Steroidogenic Acute Regulatory Protein Research Articles

Selenium nanoparticles improve nickel-induced testosterone synthesis disturbance by down-regulating miR-708-5p/p38 MAPK pathway in Leydig cells.

The present study was designed to investigate the role of miR-708-5p/p38 mitogen-activated protein kinase (MAPK) pathway during the mechanism of selenium nanoparticles (Nano-Se) against nickel (Ni)-induced testosterone synthesis disorder in rat Leydig cells. We conducted all procedures based on in vitro culture of rat primary Leydig cells. After treating Leydig cells with Nano-Se and NiSO4 alone or in combination for 24 h, we determined the cell viability, reactive oxygen species (ROS) levels, testosterone production, and the protein expression of key enzymes involved in testosterone biosynthesis: steroidogenic acute regulatory (StAR) and cytochrome P450 cholesterol side chain cleavage enzyme (CYP11A1). The results indicated that Nano-Se antagonized cytotoxicity and eliminated ROS generation induced by NiSO4 , suppressed p38 MAPK protein phosphorylation and reduced miR-708-5p expression. Importantly, we found that Nano-Se upregulated the expression of testosterone synthase and increased testosterone production in Leydig cells. Furthermore, we investigated the effects of p38 MAPK and miR-708-5p using their specific inhibitor during Nano-Se against Ni-induced testosterone synthesis disorder. The results showed that Ni-inhibited testosterone secretion was alleviated by Nano-Se co-treatment with p38 MAPK specific inhibitor SB203580 and miR-708-5p inhibitor, respectively. In conclusion, these findings suggested Nano-Se could inhibit miR-708-5p/p38 MAPK pathway, and up-regulate the key enzymes protein expression for testosterone synthesis, thereby antagonizing Ni-induced disorder of testosterone synthesis in Leydig cells.

Effects of chronic dichlorodiphenyldichloroethylene exposure on testosterone secretion and steroidogenic pathway in the male gonad.

Endocrine disrupting chemicals are present in the environment and/or in consumer products. These agents have the capacity to mimic and/or antagonize endogenous hormones and thus perturb the endocrine axis. The male reproductive tract expresses steroid hormone (androgen and estrogen) receptors at high levels and is a major target for endocrine disrupting chemicals. In this study, Long-Evans male rats were exposed to dichlorodiphenyldichloroethylene, a metabolite of dichlorodiphenyltrichloroethane and a chemical present in the environment, in drinking water at 0.1 and 10μg/L for 4 weeks. At the end of exposure, we measured steroid hormone secretion and analyzed steroidogenic proteins, including 17β-hydroxysteroid dehydrogenase, 3β-hydroxysteroid dehydrogenase, steroidogenic acute regulatory protein, aromatase, and the LH receptor. We also analyzed Leydig cell apoptosis (poly-(ADP-ribose) polymerase) and caspase-3 in the testes. Testicular testosterone (T) and 17β-estradiol (E2) were both affected by exposure to dichlorodiphenyldichloroethylene by displaying altered steroidogenic enzyme expression. Dichlorodiphenyldichloroethylene exposure also increased the expression of enzymes mediating the pathway for programmed cell death, including caspase 3, pro-caspase 3, PARP, and cleaved PARP. Altogether, the present results demonstrate that dichlorodiphenyldichloroethylene directly and/or indirectly can target specific proteins involved in steroid hormone production in the male gonad and suggest that exposure to environmentally relevant dichlorodiphenyldichloroethylene levels has implications for male reproductive development and function.

Hypothalamus-pituitary-interrenal (HPI) axis signaling in Atlantic sturgeon (Acipenser oxyrinchus) and sterlet (Acipenser ruthenus)

In vertebrates, the hypothalamic–pituitaryadrenal/interrenal (HPA/HPI) axis is a highly conserved endocrine axis that regulates glucocorticoid production via signaling by corticotropin releasing hormone (CRH) and adrenocorticotropic hormone (ACTH). Once activated by ACTH, Gs protein-coupled melanocortin 2 receptors (Mc2r) present in corticosteroidogenic cells stimulate expression of steroidogenic acute regulatory protein (Star), which initiates steroid biosynthesis. In the present study, we examined the tissue distribution of genes involved in HPI axis signaling and steroidogenesis in the Atlantic sturgeon (Acipenser oxyrinchus) and provided the first functional characterization of Mc2r in sturgeon. Mc2r of A. oxyrinchus and the sterlet sturgeon (Acipenser ruthenus) are co-dependent on interaction with the melanocortin receptor accessory protein 1 (Mrap1) and highly selective for human (h) ACTH over other melanocortin ligands. A. oxyrinchus expresses key genes involved in HPI axis signaling in a tissue-specific manner that is indicative of the presence of a complete HPI axis in sturgeon. Importantly, we co-localized mc2r, mrap1, and star mRNA expression to the head kidney, indicating that this is possibly a site of ACTH-mediated corticosteroidogenesis in sturgeon. Our results are discussed in the context of other studies on the HPI axis of basal bony vertebrates, which, when taken together, demonstrate a need to better resolve the evolution of HPI axis signaling in vertebrates.

New molecular insights into the A218V variant impact on the steroidogenic acute regulatory protein (STAR) associated with 46, XY disorders of sexual development

Disorders of sexual development (DSD) are an abnormal congenital conditions associated with atypical development of the urogenital tract and external genital structures. The steroidogenic acute regulatory (STAR) gene, associated with congenital lipoid adrenal hyperplasia (CLAH), is included in the targeted gene panel for the DSD diagnosis. Therefore, the genetic alterations of the STAR gene and their molecular effect were examined in the CLAH patients affected with DSD. Ten different Iranian families including twelve male pseudo-hermaphroditism patients with CLAH phenotype were studied using genetic linkage screening and STAR gene sequencing in the linked families to the STAR locus. Furthermore, the structural, dynamical, and functional impacts of the variants on the STAR in silico were analyzed. Sanger sequencing showed the pathogenic variant p.A218V in STAR gene, as the first report in Iranian population. Moreover, modeling and simulation analysis were performed using tools such as radius of gyration, root mean square deviation (RMSD), root mean square fluctuation (RMSF), and molecular docking showed that p.A218V variant affects the residues interaction in cholesterol-binding site and the proper folding of STAR through increasing H-bound and the amount of α-Helix, deceasing total flexibility and changing fluctuations in some residues, resulting in reduced steroidogenic activity of the STAR protein. The study characterized the structural and functional changes of STAR caused by pathogenic variant p.A218V. It leads to limited cholesterol-binding activity of STAR, ultimately leading to the CLAH disease. Molecular dynamics simulation of STAR variants could help explain different clinical manifestations of CLAH disease.

Cuscuta chinensis flavonoids reducing oxidative stress of the improve sperm damage in bisphenol A exposed mice offspring

Bisphenol A (BPA) is a common environmental endocrine disruptor, and overexposure is a threat to male reproduction. Although studies have confirmed that BPA exposure causes a decrease in sperm quality in offspring, the dosage used, and the underlying mechanism is not clear. The purpose of this study is to investigate whether Cuscuta chinensis flavonoids (CCFs) can antagonize or alleviate BPA-induced reproductive injury by analyzing the processes associated with BPA's impairment of sperm quality. BPA and 40 mg/kg bw/day of CCFs were administered to the dams at gestation day (GD) 0.5–17.5. Testicles and serum of male mice are collected on postnatal day 56 (PND56), and spermatozoa are collected to detect relevant indicators. Our results showed that compared with the BPA group, CCFs could significantly increase the serum contents of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T) in males at PND 56, as well as the transcription levels of estrogen receptor alpha (ERα), steroidogenic acute regulatory protein (StAR) and Cytochrome P450 family 11, subfamily A, and member 1 (CYP11A1). CCFs also significantly inhibit the production of reactive oxygen species (ROS), reduce oxidative stress, increase mitochondrial membrane potential, and reduce sperm apoptosis. It also has a certain regulatory effect on sperm telomere length and mitochondrial DNA copy number. These results suggest that CCFs can increase reproductive hormone and receptor levels in adult males by regulating the expression of oxidative stress correlated factors, and ultimately mitigate the negative effects of BPA on sperm quality in male mice.

The co-expression of steroidogenic enzymes with T1R3 during testicular development in the Congjiang Xiang pig

Testosterone is a key crucial hormone synthesized by steroidogenic enzymes that initiate and maintain spermatogenesis and secondary sexual characteristics in adult males. The taste receptor family 1 subunit 3 (T1R3) is reported to be associated with male reproduction. T1R3 can regulate the expressions of steroidogenic enzymes and affect testosterone synthesis. In this study, we addressed the question of whether the expression of steroid synthase was associated with T1R3 and its downstream-tasting molecules during testicular development. The results showed an overall upward trend in testosterone and morphological development in testes from Congjiang Xiang pigs from pre-puberty to sexual maturity. Gene expression levels of testicular steroidogenic acute regulatory protein (StAR), 3β-hydroxysteroid dehydrogenase (3β-HSD), cytochrome P450c17 (CYP17A1) and 17β-hydroxysteroid dehydrogenase (17β-HSD) were increased from pre-puberty to sexual maturity. Protein expression changes of CYP17A1 and 3β-HSD were consistent with mRNA. The relative abundance of tasting molecules (TAS1R3, phospholipase Cβ2, PLCβ2) was increased from pre-puberty to puberty (P < 0.05), with no further significant changes in expression from puberty to sexual maturity. Steroidogenic enzymes (3β-HSD and CYP17A1) were strongly detected in Leydig cells from pre-puberty to sexual maturity, while tasting molecules were localized in Leydig cells and spermatogenic cells. Correlation analysis showed that the genes mentioned above (except for PLCβ2) were positively correlated with testosterone levels and morphological characteristics of the testes at different developmental stages of Congjiang Xiang pigs. These results suggest that steroidogenic enzymes regulate testosterone synthesis and testicular development, and that taste receptor T1R3, but not PLCβ2, may associate with this process.

Dietary arachidonic acid supplementation promotes the growth, steroidogenesis and ovarian development in mud crab Scylla paramamosain

A 22-week feeding experiment was executed to assess the impact of dietary ARA supplementation on the growth, tissue proximate composition, hemolymph biochemical, tissue fatty acids profile, ARA metabolite and sex steroid hormone, and the mRNA expression of ovarian development related gene of female mud crab in the pre-devolopmental stage. Crabs were fed with six diets containing different ARA levels (0.04%, 0.25%, 0.46%, 0.89%, 1.25%, and 1.57%, dry matter). The results showed that the increasing dietary ARA levels had significant effects on survival, percent weight gain (PWG), specific growth rate (SGR), and feed efficiency (FE), which showed an increasing trend initially and then showed a decreasing trend. According to the two-slope broken-line model and quadratic curve model for the relationship between PWG and dietary ARA levels, optimal dietary ARA levels were estimated to be 0.46% and 0.70%. The lipid contents in muscle and hepatopancreas decreased significantly with the increasing dietary ARA levels. Similarly, total cholesterol (T-CHO) and triglyceride (TG) concentrations were significantly decreased with the increasing dietary ARA levels. In the hepatopancreas and muscle, ARA contents significantly increased with the increasing dietary ARA levels. In hemolymph and hepatopancreas, progesterone (Prog), pregnenolone (Preg), estradiol (E2) and vitellogen (Vtg) concentrations presented earlier increase and later decrease trend. In the hepatopancreas, prostaglandin E2 (PGE2) and 5-hydroxyeicosatetraenoic acid (5-HETE) concentrations showed an upward trend. In the muscle, PGE2 concentration presented earlier increasing and later decreasing trend. In the ovary, the increasing dietary ARA levels produced similar effects on the expressions of genes, including steroidogenic acute regulatory protein (star), 3β-hydroxysteroid dehydrogenase (3βhsd), vitellogenin (vtg) and vitellogenin receptor (vtgr). Thus, moderate dietary ARA levels promoted growth, improved ARA deposition and metabolism, increased sex steroid hormone and Vtg levels, and enhanced ovarian development of female mud crab in the pre-developmental stage.

Polystyrene nanoplastics inhibit StAR expression by activating HIF-1α via ERK1/2 MAPK and AKT pathways in TM3 Leydig cells and testicular tissues of mice.

Microplastics (MPs) and nanoplastics (NPs) are widely found in water, food and air, and have been found in human blood, lung and feces. Several studies in vivo have shown that MPs and NPs decrease testosterone level. However, the molecular mechanism of MPs and NPs leading to testosterone reduction remains unclear. In the present study, mice were treated with 50 μg/kg·day polystyrene (PS)-NPs by tail vein injection once daily for two consecutive days, the mRNA and protein levels of steroidogenic acute regulatory protein (StAR) decreased significantly in testis. TM3 Leydig cells were treated with non-toxic doses of PS-NPs, hypoxia-inducible factor-1α (HIF-1α) mRNA translation was induced by PS-NPs through mTOR/4E-BP1 pathway, which was activated by the ERK1/2 MAPK and AKT pathways. Simultaneously, increased HIF-1α protein inhibited StAR transcription. Additionally, reactive oxygen species production induced by PS-NPs played a central role in the activation of ERK1/2 MAPK/mTOR and AKT/mTOR signaling pathways. These results suggest that PS-NPs down-regulate StAR expression by increasing HIF-1α, which is induced by activation of mTOR/4E-BP1 through the ERK1/2 MAPK and AKT signaling pathways. Our findings provide new insight into the potential molecular mechanism by which PS-NPs impair testosterone synthesis and male reproductive function.

Rhamnazin ameliorates 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin-evoked testicular toxicity by restoring biochemical, spermatogenic and histological profile in male albino rats.

2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) is a potential environmental toxin that has the ability to affect male reproductive tract. Rhamnazin is a naturally present flavone that displays multiple medicinal properties. Therefore, the current study was designed to determine the mitigative role of rhamnazin against TCDD induced reproductive damage. 48 adult male albino rats were randomly separated into four groups: control, TCDD (10µgkg-1), TCDD + rhamnazin (10µgkg-1 + 5mgkg-1 respectively) and rhamnazin (5mgkg-1). The trial was conducted for 56days. TCDD intoxication notably affected superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GSR) and catalase (CAT) activities, besides reactive oxygen species (ROS) and malondialdehyde (MDA) concentrations were augmented. TCDD administration also lowered sperm motility, viability, sperm number, while it augmented the sperm morphological (tail, neck/midpiece and head) anomalies. Moreover, it decreased the levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and plasma testosterone. Moreover, TCDD reduced steroidogenic enzymes i.e., 17-beta hydroxysteroid dehydrogenase (17β-HSD), steroidogenic acute regulatory protein (StAR) and 3-beta hydroxysteroid dehydrogenase (3β-HSD) as well as B-cell lymphoma 2 (Bcl-2) expressions, but increased the expressions of Bcl-2-associated X protein (Bax) and cysteine-aspartic acid protease (Caspase-3). Furthermore, TCDD exposure also induced histopathological anomalies in testicular tissues. However, the supplementation of rhamnazin recovered all the mentioned damages in the testicles. The outcomes revealed that rhamnazin can ameliorate TCDD induced reproductive toxicity due to its anti-oxidant, anti-apoptotic and androgenic nature.

Rhamnetin alleviates polystyrene microplastics-induced testicular damage by restoring biochemical, steroidogenic, hormonal, apoptotic, inflammatory, spermatogenic and histological profile in male albino rats.

The current research was performed to evaluate the ameliorative effects of Rhamnetin (RHM) on polystyrene microplastics (PS-MPs)-instigated testicular dysfunction in male albino rats. 48 albino rats were distributed in four groups, i.e., control, PS-MPs treated, PS-MPs + RHM co-treated and RHM only supplemented group. PS-MPs exposure considerably reduced anti-oxidant enzymes i.e., catalase (CAT), glutathione peroxidase (GSR), superoxide dismutase (SOD) and glutathione reductase (GPx) activities. Whereas, reactive oxygen species (ROS) level along with malondialdehyde (MDA) was considerably escalated in PS-MPs treated rats as well as a potential decline was observed in sperm progressive motility. Additionally, a substantial upsurge was noticed in the count of dead sperms, deformity in the tail, mid-piece and head of sperms in PS-MPs treated rats. PS-MPs exposure also decreased steroidogenic enzymes, 17β-hydroxysteroid dehydrogenase (17β-HSD), steroidogenic acute regulatory protein (StAR) and 3β-hydroxysteroid dehydrogenase (3β-HSD) expressions. Moreover, the levels of inflammatory indices i.e., Interleukin-6 (IL-6), Nuclear factor kappa-B (NF-κB), Interleukin-1β (IL-1β), tumour necrosis factor-α (TNF-α) and cyclooxygenase-2 (COX-2) activity were also increased in PS-MPs administrated group. Besides it increased the expression of apoptotic markers (Bax and caspase-3) expression. Whereas, anti-apoptotic marker i.e., Bcl-2 expression was reduced. Moreover, luteinizing hormone (LH), follicle-stimulating hormone (FSH) as well as plasma testosterone levels were also decreased. PS-MPs exposure also led to a substantial histopathological damage in testicular tissues. However, RHM supplementation potentially reduced the damaging effects of PS-MPs in the reproductive tissues of male albino rats. Thus, the current study revealed, RHM possesses potential to prevent PS-MPs-induced testicular damage due to its anti-oxidant anti-apoptotic, anti-inflammatory as well as androgenic properties.

Mitoguardin2 Is Associated With Hyperandrogenism and Regulates Steroidogenesis in Human Ovarian Granulosa Cells.

Polycystic ovary syndrome (PCOS) is an endocrinopathy characterized by hyperandrogenism, anovulation, and polycystic ovaries, in which hyperandrogenism manifests by excess androgen and other steroid hormone abnormalities. Mitochondrial fusion is essential in steroidogenesis, while the role of mitochondrial fusion in granulosa cells of hyperandrogenic PCOS patients remains unclear. In this study, mRNA expression of mitochondrial fusion genes mitoguardin1, -2 (MIGA 1, -2) was significantly increased in granulosa cells of hyperandrogenic PCOS but not PCOS with normal androgen levels, their mRNA expression positively correlated with testosterone levels. Dihydrotestosterone (DHT) treatment in mice led to high expression of MIGA2 in granulosa cells of ovulating follicles. Testosterone or forskolin/ phorbol 12-myristate 13-acetate treatments increased expression of MIGA2 and the steroidogenic acute regulatory protein (StAR) in KGN cells. MIGA2 interacted with StAR and induced StAR localization on mitochondria. Furthermore, MIGA2 overexpression significantly increased cAMP-activated protein kinase A (PKA) and phosphorylation of AMP-activated protein kinase (pAMPK) at T172 but inhibited StAR protein expression. However, MIGA2 overexpression increased CYP11A1, HSD3B2, and CYP19A1 mRNA expression. As a result, MIGA2 overexpression decreased progesterone but increased estradiol synthesis. Besides the androgen receptor, testosterone or DHT might also regulate MIGA2 and pAMPK (T172) through LH/choriogonadotropin receptor-mediated PKA signaling. Taken together, these findings indicate that testosterone regulates MIGA2 via PKA/AMP-activated protein kinase signaling in ovarian granulosa cells. It is suggested mitochondrial fusion in ovarian granulosa cells is associated with hyperandrogenism and potentially leads to abnormal steroidogenesis in PCOS.

Long-term consumption of fermented pork fat-based diets differing in calorie, fat content, and fatty acid levels mediates oxidative stress, inflammation, redox imbalance, germ cell apoptosis, disruption of steroidogenesis, and testicular dysfunction in Wistar rats.

There is a dearth of experimental evidence available as to whether the consumption of fermented pork fat (FPF) food has any harmful effects on metabolism and reproduction due to its excessive calories, high fat content, and fatty acid methyl ester (FAME) levels. We hypothesized that exposure to a FPF-diet with excessive calories, a high fat content, and high FAME levels alters testicular physiology and metabolism, leading to permanent damage to the testicular system and its function. Thirteen-week-old male rats (n = 20) were assigned to a high-calorie, high-fat diet (FPF-H, fat-60%, 23kJ/g), a moderate-calorie, moderate-fat diet (FPF-M, fat-30%, 17.5kJ/g), a low-calorie and low-fat diet (FPF-L, fat-15%, 14.21kJ/g) compared to the standard diet (Control, fat-11%, 12.56kJ/g) orally for 90days. GC-MS analysis of the three FPF-diets showed high quantities of saturated fatty acids (SFAs) and polyunsaturated fatty acids-ω6 (PUFA-ω6) and low levels of monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids-ω3 (PUFA-ω3) compared to the control diet. Consequently, the levels of serum FAMEs of the FPF-diet fed rats were significantly increased. In addition, a high level of n-6:n-3 PUFA towards PUFA-ω6 was observed in the serum of FPF-diet fed rats due to the high content of linoleic, γ-linolenic, and arachidonic acid. Long-term consumption of FPF-diets disturbed the anthropometrical, nutritional, physiological, and metabolic profiles. Furthermore, administration of FPF-diets generated metabolic syndrome (dyslipidemia, leptinemia, insulin resistance, obesity, hepato-renal disorder and function), increased the cardiovascular risk factors, and triggered serum and testis inflammatory markers (interleukin-1↑, interleukin-6↑, interleukin-10↓, leukotriene B4↑, prostaglandin↑, nitric oxide↑, myeloperoxidase↑, lactate dehydrogenase↑, and tumor necrosis factor-α↑). Activated testis oxidative stress (conjugated dienes↑, lipid hydroperoxides↑, malondialdehyde↑, protein carbonyl↑, and fragmented DNA↑) and depleted antioxidant reserve (catalase↓, superoxide dismutase↓, glutathione S-transferase↓, reduced glutathione↓, glutathione disulfide↑, and GSH:GSSG ratio↓) were observed in FPF-diet fed rats. Disrupted testis histoarchitecture, progressive deterioration of spermatogenesis, poor sperm quality and functional indices, significant alterations in the reproductive hormones (serum and testis testosterone↓, serum estradiol↑, serum luteinizing hormone↓, and follicle-stimulating hormone↑), were noted in rats fed with FPF diets than in the control diet. Severe steroidogenic impairment (steroidogenic acute regulatory protein, StAR↓; 3β-hydroxysteroid dehydrogenase, 3β-HSD↓; and luteinizing hormone receptor, LHR↓), deficiency in germ cells proliferation (proliferating cell nuclear antigen, PCNA↓), and abnormally enhanced testicular germ cell apoptosis (terminal deoxynucleotidyl transferase dUTP nick end labeling, TUNEL assay↑; B-cell lymphoma-2, BCL-2↓; Bcl-2-associated X protein, BAX↑; and BAX/BCL-2 ratio↑) were remarked in the FPF-diet administered rats in comparison with the control diet. In conclusion, the long-term feeding of an FPF-diet with excessive calories, a high fat content, and high FAME levels induced oxidative stress, inflammation, and apoptosis, resulting in metabolic syndrome and hampering male reproductive system and functions. Therefore, the adoption of FPF diets correlates with irreversible changes in testis metabolism, steroidogenesis, germ cell proliferation, and apoptosis, which are related to permanent damage to the testicular system and function later in life.

Mechanistic insight into the protective effects of fisetin against arsenic-induced reproductive toxicity in male rats

Arsenic is one of the most hazardous environmental contaminants, which adversely affects the dynamics of male reproductive system. Fisetin (FIS) is a bioactive flavonoid, which is known to exert strong antioxidative effects. Therefore, the current research was planned to evaluate the alleviative efficacy of FIS against arsenic-induced reproductive damages. Forty-eight male albino rats were divided into 4 groups (n = 12), which were treated as follows: (1) Control, (2) Arsenic-intoxicated group (8 mg kg−1), (3) Arsenic + FIS-treated group (8 mg kg−1 + 10 mg kg−1), and (4) FIS-treated group (10 mgkg−1). After 56 days of treatment, the biochemical, lipidemic, steroidogenic, hormonal, spermatological, apoptotic and histoarchitectural profiles of rats were analyzed. Arsenic intoxication reduced the enzymatic activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GSR), in addition to glutathione (GSH) level. Conversely, the levels of thiobarbituric acid reactive substance (TBARS) and reactive oxygen species (ROS) were increased. Moreover, it escalated the level of low-density lipoprotein (LDL), triglycerides and total cholesterol, while declining the level of high-density lipoprotein (HDL). Furthermore, steroidogenic enzymes expressions, 3β-hydroxysteroid dehydrogenase (HSD), 17β-HSD, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (CYP11A1) and 17α-hydroxylase/17, 20-lyase (CYP17A1), were found to be reduced, which brought down the level of testosterone. Besides, the levels of gonadotropins (LH and FSH) were decreased. Additionally, a decline in sperm mitochondrial membrane potential (MMP), motility, epididymal sperm count and hypo-osmotic swelling (HOS) coil-tailed sperms was observed, whereas the dead sperms and structural damages (head, midpiece and tail) of sperms were escalated. Moreover, arsenic exposure up-regulated the mRNA expressions of apoptotic markers, namely Bax and caspase-3, whereas lowered the expression of anti-apoptotic marker, Bcl-2. In addition, it induced histoarchitectural changes in testes of rats. However, FIS treatment resulted in remarkable improvements in testicular and sperm parameters. Therefore, it was inferred that FIS could serve as a therapeutic candidate against arsenic-generated male reproductive toxicity attributing to its anti-oxidant, anti-lipoperoxidative, anti-apoptotic, and androgenic efficacy.

Effects of IL-11/IL-11 Receptor Alpha on Proliferation and Steroidogenesis in Ovarian Granulosa Cells of Dairy Cows.

Granulosa cells (GCs) are essential for follicular growth, oocyte maturation, and steroidogenesis in the ovaries. Interleukin (IL)-11 is known to play a crucial role in the decidualization of the uterus, however, the expression of the IL-11 system (IL-11, IL-11Rα, and gp130) in the bovine ovary and its exact role in GCs have not been extensively studied. In this study, we identified the IL-11 signaling receptor complex in the bovine ovary and investigated the regulatory effects and underlying mechanism of IL-11Rα on the proliferation and steroidogenesis of GCs. We observed that the IL-11 complex was highly expressed in the GCs of large follicles. IL-11Rα knockdown significantly inhibited GC proliferation by inducing cell cycle arrest at the G1 phase, along with a significant downregulation of proliferating cell nuclear antigen (PCNA) and Cyclin D1 (CCND1) protein, and induced GC apoptosis by significantly upregulating the ratio of BCL-2-associated X protein (BAX) and B-cell lymphoma-2 (BCL-2). In addition, IL-11Rα knockdown attenuated the Janus kinase (JAK) 1-signal transducer and activator of transcription 3 (STAT3) signaling, which is related to cell proliferation and apoptosis. Furthermore, the enzyme-linked immunosorbent assay (ELISA) indicated that IL-11Rα silencing decreased the basal and forskolin (FSK)-stimulated secretions of estradiol and progesterone in GC culture medium concomitantly with a remarkable decrease in cytochrome P450 family 19 subfamily A member 1 (CYP19A1) and steroidogenic acute regulatory protein (StAR). We subsequently determined that this reduction in steroidogenesis was in parallel with the decrease in phosphorylations of protein kinase A (PKA) substrates, cAMP-response element binding protein (CREB), extracellular regulated protein kinase (ERK) 1/2, and p38 mitogen-activated protein kinase (MAPK). Taken together, these data indicate that the effects of IL-11/IL-11Rα on the proliferation and steroidogenesis in bovine GCs is mediated by the JAK1-STAT3, PKA-CREB, p38MAPK, and ERK1/2 signaling pathways. Our findings provide important insights into the local action of the IL-11 system in regulating ovarian function.

Anthocyanins Prevent AAPH-Induced Steroidogenesis Disorder in Leydig Cells by Counteracting Oxidative Stress and StAR Abnormal Expression in a Structure-Dependent Manner

Testosterone deficiency may increase the risk of sexual dysfunction and the failure of spermatogenesis. Oxidative stress that is derived from the destruction of homeostasis, disease, and exposure to contaminants can damage the steroidogenicity process in Leydig cells, resulting in a reduction in testosterone synthesis. Anthocyanins are a group of innoxious antioxidants widely recognized in food sources, and are an ideal candidate to relieve oxidative stress-related steroidogenesis disorder. However, there is still a major gap in our knowledge of the structure-function relationship of anthocyanin on the activity mentioned above. In the present study, four anthocyanins including cyanidin-3-glucoside (Cy-3-glu), delphinidin-3-glucoside (Dp-3-glu), pelargonidin-3-glucoside (Pg-3-glu), and cyanidin-3,5-diglucoside (Cy-3,5-diglu) were applied to reverse testosterone generation after employing 2,2'-Azobis(2-amidinopropane)-dihydrochloride (AAPH) as the inducer of oxidative stress in R2C cells. The results demonstrated that all four kinds of anthocyanins can inhibit ROS generation, alleviate mitochondrial membrane potential damage, and contribute to increased testosterone. Among them, Cy-3,5-diglu with diglycoside performed best on antioxidative ability and improved cell dysfunction and upregulated the expression of the steroidogenic acute regulatory protein (StAR). The molecular docking further revealed the direct combination between anthocyanins and StAR, suggesting that anthocyanins with monosaccharide were more likely to interact with StAR than with diglycoside. Taken together, these data indicate that recipient R2C cells under oxidative stress submitted to anthocyanins exhibited improved steroidogenesis in a structure-dependent manner. Anthocyanins could be considered the ideal ingredients against oxidative stress-induced testosterone deficiency.

Impact of Chronic Multi-Generational Exposure to an Environmentally Relevant Atrazine Concentration on Testicular Development and Function in Mice.

A common herbicide, atrazine, is associated with poor health. Atrazine acts as an endocrine disruptor at supra-environmental levels. Little research, however, has been conducted regarding chronic exposure to environmental atrazine concentrations across generations. This study utilized comprehensive endpoint measures to investigate the effects of chronic exposure to a conservative atrazine concentration (0.02 ng/mL), measured in Australian waterways, on male mice fertility across two generations. Mice were exposed through the maternal line, from the pre-conception period and through the F1 and F2 generations until three or six months of age. Atrazine did not impact sperm function, testicular morphology nor germ cell parameters but did alter the expression of steroidogenic genes in the F1, down-regulating the expression of Cyp17a1 (Cytochrome P450 family 17, subfamily A member 1; p = 0.0008) and Ddx4 (DEAD-box helicase 4; p = 0.007), and up-regulating the expression of Star (Steroidogenic acute regulatory protein; p = 0.017). In the F2, atrazine induced up-regulation in the expression of Star (p = 0.016). The current study demonstrates that chronic exposure to an environmentally relevant atrazine concentration perturbs testicular steroid-associated gene expression that varies across generations. Future studies through the paternal and combined parental lineages should be undertaken to further elucidate the multigenerational effects of atrazine on male fertility.

Long-read sequencing reveals the effect of follicle-stimulating hormone on the mRNA profile of chicken granulosa cells from prehierarchical follicles

Follicle selection is an important step in the laying process of chicken, which is closely related to the laying performance and fecundity of hens. Follicle selection mainly depends on the regulation of follicle-stimulating hormone (FSH) secreted by pituitary gland and the expression of follicle stimulation hormone receptor. To uncover the role of FSH in chicken follicle selection, in this study, we analyzed the changes in the mRNA transcriptome profiles of FSH-treated chicken granulosa cells from prehierarchical follicles by long-read sequencing Oxford Nanopore Technologies (ONT) approach. Among the 10,764 genes detected, 31 differentially expressed (DE) transcripts of 28 DE genes were significantly upregulated by FSH treatment. These DE transcripts (DETs) were mainly related to the steroid biosynthetic process by GO analysis and enriched in pathways of ovarian steroidogenesis and aldosterone synthesis and secretion by KEGG analysis. Among these genes, the mRNA and protein expression of TNF receptor associated factor 7 (TRAF7) was upregulated after FSH treatment. Further study revealed that TRAF7 stimulated the mRNA expression of steroidogenic enzymes steroidogenic acute regulatory protein (StAR) and cytochrome P450 family 11 subfamily A member 1 (CYP11A1) genes and the proliferation of granulosa cells. This is the first study to investigate differences in chicken prehierarchical follicular granulosa cells before and after FSH treatment by using ONT transcriptome sequencing, which provides a reference for a more comprehensive understanding of the molecular mechanism of follicle selection in chicken.

Effect of maternal periconceptional undernutrition in sheep on cortisol regulation in offspring from mid-late gestation, through to adulthood.

Maternal periconceptional undernutrition (PCUN) alters fetal hypothalamic-pituitary-adrenal axis (HPAA) function and placental glucocorticoid metabolism in sheep. The effects of PCUN on HPAA function in adult life are not known. We investigated the effects of PCUN on fetal adrenal development across gestation and on cortisol regulation in adult offspring. Ewes were undernourished from 61 days before to 30 days after conception ('PCUN') or fed ad libitum ('N'). mRNA expression in the fetal adrenal gland of ACTH receptor (ACTHR), steroidogenic acute regulatory protein (STAR), cytochrome P450 17A1 (CYP17A1), 11beta-hydroxysteroid-dehydrogenase type 2 (11βHSD2), insulin-like growth factor-2 (IGF2), and in the fetal hippocampus of 11βHSD1, 11βHSD2, mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) was determined at 50 (adrenal only), 85, 120 and 131 days of gestation (term=148 days). In adult offspring (≥ 3 years, N; 10 female, 5 male, PCUN; 10 female, 10 male) a combined arginine vasopressin (AVP, 0.1 μg/kg) and corticotropin-releasing hormone (CRH, 0.5 μg/kg) challenge and a metyrapone (40 mg/kg) challenge were undertaken. mRNA expression of ACTHR, STAR and CYP17A1 were determined in adult adrenals. Fetal adrenal STAR, CYP17A1 and IGF2 mRNA expression were not different between groups in early gestation but were higher in PCUN than N at 131 days' gestation (all p<0.01). PCUN reduced fetal hippocampal MR and GR mRNA expression by 50% at 85 day, but not in later gestation. Adult offspring plasma cortisol responses to AVP+CRH or metyrapone were not different between groups. Plasma ACTH response to AVP+CRH was lower in PCUN males but ACTH response to metyrapone was not different between groups. Adult adrenal ACTHR, STAR, and CYP17A1 mRNA expression were not affected by PCUN. We conclude that the effects of PCUN on fetal HPAA function that became apparent in late gestation, are not reflected in adrenal cortisol secretion in mid-adulthood.

Effects of dietary theabrownins on production performance, egg quality, and ovarian function of laying hens with different ages

This experiment was conducted to investigate the effect of theabrownins (TB) on production performance, egg quality, and ovarian function of laying hens at different ages. A total of 240 Lohmann laying hens were assigned in a 2×2 factorial design, which encompassed 2 layers ages (47-wk-old and 67-wk-old) and 2 dietary levels of TB (0 and 100 mg/kg) for 12 wk. Results showed that older layers had lower laying rate, egg mass, and higher feed-to-egg ratio (F/E), egg weight and unqualified egg rate than the younger layers (P(AGE) < 0.01) during all the experimental period. The effect of TB was found to increase egg laying rate and feed efficiency during 5 to 8 wk, 9 to 12 wk and the overall phases and decreased unqualified egg rate during 1 to 4 wk and the overall phases (P(TB) ≤ 0.05). The eggshell quality (strength, thickness), albumen quality (albumen height and Haugh unit) of eggs from older layers were decreased during overall phases (P(AGE) ≤ 0.05). TB increased eggshell strength during all phases and enhanced eggshell thickness at the end of wk 4 and 8 and increased albumen height and Haugh unit at the end of wk 8 and 12 of older layers (P(Interaction) ≤ 0.05). In addition, TB also increased egg quality of older layers after 14 d storage. A decrease in the serum concentration of progesterone, melatonin, follicle stimulating hormone, estradiol was observed in the older compared to the younger ones (P(AGE) < 0.05), while the increase in serum concentration of progesterone, melatonin, anti-Müllerian hormone (AMH) were more emphasized when older hens received TB supplemented diet (P(Interaction) < 0.05). The older layer demonstrated lower the concentration of glutathione (GSH) (P(AGE) < 0.05). And the activity of glutathione-s-transferase (GST) was significantly decreased in layers under 67-wk-old (P(AGE) <0.05). The increase in concentration of GSH and the decrease in concentration of malondialdehyde (MDA) were more pronounced when TB were supplemented in 67-wk-old layers (P(Interaction) ≤ 0.05). Layers at 67-wk-old had lower mRNA expression of Heme oxygenase 1 (HO-1) (P(AGE) < 0.01) in ovary. Dietary TB supplementation upregulated mRNA gene expression of HO-1, Nuclear factor E2 related factor 2 (Nrf2), Quinone oxidoreductase 1 (NQO1) (P(TB) < 0.01). Dietary TB upregulated mRNA expression of ovarian reproductive hormone receptor (estrogen receptor 1 [ESR1] and steroidogenic acute regulatory protein 1 [StAR1]]; P(TB) < 0.01). The results suggest feeding TB (100 mg/kg) could improve the egg production rate, egg quality, and antioxidant capacity of the ovary. Moreover, the effect of TB was more pronounced in older layers (64-wk-old vs. 47-wk-old).

Male reproductive toxicity of polystyrene microplastics: Study on the endoplasmic reticulum stress signaling pathway.

Microplastics (MPs) have raised health concerns in public for its potential reproductive toxicity. In this study, we subjected the Kunming mice to 0.01, 0.1 and 1.0mg/day polystyrene MPs (10μm, PS-MPs) for 35 days, aiming to investigate the relevant male reproductive toxicity and latent molecular mechanism. The results showed the decreased sperm counts and motility, while the elevated sperm abnormality in PS-MPs-exposed mice. Testicular H&E staining displayed the vacuolization, atrophy, and even shedding of germ cells in seminiferous tubule. And the testosterone content in serum also decreased with PS-MPs treatment. Moreover, molecular analysis indicated that PS-MPs upregulated the expression trait factors for ERS (e.g., immunoglobulin-binding protein [BIP], inositol-requiring protein 1α [IRE1α], X-box-binding protein 1 splicing [XBP1s], Jun kinase [JNK], and the transcription of CCAAT/enhancer-binding protein (C/EBP) homologous protein [CHOP]) and downstream apoptotic modulator (e.g., Caspase-12, -9, and -3) in the testis. The steroidogenic acute regulatory protein (StAR), the testosterone synthetic initiator, was also downregulated. With the supplementation of ERS inhibitor, the MPs-induced testicular damage and decreased testosterone were improved to almost normal level. Overall, this study suggested that PS-MPs generate reproductive toxicity possibly via activating ERS and apoptosis signaling pathway.