviate the increased risk of high SHBG and the abnormal changes of estradiol and SHBG caused by higher fluoride exposure.

The ROS/SIRT1/STAR axis as a target for melatonin ameliorating atrazine-induced mitochondrial dysfunction and steroid disorders in granulosa cells

The granulosa cells (GCs) of birds are essential for the reproduction and maintenance of populations in nature. Atrazine (ATR) is a potent endocrine disruptor that can interfere with reproductive function in females and Diaminochlorotriazine (DACT) is the primary metabolite of ATR in the organism. Melatonin (MT) is an endogenous hormone with antioxidant properties that plays a crucial role in development of animal germ cells. However, how ATR causes mitochondrial dysfunction, abnormal secretion of steroid hormones, and whether MT prevents ATR-induced female reproductive toxicity remains unclear. Thus, the purpose of this study is to investigate the protective effect of MT against ATR-induced female reproduction. In the present study, the GCs of quail were divided into 6 groups, as follows: C (Serum-free medium), MT (10 µM MT), A250 (250 µM ATR), MA250 (10 µM MT+250 µM ATR), D200 (200 µM DACT) and MD200 (10 µM MT+200 µM DACT), and were cultured for 24 h. The results revealed that ATR prevented GCs proliferation and decreased cell differentiation. ATR caused oxidative damage and mitochondrial dysfunction, leading to disruption of steroid synthesis, which posed a severe risk to GC's function. However, MT supplements reversed these changes. Mechanistically, our study exhibited that the ROS/SIRT1/STAR axis as a target for MT to ameliorate ATR-induced mitochondrial dysfunction and steroid disorders in GCs, which provides new insights into the role of MT in ATR-induced reproductive capacity and species conservation in birds.

Effect of Notch Signal Pathway on Steroid Synthesis Enzymes in TM3 Cells.

Studies have indicated that the conservative Notch pathway contributes to steroid hormone synthesis in the ovaries; however, its role in hormone synthesis of the testis remains unclear. We have previously reported Notch 1, 2, and 3 to be expressed in murine Leydig cells and that inhibition of Notch signaling caused G0/G1 arrest in TM3 Leydig cells. In this study, we have further explored the effect of different Notch signal pathways on key steroidogenic enzymes in murine Leydig cells. TM3 cells were treated with Notch signaling pathway inhibitor MK-0752, and different Notch receptors were also overexpressed in TM3 cells. We evaluated the expression of key enzymes of steroid synthesis, including p450 cholesterol side-chain cleavage enzyme (P450Scc), 3β-hydroxysteroid dehydrogenase (3β-HSD) and steroidogenic acute regulatory protein (StAR), and key transcriptional factors for steroid synthesis, including steroidogenic factor 1 (SF1), GATA-binding protein 4 (GATA4) and GATA6. We found the level of P450Scc, 3β-HSD, StAR and SF1 to be decreased after treatment with MK-0752, while overexpression of Notch1 up-regulated the expression of 3β-HSD, P450Scc, StAR and SF1. MK-0752 and overexpression of different Notch members had no influence on the expression of GATA4 and GATA6. In conclusion, Notch1 signaling may contribute to the steroid synthesis in Leydig cells through regulating SF1 and downstream steroidogenic enzymes (3β-HSD, StAR and P450Scc).

Early Castration in Horses Does Not Impact Osteoarticular Metabolism

The castration of stallions is traditionally performed after puberty, at around the age of 2 years old. No studies have focused on the effects of early castration on osteoarticular metabolism. Thus, we aimed to compare early castration (3 days after birth) with traditional castration (18 months of age) in horses. Testosterone and estradiol levels were monitored from birth to 33 months in both groups. We quantified the levels of biomarkers of cartilage and bone anabolism (CPII and N-MID) and catabolism (CTX-I and CTX-II), as well as of osteoarthritis (HA and COMP) and inflammation (IL-6 and PGE2). We observed a lack of parallelism between testosterone and estradiol synthesis after birth and during puberty in both groups. The extra-gonadal synthesis of steroids was observed around the 28-month mark, regardless of the castration age. We found the expression of estrogen receptor (ESR1) in cartilage and bone, whereas androgen receptor (AR) expression appeared to be restricted to bone. Nevertheless, with respect to osteoarticular metabolism, steroid hormone deprivation resulting from early castration had no discernable impact on the levels of biomarkers related to bone and cartilage metabolism, nor on those associated with OA and inflammation. Consequently, our research demonstrated that early castration does not disrupt bone and cartilage homeostasis.

Mitochondrial morphology in the mouse adrenal cortex: Influence of chronic psychosocial stress

Mitochondria within the adrenal cortex play a key role in synthesizing steroid hormones. The adrenal cortex is organized in three functionally specialized zones (glomerulosa, fasciculata, and reticularis) that produce different classes of steroid hormones in response to various stimuli, including psychosocial stress. Given that the functions and morphology of mitochondria are dynamically related and respond to stress, we applied transmission electron microscopy (TEM) to examine potential differences in mitochondrial morphology under basal and chronic psychosocial stress conditions. We used the chronic subordinate colony housing (CSC) paradigm, a murine model of chronic psychosocial stress. Our findings quantitatively define how mitochondrial morphology differs among each of the three adrenal cortex zones under basal conditions, and show that chronic psychosocial stress mainly affected mitochondria in the zona glomerulosa, shifting their morphology towards the more typical glucocorticoid-producing zona fasciculata mitochondrial phenotype. Analysis of adrenocortical lipid droplets that provide cholesterol for steroidogenesis showed that chronic psychosocial stress altered lipid droplet diameter, without affecting droplet number or inter-organellar mitochondria-lipid droplet interactions. Together, our findings support the hypothesis that each adrenal cortex layer is characterized by morphologically distinct mitochondria and that this adrenal zone-specific mitochondrial morphology is sensitive to environmental stimuli, including chronic psychosocial stressors. Further research is needed to define the role of these stress-induced changes in mitochondrial morphology, particularly in the zona glomerulosa, on stress resilience and related behaviors.

The multistep oxidation of cholesterol to pregnenolone by human cytochrome P450 11A1 is highly processive

Cytochrome P450 (P450, CYP) 11A1 is the classical cholesterol side chain cleavage enzyme (P450scc) that removes six carbons of the side chain, the first and rate-limiting step in the synthesis of all mammalian steroids. The reaction is a 3-step, 6-electron oxidation that proceeds via formation of 22R-hydroxy (OH) and 20R,22R-(OH)2 cholesterol, yielding pregnenolone. We expressed human P450 11A1 in bacteria, purified the enzyme in the absence of nonionic detergents, and assayed pregnenolone formation by HPLC-mass spectrometry of the dansyl hydrazone. The reaction was inhibited by the nonionic detergent Tween 20, and several lipids did not enhance enzymatic activity. The 22R-OH and 20R,22R-(OH)2 cholesterol intermediates were bound to P450 11A1 relatively tightly, as judged by steady-state optical titrations and koff rates. The electron donor adrenodoxin had little effect on binding; the substrate cholesterol showed a ∼5-fold stimulatory effect on the binding of adrenodoxin to P450 11A1. Presteady-state single-turnover kinetic analysis was consistent with a highly processive reaction with rates of intermediate oxidation steps far exceeding dissociation rates for products and substrates. The presteady-state kinetic analysis revealed a second di-OH cholesterol product, separable by HPLC, in addition to 20R,22R-(OH)2 cholesterol, which we characterized as a rotamer that was also converted to pregnenolone at a similar rate. The first oxidation step (at C-22) is the slowest, limiting the overall rate of cleavage. d3-Cholesterol showed no kinetic deuterium isotope effect on C-22, indicating that C-H bond cleavage is not rate-limiting in the first hydroxylation step.

New insights into endocrine reproductive toxicity of Microcystis aeruginosa combined with ammonia exposure in zebrafish

The ecological risk posed by MCs-producing M. aeruginosa and elevated ammonia to fish in actual aquatic environments remains uncertain. To address this knowledge gap, we conducted simulations to investigate the endocrine-reproductive toxicity of prolonged exposure (45 d) to Microcystis aeruginosa (2 × 10^6 cells/mL) and 30 mg/L total ammonia nitrogen (TAN) in zebrafish under environmentally relevant conditions. Our results showed that exposure to M. aeruginosa significantly inhibited the body weight, increased gonadosomatic index (GSI), delayed oocyte development, and disrupted endocrine hormonal balance (reduced gonadotropin-releasing hormone (GnRH), and increased estradiol (E2) and testosterone (T)). Mechanistically, it should be attributed to the over-expression of hypothalamic-pituitary-gonadal-liver (HPGL) axis-related genes (cyp11a and cyp17) induced by M. aeruginosa. On the other hand, TAN exposure caused mild damage to zebrafish ovarian tissue and promoted an increase of T levels by inducing the upregulation of steroid hormone synthesis gene (3βhsd) expression in the ovary. It is worth noting that the dysregulation of E2/T ratio in zebrafish ovaries may be attributed to the inhibition of cyp19a1a by both M. aeruginosa and TAN. These results were further confirmed by changes in steroidogenic enzymes activities in the M. aeruginosa or TAN treated groups. Our findings indicated that exposure to M. aeruginosa and TAN had adverse impacts on the reproductive system of zebrafish. And the combined exposure of M. aeruginosa and TAN had more severe effects on the body weight, GSI, pathological changes, hormone levels and HPGL-axis related gene expression in female zebrafish. These results provide compelling evidence regarding the potential risks for reproductive health associated with M. aeruginosa and TAN in eutrophic water bodies experiencing M. aeruginosa blooms, and contribute to the development of effective strategies for monitoring and managing these toxins in aquatic ecosystems.

Effects of environmentally relevant concentrations of niclosamide on lipid metabolism and steroid hormone synthesis in adult female zebrafish

Niclosamide (NIC) is a commonly used molluscicide that reportedly disrupts the endocrine system and may lead to lipid metabolism disorders. However, few studies have investigated the mechanism by which NIC affects the endocrine system from the perspective of lipid metabolism. Adult female zebrafish were fed either a normal-fat diet (NFD) or a high-fat diet (HFD) and then exposed for 28 days to environmentally relevant concentrations of NIC. NIC accumulated most in the liver followed by the brain and then the gonads in both feeding conditions. Somatic index changes confirmed that HFD promotes fish growth, and NIC administration inhibits it. Lipid metabolites were decreased by NIC, as were levels of pregnenolone, androstenedione, estrogen, testosterone, and estradiol, suggesting that NIC impacted steroidogenesis. In addition, gene transcription changes related to the hypothalamic-pituitary-gonad-liver (HPGL) axis and altered ovarian histology strongly suggest that environmental relevant concentrations of NIC exposure may disrupt endocrine function. These findings highlighted that NIC exposure at environmentally relevant concentrations elicited endocrine-disruption effects may through impairing of steroid hormone synthesis.

Transcriptome analysis of three medicinal plants of the genus Polygonatum: identification of genes involved in polysaccharide and steroidal saponins biosynthesis.

Polysaccharides and saponins are the main active components of Polygonati Rhizoma. Studying the molecular mechanism of their synthesis pathway is helpful in improving the content of active components at the molecular level. At present, transcriptome analysis of three Polygonatum species (Polygonatum sibiricum Red., Polygonatum cyrtonema Hua, Polygonatum kingianum Coll. et Hemsl.) has been reported, but no comparative study has been found on the transcriptome data of the three species. Transcriptome sequencing was performed on the rhizomes of three Polygonatum species based on high-throughput sequencing technology, and all transcripts were assembled. A total of 168,108 unigenes were generated after the removal of redundancy, of which 121,642 were annotated in seven databases. Through differential analysis and expression analysis of key enzyme genes in the synthesis pathway of three Polygonatum polysaccharides and steroidal saponins, 135 differentially expressed genes encoding 18 enzymes and 128 differentially expressed genes encoding 28 enzymes were identified, respectively. Numerous transcription factors are involved in the carbohydrate synthesis pathway. Quantitative real-time PCR was used to further verify the gene expression level. In this paper, we present a public transcriptome dataset of three medicinal plants of the genus Polygonatum, and analyze the key enzyme genes of polysaccharide and steroidal saponins synthesis pathway, which lays a foundation for improving the active component content of Polygonati Rhizoma by molecular means.

Adipolin (C1QTNF12) is a new adipokine in female reproduction: expression and function in porcine granulosa cells.

Adipolin (C1QTNF12) has been described as a regulator of metabolism and is linked with the pathophysiology of PCOS. In this study, for the first time, we show the expression of C1QTNF12 in granulosa cells and its positive effect on porcine granulosa cell proliferation and steroid synthesis. Adipolin (C1QTNF12) is a recently discovered adipokine that plays an important role in glucose and insulin level regulation. Previous studies showed its reduced level in serum of women suffering from polycystic ovarian syndrome; however, whether C1QTNF12 regulates ovary function is still unknown. The aim of the study was first to determine the level of C1QTNF12 in the porcine ovarian follicles granulosa cells (Gc) and then its in vitro effect on proliferation and steroidogenesis as well as phosphorylation of several signalling pathways. Our results showed that the expression of C1QTNF12 was dependent on follicle size and was higher at the mRNA and protein level in Gc of small than large follicles from both prepubertal and mature animals. Similar pattern was observed for C1QTNF12 concentration in porcine follicular fluid. Additionally, we observed immunolocalisation of C1QTNF12 in Gc, theca cells and oocytes. We found that C1QTNF12 stimulated porcine Gc proliferation via the activation of protein kinase B (AKT). Moreover, C1QTNF12 enhanced progesterone, testosterone and oestradiol secretion by elevating STAR, CYP11A1, HSD3B and CYP19A1 mRNA expression and by activation of MAP3/1 pathway. Additionally, C1QTNF12 increased pMAP3/1-to-MAP3/1 protein expression ratio and enhanced IGF1-induced pTyr-IGF1Rβ-to-IGFR1β and pMAP3/1-to-MAP3/1 protein ratios. Taken together, C1QTNF12 could act directly on proliferation and steroid synthesis and serve as an important factor in in vivo ovarian follicle function, possibly regulating the course of folliculogenesis.

Sex steroid hormones and steroid secretory cells in the gonads of cyclostomes and fish

. The purpose of this work is to generalize data on the content of sex steroid hormones in cyclostomes and fish in connection with the different state of the gonads, as well as on the localization and ultrastructural organization of steroid secretory cells (SCs). Method used: generalization of literature data. Novelty: for the first time, data on the development of SC in the gonads of juvenile fish are presented on the scale of a literature review; changes in their localization and functional activity are shown. Results: Most of the data on the steroidogenic function of the gonads in fish were obtained in connection with the study of sex differentiation (inversion) and sexual maturation (spawning). The issues of the synthesis of sex steroid hormones and their molecular structure, blood levels in various states of the gonads, as well as the localization and ultrastructural organization of SCs are considered. It has been noted that SCs appear in the gonads long before sex differentiation and can be present among stromal, granulosa, and theca cells. In cyclostomes and fish — juvenile protogynous hermaphrodites in the gonads of genetic males, SCs do not appear in the membranes around the oocytes of the previtellogenesis period, which determines their deficiency of estrogen hormones and, as a result, sex inversion. With induced sex reversal, the development of the steroidogenic function of the gonads proceeds in the opposite direction to its natural development: in males, oocytes appear and steroid activity shifts from the stroma of the gonads into granulosa and theca; in females, on the contrary, secretory activity in the follicular membranes decreases and increases in the stroma of the gonads. Practical significance: the results of the analysis will give insight into the nature of sex steroid hormones, their dynamics in the blood, and their role in the implementation of various processes of gonadogenesis and gametogenesis before their practical use in fish farms.

Thyroid-gonadal hormonal interplay in zebrafish exposed to sodium perchlorate: Implications for reproductive health

Perchlorate, a widespread environmental contaminant originating from various industrial applications, agricultural practices, and natural sources, poses potential risks to ecosystems and human health. While previous studies have highlighted its influence on the thyroid endocrine system and its impact on gonadal maturation, reproduction, and sex hormone synthesis, the specific interplay between thyroid and steroid hormones, in this context, remains largely unexplored. Therefore, this study was undertaken to investigate the adverse effects and underlying mechanisms triggered by exposure to sodium perchlorate (SP) on reproductive endocrine activity in zebrafish. For 21 d, the fish were exposed to test SP concentrations (0, 3, 30, 300 mg/L), which were determined based on the exposure concentrations that induced various toxic effects in the fish, considering naturally occurring concentrations. Exposure to SP, except at 3 mg/L in males, significantly decreased the production of thyroid hormone (TH) in both female and male zebrafish. Moreover, gonadal steroid levels were markedly reduced in both sexes. The expression of hepatic vitellogenin (VTG) mRNA in female zebrafish was significantly decreased, whereas aromatase activity in male zebrafish was significantly elevated in the SP exposure groups. The reduced levels of THs and gonadal steroid hormones were strongly correlated. Abnormal responses to SP exposure led to reduced reproductive success in the 300 mg/L SP exposure group. These findings indicate that prolonged and continuous exposure to a specific concentration of SP may lead to long-term reproductive problems in zebrafish, primarily through hormonal imbalances and suppression of hepatic VTG mRNA expression.

Activation or inhibition of NF-κB from the juvenile stage results in skewed sex ratios in common carp (Cyprinus carpio L.)

Due to sexual dimorphism in growth traits, all-female populations and populations with a high proportion of females hold significant economic value in common carp (Cyprinus carpio L.). However, the underlying mechanisms governing gonadal differentiation remain inadequately understood. NF-κB, as a crucial regulator of anti-apoptotic processes, plays a significant role in mammalian gonad formation and zebrafish oocyte maintenance. To investigate the potential roles of the NF-κB pathway in the gonad differentiation of common carp, juveniles with undifferentiated gonads were treated with an activator (DOC) or inhibitor (NAI) of NF-κB. Our findings revealed that juveniles exposed to DOC exhibited a significantly increased female-to-male ratio (from 0.97:1 to 4.30:1) and substantial increases in weight and body length. Conversely, inhibiting NF-κB with NAI resulted in a decreased female-to-male ratio (from 1.06:1 to 0.33:1). Furthermore, NAI also induced the occurrence of ovotestis. Additional in vivo and ex vivo experiments demonstrated that DOC activation of the NF-κB pathways led to inhibition of apoptosis, significant increases in serum 17β-estradiol (E2) levels and the E2/T ratio, and upregulation of the female-biased genes foxl2 and cyp19a1a. Simultaneously, it significantly suppressed serum 11-ketotestosterone (11-KT) levels and the expression of male-biased genes amh, dmrt1, and gsdf and induced VTG levels in the hepatopancreas of male carp. However, NAI exhibited contrasting effects. Both DOC and NAI disrupted the balance of the hypothalamus-pituitary-gonad (HPG) reproductive axis at multiple potential loci, including changes in the expression levels of steroid hormone synthesis-related genes in gonads. Transcriptomic analysis revealed comprehensive molecular changes in undifferentiated gonads during DOC-induced feminization and NAI-induced masculinization. The transcriptome results were consistent with the observed alterations in hormone content and the expression of genes involved in sex hormone synthesis, inflammation, and apoptosis following exposure to DOC or NAI. These findings suggest the involvement of NF-κB in the sex differentiation of common carp by regulating apoptosis and disrupting the HPG reproductive axis. Our study also explores a DOC-induced method of producing high ratios of females and discovers the underlying mechanism of NF-κB's contribution to sex differentiation in common carp.

Unilateral adrenalectomy for a drug-resistant bilateral primary aldosteronism with heart failure: pathophysiology and surgical indication

BackgroundPatients with bilateral primary aldosteronism (PA) generally are treated with antihypertensive drugs, but optimal treatment for patients with complications due to refractory hypertension has not been established. In this report, we present a case with bilateral PA who presented with persistent hypertension, despite treatment with 6 drugs, and left-dominant heart failure, which was improved after unilateral adrenalectomy.Case presentationA 61-year-old man was admitted to our hospital because of severe left-dominant heart failure. His heart rhythm was atrial fibrillation and the left ventricle was diffusely hypertrophic and hypokinetic. Coronary arteries were normal on coronary arteriogram. Primary aldosteronism was suspected based on severe hypokalemia (2.5 mEq/L) and plasma aldosterone concentration (PAC; 1,410 pg/mL). Although computed tomography (CT) showed a single left cortical nodule, adrenal vein sampling (AVS) indicated bilateral PA. Early in the case, heart failure and hyperkalemia in this patient were improved by treatment with a combination of 6 antihypertensive drugs (spironolactone 25 mg/day, eplerenone 100 mg/day, azosemide 60 mg/day, tolvaptan 7.5 mg/day, enalapril 5 mg/day, and bisoprolol fumarate 10 mg/day); however, heart failure relapsed after four months of treatment. We hypothesized that hypertension caused by excess aldosterone was inducing the patient’s heart failure. In order to reduce aldosterone secretory tissue, a laparoscopic adrenalectomy was performed for the left adrenal gland, given the higher level of aldosterone from the left gland compared to the right. Following surgery, the patient’s heart failure was successfully controlled despite the persistence of high PAC. Treatment with anti-hypertensive medications was reduced to two drugs (eplerenone 100 mg/day and bisoprolol fumarate 10 mg/day). In order to elucidate the mechanism of drug resistance, immunohistochemistry (IHC) and real time-polymerase chain reaction (RT-PCR) assays were performed to assess the expression of steroidogenic factor 1 (SF-1), a regulator of steroid synthesis in adrenal tissue. IHC and RT-PCR demonstrated that the expression of SF-1 in this patient (at both the protein and mRNA levels) was higher than that observed in unilateral PA cases that showed good responsivity to drug treatment.ConclusionsUnilateral adrenalectomy to reduce aldosterone secretory tissue may be useful for patients with drug-refractory, bilateral PA. Elevated expression of SF-1 may be involved in drug resistance in PA.