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Background: In recent years, the use of endocrine-disrupting chemicals (EDCs) has become increasingly common, leading to severe environmental pollution and harm to aquatic organisms. 17α-Methyltestosterone (MT) is a synthetic androgen that can cause immunotoxicity in aquaculture, affecting fish health. To address this issue, this study aimed to investigate the effect of Vitamin C (VC) on MT-induced immunotoxicity and determine the optimal VC supplementation. Results: Carassius auratus was exposed to 50 ng/L MT and treated with 25, 50, and 150 mg/kg VC for 7, 14, and 21 d. Morphological indicators, histological characteristics, hepatic antioxidant capacity, and immune-related gene expression were analyzed. Additionally, RNA-seq was performed on the liver tissues of the control, MT, and MT + 25 mg/kg VC groups after 21 d. Results showed that, MT treatment significantly increased liver malondialdehyde content and inhibited immune-related gene expression (TNF-α, IL-8, INF-γ, IL-10, Caspase-9, and IGF-I), causing oxidative stress and immunotoxicity, leading to hepatic steatosis. However, supplementation with 25–50 mg/kg VC effectively alleviated the MT-induced damage to the hepatic structure and immune system. RNA-seq revealed significant enrichment of differentially expressed genes in multiple signaling pathways, including the mTOR, MAPK, and Wnt pathways. Conclusions: In summary, 25–50 mg/kg VC alleviated inhibitory effect of MT on immune-related genes in C. auratus liver, reducing MT-induced tissue damage. VC not only alleviated inflammation, oxidative stress, and immunotoxicity induced by MT through the regulation of the mTOR, MAPK, and Wnt signaling pathways, but also indirectly enhanced cellular antioxidant defense mechanisms by regulating the NRF2 pathway. This provides a theoretical basis for VC application in aquaculture, improving fish health and increasing efficiency. [ABSTRACT FROM AUTHOR]

Abstract Background In recent years, the use of endocrine-disrupting chemicals (EDCs) has become increasingly common, leading to severe environmental pollution and harm to aquatic organisms. 17α-Methyltestosterone (MT) is a synthetic androgen that can cause immunotoxicity in aquaculture, affecting fish health. To address this issue, this study aimed to investigate the effect of Vitamin C (VC) on MT-induced immunotoxicity and determine the optimal VC supplementation. Results Carassius auratus was exposed to 50 ng/L MT and treated with 25, 50, and 150 mg/kg VC for 7, 14, and 21 d. Morphological indicators, histological characteristics, hepatic antioxidant capacity, and immune-related gene expression were analyzed. Additionally, RNA-seq was performed on the liver tissues of the control, MT, and MT + 25 mg/kg VC groups after 21 d. Results showed that, MT treatment significantly increased liver malondialdehyde content and inhibited immune-related gene expression (TNF-α, IL-8, INF-γ, IL-10, Caspase-9, and IGF-I), causing oxidative stress and immunotoxicity, leading to hepatic steatosis. However, supplementation with 25–50 mg/kg VC effectively alleviated the MT-induced damage to the hepatic structure and immune system. RNA-seq revealed significant enrichment of differentially expressed genes in multiple signaling pathways, including the mTOR, MAPK, and Wnt pathways. Conclusions In summary, 25–50 mg/kg VC alleviated inhibitory effect of MT on immune-related genes in C. auratus liver, reducing MT-induced tissue damage. VC not only alleviated inflammation, oxidative stress, and immunotoxicity induced by MT through the regulation of the mTOR, MAPK, and Wnt signaling pathways, but also indirectly enhanced cellular antioxidant defense mechanisms by regulating the NRF2 pathway. This provides a theoretical basis for VC application in aquaculture, improving fish health and increasing efficiency.

The sex determination system of largemouth bass (Micropterus salmoides, LMB) is XX/XY; however, the underlying molecular mechanisms involved in early sex differentiation, gonadal development, and exogenous hormone-induced sex reversal remain unknown. In this study, LMB at 15 days post-hatching (dph) were fed diets containing 20 mg/kg of 17a-methyltestosterone (17a-MT) or 30 mg/kg of 17ß-estradiol (17ß-E2) for 60 days, respectively. Serum steroid levels, histological observations of the gonads, and identification of sex-specific markers were employed to screen the gonads of 60-day-old normal female fish (XX-F), normal male fish (XY-M), 17ß-E2 induced pseudo-female fish (XY-F), and 17a-MT-induced pseudo-male fish (XX-M) for transcriptome sequencing in order to uncover genes and pathway involved in the process of sexual reversal. The results from histology and serum sex steroid hormone analysis showed that both 17a-MT and 17ß-E2 were capable of inducing sex reversal of LMB at 15 dph. Transcriptome results revealed a total of 2,753 genes exhibiting differential expression, and the expression pattern of these genes in the gonads of XX-M or XY-F resembled that of normal females or males. The male sex-biased genes that are upregulated in XX-M and downregulated in XY-F are referred to as key genes for male reversal, while the female sex-biased genes that are upregulated in XY-F and downregulated in XX-M are referred to as key genes for female reversal. Finally, 12 differentially expressed genes (DEGs) related to male sex reversal were screened, including star2, cyp17a, cyp11b1, dmrt1, amh, sox9a, katnal1, spata4, spata6l, spata7, spata18 and foxl3. 2DEGs (foxl2a and cyp19a1b) were found to be associated with female sex reversal. The changes in these genes collectively influence the direction of sex differentiation of LMB. Among them, star2, dmrt1 and cyp19a1b with significantly altered expression levels may play potentially crucial role in the process of gender reversal. The expression patterns of 21 randomly selected genes were verified using qRT-PCR which confirmed the reliability and accuracy of the RNA-seq results. These findings not only enhance our understanding of the molecular basis underlying sex reversal but also provide crucial data support for future breeding research on unisexual LMB. [ABSTRACT FROM AUTHOR]

17α-Methyltestosterone (MT) is a widely used androgen for all-male fish production in aquaculture. However, the molecular mechanism underlying MT-induced masculinization remains unclear. In this study, we aim to identify the key gene responsible for MT-induced masculinization using the Nile tilapia (Oreochromis niloticus) amhy, dmrt1, and gsdf mutants, which exhibit male-to-female sex reversal. Nile tilapia fry from these three mutant lines were treated with 50 μg/g MT from 5 to 30 days after hatching (dah). The results showed that amhy and gsdf mutants, but not dmrt1 mutants, were masculinized by the MT treatment. Gonadal transcriptome analysis revealed that genes involved in steroidogenesis and germ cell development in MT-treated dmrt1 mutants exhibited a similar expression pattern to that of the wild type (WT) XX. In addition, the dmrt1 mutants cannot be masculinized by co-treatment with MT and the aromatase inhibitor fadrozole. The MT treatment completely blocked early steroidogenic enzyme (Star2, Cyp17a2, and Cyp19a1a) expression independent of amhy, gsdf, and dmrt1. A luciferase analysis showed that MT directly suppressed basal and Sf-1-activated cyp19a1a promoter activity through ara and arb in cultured HEK293 cells. Furthermore, MT treatment inhibited germ cell proliferation in amhy and gsdf mutants but not in dmrt1 mutants. Consistently, dmrt1 expression was induced in MT-treated WT XX, -amhy, and -gsdf mutants. Taken together, these results suggest that dmrt1 is indispensable for MT-induced masculinization in Nile tilapia and that MT functions by inhibiting early steroid synthesis and activating dmrt1 to promote testis development. [ABSTRACT FROM AUTHOR]

Novel metabolites of the anabolic androgenic steroid 17α‐methyltestosterone have been detected in HepG2 cell in vitro metabolic model and in human urine. Their detection was accomplished through targeted gas chromatography–(tandem) mass spectrometry analysis that has been based on microscale synthesized standards. The related synthesis and the gas chromatography–(tandem) mass spectrometry characterization of the analytical standards are described. All newly presented metabolites have a fully reduced steroid A‐ring with either an 17,17‐dimethyl‐18‐nor‐Δ13 structure or they have been further oxidized at position 16 of the steroid backbone. Metabolites with 17,17‐dimethyl‐18‐nor‐Δ13 structure may be considered as side products of phase II metabolic sulfation of the 17β‐hydroxy group of methyltestosterone or its reduced tetrahydro‐methyltestosterone metabolites 17α‐methyl‐5β‐androstane‐3α,17β‐diol and 17α‐methyl‐5α‐androstane‐3α,17β‐diol that produce the known epimeric 17β‐methyl‐5β‐androstane‐3α,17α‐diol and 17β‐methyl‐5α‐androstane‐3α,17α‐diol metabolites. The prospective of these new metabolites to increase detection time windows and improve identification was investigated by applying the World Anti‐doping Agency TD2021IDCR criteria. The new metabolites, presented herein, complement the current knowledge on the 17α‐methyltestosterone metabolism and in some cases can be used as additional long‐term markers in the frame of sport doping drug testing. [ABSTRACT FROM AUTHOR]

It is widely known that all-female fish production holds economic value for aquaculture. Sebastes schlegelii, a preeminent economic species, exhibits a sex dimorphism, with females surpassing males in growth. In this regard, achieving all-female black rockfish production could significantly enhance breeding profitability. In this study, we utilized the widely used male sex-regulating hormone, 17α-methyltestosterone (MT) at three different concentrations (20, 40, and 60 ppm), to produce pseudomales of S. schlegelii for subsequent all-female offspring breeding. Long-term MT administration severely inhibits the growth of S. schlegelii, while short term had no significant impact. Histological analysis confirmed sex reversal at all MT concentrations; however, both medium and higher MT concentrations impaired testis development. MT also influenced sex steroid hormone levels in pseudomales, suppressing E2 while increasing T and 11-KT levels. In addition, a transcriptome analysis revealed that MT down-regulated ovarian-related genes (cyp19a1a and foxl2) while up-regulating male-related genes (amh) in pseudomales. Furthermore, MT modulated the TGF-β signaling and steroid hormone biosynthesis pathways, indicating its crucial role in S. schlegelii sex differentiation. Therefore, the current study provides a method for achieving sexual reversal using MT in S. schlegelii and offers an initial insight into the underlying mechanism of sexual reversal in this species. [ABSTRACT FROM AUTHOR]

There is an interest in mono-sex fish production in a variety of aquatic species, particularly to achieve higher growth, limit unwanted reproduction, increase survival, prevent sexual/territorial behavior, avoid size diversity in fish harvest, and reduce environmental risks from escape into the wild. In this project, rainbow trout (Oncorhynchus mykiss) larvae (15 days old) were fed diets containing 75 mg letrozole kg−1, 150 mg letrozole kg−1, 70 mg 17α-methyltestosterone kg−1, 140 mg 17α-methyltestosterone kg−1, 200 mg Tribulus terrestris extract kg−1, 400 mg T. terrestris extract kg−1, 200 mg Asparagus racemosus extract kg−1, 400 mg A. racemosus extract kg−1, 200 mg combination of both extracts in equal proportions kg−1, and 400 mg combination of both extracts in equal proportions kg−1. The control group was fed basal diet for 120 days. The fish were then fed a commercial diet until approximately 11 months of age. At the end of the experiment, the growth, survival rate, and percentage of male, female, and intersex fish were evaluated. The results showed that the groups with 17α-methyltestosterone (70 and 140 mg kg−1 diet), letrozole (150 mg kg−1 diet), and a combination of T. terrestris and A. racemosus extracts (400 mg kg−1 diet) had a higher percentage of induction of masculinization than the other groups (P < 0.05). In general, letrozole and a mixture of T. terrestris and A. racemosus extracts are recommended as alternatives to 17α-methyltestosterone. [ABSTRACT FROM AUTHOR]

17α-methyltestosterone (MT), as a synthetic androgen, has been widely used in medicine and aquaculture industry. However, the related molecular regulating mechanism of MT effects on hypothalamic-pituitary-gonadal axis has not been well addressed. This study first evaluated the effects of MT treatment on pituitary of mandarin fish (Siniperca chuatsi). Transcriptome analysis revealed a lot of differentially expressed genes were found in pituitary after MT treatment in early stage (10 and 20 dat), with a relatively few genes found in late stage (30 and 40 dat). Among them, several reproduction related genes were significantly upregulated after MT treatment, while only the expression of follicle-stimulating hormone beta submit (fshb) was significantly inhibited in all stages. Moreover, in vitro experiment also revealed MT significantly inhibited the expression of fshb and promoted the expression of androgen receptors in pituitary. However, dual-luciferase assay revealed that MT activated the promotor activity of fshb via androgen receptors. Finally, we also demonstrated FSH could directly activate the promotor of cyp19a1a by several ways. In all, we first demonstrated exogenous MT could inhibit the expression of fshb in pituitary and further decrease the production of estrogen in gonad by FSH signaling pathway in mandarin fish.

Sex steroid hormones are closely related to biological reproduction, gamete emission and gonadal development. And Srd5a1 plays an important role in the synthesis of sex steroid hormones. In order to investigate the effect of the sex steroid hormone synthesis-related enzyme gene Srd5a1 on ovarian development in female Hyriopsis cumingii, Srd5a1 was obtained from the gonads of H. cumingii by RACE （Rapid-amplificiantion of cDNA ends）. The expression characteristics of Srd5a1 were investigated by realtime fluorescence quantification （qRT-PCR）, in situ hybridization and in vitro induction of 17β-estradiol （E2） and 17α-methyltestosterone （MT） at different concentrations for 24 days. The results showed that the full-length cDNA of Srd5a1 was 2 224 bp, including ORF 825 bp, 3′-UTR 953 bp, and 5′-UTR 446 bp, encoding 274 amino acids. The expression of Srd5a1 in the gonad was bimodal, with higher expression in the ovary than that in the testis; Srd5a1 had the highest expression level in the ovarian discharge period, which was extremely significantly higher than that in all other periods. In situ hybridization results showed that Srd5a1 detected positive signals in ovarian oocytes and follicles as well as spermatocytes of the testis. The expression of Srd5a1 was changed after 24 days of treatment with different concentrations of E2 and MT. In conclusion, Srd5a1 may play a role in the developmental maturation of the ovary oocyte and gamete emission in the female H. cumingii. This study is important for exploring the gonadal growth and development of H. cumingii, and also provides a theoretical reference value for the realization of monogamous breeding of H. cumingii. [ABSTRACT FROM AUTHOR]

As a synthetic androgen, 17α‐methyltestosterone (MT) is widely used in aquaculture to induce sex reversal and may pose a potential risk to aquatic organisms. This ecological risk has attracted the attention of many scholars, but it is not comprehensive enough. Thus, the adverse effects of MT on zebrafish (Danio rerio) were comprehensively evaluated from gonadal histology, as well as the mRNA expression levels of 47 genes related to hypothalamic‐pituitary‐gonadal (HPG) axis, germ cell differentiation, sex determination, and hypothalamus‐pituitary‐thyroid (HPT) axis. Adult zebrafish with a female/male ratio of 5:7 were exposed to a solvent control (0.001% dimethyl sulfoxide) and three measured concentrations of MT (5, 51 and 583 ng/L) for 50 days. The results showed that MT had no significant histological effects on the ovaries of females, but the frequency of late‐mature oocytes (LMO) showed a downward trend, indicating that MT could induce ovarian suppression to a certain extent. The transcriptional expression of activating transcription factor 4b1 (atf4b1), activating transcription factor 4b2 (atf4b2), calcium/calmodulin‐dependent protein kinase II delta 1 (camk2d1), calcium/calmodulin‐dependent protein kinase II delta 2 (camk2d2) and calcium/calmodulin‐dependent protein kinase II inhibitor 2 (camk2n2) genes in the brain of females increased significantly at all treatment groups of MT, and the mRNA expression of forkhead box L2a (foxl2) and ovarian cytochrome P450 aromatase (cyp19a1a) genes in the ovaries were down‐regulated by 5 and 583 ng/L group, which would translate into inhibition of oocyte development. As compared to females, MT had relatively little effects on the reproductive system of males, and only the transcriptional alterations of synaptonemal complex protein 3 (sycp3) and 17‐alpha‐hydroxylase/17,20‐lyase (cyp17) genes were observed in the testes, not enough to affect testicular histology. In addition, MT at all treatments strongly increased corticotropin‐releasing hormone (crh) transcript in the brain of females, as well as deiodinase 2 (dio2) transcript in the brain of males. The paired box protein 8 (pax8) gene was significantly decreased at 51 or 583 ng/L of MT in both female and male brains. The above results suggest that MT can pose potential adverse effects on the reproductive and thyroid endocrine system of fish. [ABSTRACT FROM AUTHOR]

This study was carried out to examine the effect of combining factors for immersion technique on masculinization of red tilapia (Oreochromis spp.). Three selected doses (1000, 1,500, and 2000 μg/L) of 17α-methyltestosterone (17 MT) immersion, 2 exposure times (3 and 6 h) once or twice a week, and interaction of these factors were tested. A total of 12 treatment combinations under 3 × 2 × 2 factorial experiments in randomized complete block design were conducted to compare with the negative control group (no 17 MT-treated) and positive control group (17 MT-treated feed administration). One hundred red tilapia larvae (14 days post-hatch) were randomly allocated in each treatment and were operated in a 10-L glass jar for 21 days. At the end of the treatment regime, the survival rate was determined. Then, fish were raised in a 150-L glass tank using normal commercial feed for 2 months and sex ratios were determined via the gonad squash technique. None of the immersion treatments tested resulted in a statistically significant (p > 0.05) decrease in survival rate. All 17 MT-treated groups produced progenies with a significantly higher (p

Simple Summary: The steroid 17α-methyltestosterone (MT) inhibits ovarian function and is often used to induce sex reversal artificially in vertebrates. In oriental river prawn, sex reversal through vertebrate sex hormones can be observed. Neo-males (sex-reversed female prawns) were maintained by exogenous androgen, and over-reliance led to slow testis growth, small body size, and low growth rate, but sperm was still produced. In female prawns, MT inhibited ovary development and promoted growth. The steroid 17α-methyltestosterone (MT) inhibits ovarian function and is often used to induce sex reversal artificially in vertebrates. In the present study, different concentrations of MT were added as dietary supplementation, and the effects on sex ratio, growth, and gonadal development were examined. After 40 days, the sex ratio (male:female) in each group increased at different degrees with 50 (1.36:1), 100 (1.57:1), and 200 (2.61:1) mg/kg MT, and neo-males with testis–ovary coexistence were observed in the 200 mg/kg MT group. Furthermore, 50 and 100 mg/kg MT could induce female reversion in neo-males. Histologically, the development of the testes in experimental groups was slower, but the ovaries of the experimental and control groups had similar developmental rates. The expression levels of DMRT11E, Foxl2, and SoxE1 in males at 200 mg/kg MT were 8.65-, 3.75-, and 3.45-fold greater than those of the control group. In crustaceans, sex reversal through vertebrate sex hormones can be observed. Neo-males (sex-reversed female prawns) were maintained by exogenous androgen, and over-reliance led to slow testis growth, small body size, and low growth rate, but sperm was still produced. In female prawns, MT inhibited ovary development and promoted growth. [ABSTRACT FROM AUTHOR]

17 α -Methyltestosterone (MT) is a synthetic androgen combining both androgenic and estrogenic effects. In this study, the biotransformation of MT was performed using a cultivated human intestinal fungal strain (Aspergillus niger RG13B1) in vitro. Twelve metabolites, including three new metabolites (1 − 3) and nine known analogs (4 −1 2), were obtained. Their structures were determined by extensive spectroscopic data, including 1D NMR, 2D NMR, and HRESIMS. Based on the structures of the MT metabolites, the main biotransformation reactions were revealed to be hydroxylation, oxidation, dehydrogenation, and reduction. In addition, the cytotoxic effects of MT and metabolites 1 − 12 were evaluated against human cancer cell lines, including glioblastoma cells U87 and LN229, cervical cells HeLa, colorectal cancer cells HCT-15, renal carcinoma cancer cells RCC-786-O, and breast cancer cells BT549. As a result, MT displayed cytotoxity against U87, LN229, and HeLa, among which a strongest inhibition rate of 82.4% against the HeLa cell line at 50 μM was observed, and metabolites 1 − 12 displayed no significant cytotoxicity at 50 μM. Therefore, MT could be metabolized by intestinal fungi and yield various metabolites displaying weak cytotoxicity. [Display omitted] • The biotransformation of 17 α -methyltestosterone was performed by Aspergillus niger RG13B1. • Twelve metabolites were obtained from the biotransformation of 17 α -methyltestosterone. • Three new structures were determined for the metabolites of 17 α -methyltestosterone. • The cytotoxic effect of MT and metabolites 1 − 12 was evaluated against human cancer cells. [ABSTRACT FROM AUTHOR]

17α-Methyltestosterone (17MT), a synthetic organic compound commonly found in sewage waters, can affect reproduction in aquatic animals, such as tilapia and yellow catfish. In the present study, male Gobiocypris rarus were exposed to 25, 50, and 100 ng/L of 17α-methyltestosterone (17MT) for 7 days. We first analyzed miRNA- and RNA-seq results to determine miRNA-target gene pairs and then developed miRNA-mRNA interactive networks after 17MT administration. Total weights, total lengths, and body lengths were not significantly different between the test groups and control groups. The paraffin slice method was applied to testes of G. rarus in the MT exposure and control groups. We found that there were more mature sperm (S) and fewer secondary spermatocytes (SSs) and spermatogonia (SGs) in the testes of control groups. As 17MT concentration increased, fewer and fewer mature sperm (S) were observed in the testes of male G. rarus. The results showed that FSH, 11-KT, and E2 were significantly higher in individuals exposed to 25 ng/L 17MT compared with the control groups. VTG, FSH, LH, 11-KT, and E2 were significantly lower in the 50 ng/L 17MT exposure groups compared to the control groups. VTG, FSH, LH, 11-KT, E2, and T were significantly lower in the groups exposed to 100 ng/L 17MT. High-throughput sequencing revealed 73,449 unigenes, 1205 known mature miRNAs, and 939 novel miRNAs in the gonads of G. rarus. With miRNA-seq, 49 (MT25-M vs. Con-M), 66 (MT50-M vs. Con-M), and 49 (MT100-M vs. Con-M) DEMs were identified in the treatment groups. Five mature miRNAs (miR-122-x, miR-574-x, miR-430-y, lin-4-x, and miR-7-y), as well as seven differentially expressed genes (soat2, inhbb, ihhb, gatm, faxdc2, ebp, and cyp1a1), which may be associated with testicular development, metabolism, apoptosis, and disease response, were assayed using qRT-PCR. Furthermore, miR-122-x (related to lipid metabolism), miR-430-y (embryonic development), lin-4-x (apoptosis), and miR-7-y (disease) were differentially expressed in the testes of 17MT-exposed G. rarus. This study highlights the role of miRNA-mRNA pairs in the regulation of testicular development and immune response to disease and will facilitate future studies on the miRNA-RNA-associated regulation of teleost reproduction. [ABSTRACT FROM AUTHOR]

17α-Methyltestosterone (MT), a synthetic environmental endocrine disruptor with androgenic effects, has been shown to disrupt the reproductive system and inhibit germ cell maturation in Gobiocypris rarus. To further investigate the regulation of gonadal development by MT through the hypothalamic-pituitary-gonadal (HPG) axis, G. rarus were exposed to 0, 25, 50, and 100 ng/L of MT for 7, 14, and 21 days. We analyzed its biological indicators, gonadotropin-releasing hormone (GnRH), gonadotropins, reproduction-related gene expression, and brain tissue transcriptome profiles. We found a significant decrease in the gonadosomatic index (GSI) in G. rarus males exposed to MT for 21 days compared to the control group. GnRH, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) levels, as well as the expressions of the gnrh3, gnrhr1, gnrhr3, fshβ, and cyp19a1b genes, were significantly reduced in the brains of both male and female fish when exposed to 100 ng/L MT for 14 days compared to the controls. Therefore, we further constructed four RNA-seq libraries from 100 ng/L MT-treated groups of male and female fish, obtaining 2412 and 2509 DEGs in male and female brain tissue, respectively. Three common pathways were observed to be affected in both sexes after exposure to MT, namely, nicotinate and nicotinamide metabolism, focal adhesion, and cell adhesion molecules. Furthermore, we found that MT affected the PI3K/Akt/FoxO3a signaling pathway through the upregulation of foxo3 and ccnd2, and the downregulation of pik3c3 and ccnd1. Therefore, we hypothesize that MT interferes with the levels of gonadotropin-releasing hormone (GnRH, FSH, and LH) in G. rarus brains through the PI3K/Akt/FoxO3a signaling pathway, and affects the expression of key genes in the hormone production pathway (gnrh3, gnrhr1 and cyp19a1b) to interfere with the stability of the HPG axis, thus leading to abnormal gonadal development. This study provides a multidimensional perspective on the damaging effects of MT on fish and confirms that G. rarus is a suitable model animal for aquatic toxicology. [ABSTRACT FROM AUTHOR]

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The 17α-methyltestosterone is the most common synthetic hormone used in male mono-sex production of Nile tilapia, Oreochromis niloticus. The current research aimed at finding out the most effective dose of 17α-methyltestosterone to produce quality Nile tilapia fry. Soon after absorbing the yolk sac, Nile tilapia fry was fed with a mixture of commercial fish feed and 17α-methyltestosterone for 28 days. Five doses of 17α-methyltestosterone, i.e., 0 mg, 50 mg, 60 mg, 70 mg, and 80 mg per kg feed, were used to treat tilapia that has been reared for additional 90 days to compare sex reversal, development, and survival rates. Both gonad histology and Squash test were performed to expose the sex percentage of O. niloticus accurately. The highest male 94.44% was obtained at 60 mg 17α-MT/kg feed dose followed by 91.67%, 88.89%, 86.11%, and 47.22% at 70, 80, 50, and 0 mg 17α-MT/kg feed dose. The groups treated with 17α-methyltestosterone hormone showed superior growth performance in comparison to the control group. The highest weight (14.62 ± 0.59 g) and length (92.18 ± 3.01 mm) were found at 60 mg dose whereas the lowest weight (8.64 ± 0.38 g) and length (70.17 ± 3.75 mm) were in the control group. The group given 60 mg 17α-MT feed represented the highest survival rate (84.10%) among other hormone-treated groups. The study disclosed that 60 mg 17α-MT/kg feed might be treated as the optimal dose for producing quality mono-sex male tilapia in the commercial hatchery.

The potential of pine pollen (PP) to masculinize sexually undifferentiated Nile tilapia was evaluated by feeding graded levels of PP (0.08–3.20 g kg-1 basal diet) to triplicate groups of three-day-old Nile tilapia for 28 days. Masculinization and associated differences in growth were compared to fish individuals fed the same basal diet with no PP (CT; negative control) and with 0.06 g 17α-methyltestosterone (MT) kg-1 basal diet (positive control). Both PP and MT treatments significantly skewed the expected 50:50 (male: female) ratio towards more male individuals. Notably, MT and 1.28 g PP kg-1 of diet produced a significantly high proportion of males (89.2 ± 2.2% and 80.0 ± 2.9% respectively), compared to 50.8 ± 2.2% in the CT treatment (P

This study aimed to investigate the effects of 17α-Methyltestosterone (MT) on hepatic lipid metabolism in Gobiocypris rarus. G. rarus was exposed to varying concentrations of MT (0, 25, 50, and 100 ng/L) for durations of 7, 14, and 21 d. Biochemical and transcriptomic analyses were conducted using methods, such as ELISA, RT-qPCR, Western Blotting, and RNA-seq, to decipher the key signals and molecular mechanisms triggered by MT in vivo. The results revealed that MT induced hepatomegaly in G. rarus and markedly increased the hepatic steatosis index (HSI). After 14 d of exposure, significant increase in PPARγ mRNA expression was observed, whereas after 21 d, PPARα mRNA expression was significantly reduced. The expression pattern of SREBP1C mRNA initially decreased before increasing, mirroring the trend observed for SREBP1C protein expression. Furthermore, MT increased the levels of key lipid synthesis enzymes, including HSL, CPT1, GPAT, and FAS, thereby fostering lipid accumulation. RNA-seq analysis revealed that MT modulated hepatic bile acid metabolism via the PPAR pathway, consequently influencing cholesterol and lipid metabolism. Considering the differential metabolic pathways of MT across genders, it is postulated that MT may undergo aromatization to estrogen within G. rarus , thereby exerting estrogenic effects. These findings provide crucial experimental insights into the detrimental effects of MT in aquatic settings, underscoring its implications for safeguarding aquatic organisms and human health. [Display omitted] • MT promotes lipid accumulation in the liver of Gobiocypris rarus. • MT increases HSI, promotes the expression of genes related to lipid synthesis, affects triglyceride synthesis and metabolism. • MT can regulate hepatic bile acid metabolism through the PPAR pathway, thereby influencing cholesterol and lipid metabolism. • The response mechanisms to MT in the liver differ between female and male G. rarus. [ABSTRACT FROM AUTHOR]

Objective(s): This study aims to enhance 17 a-methyltestosterone loaded human serum albumin nanoparticles (MT-HSA NPs) bioavailability through a desolvation technique. Dopamine (DA) molecules were conjugated on the surface of MT-HSA NPs and have the potential to act as tiny proper ligands in a unique treatment system to cope with cancer in which drug will be transmitted to the cancer area. Herein, we used HSA as an adaptable carrier of anticancer agents for methyltestosterone transport to the tumor site via DA D1-D5 receptors. In the present study, sonication of MT-HSA solution was carried out before the desolvation procedure to increase the drug loading and entrapment efficiency. Materials and Methods: Various parameters were optimized to characterize NPs including morphology, size, zeta potential, polydispersity index, drug release profile, and entrapment efficiency. Results: Under the optimum conditions of HSA and drug (1:41), at pH 9, results demonstrate sizes of 69 nm and 82 nm for MT-HSA and MT-HSA-DA NPs respectively. For MT-HSA NPs, the polydispersity index was found to be 0.3 and the average drug loading and encapsulation efficiency were 14% and 91% respectively. Anticancer activity and the release of drug was investigated through MCF-7 breast cancer cell line. Results show that targeted NPs are more effective than non-targeted NPs. Conclusion: According to these studies, the therapeutic effects against various diseases such as cancers increase through cellular targeting property of a biocompatible drug delivery system. This is the first report for methyltestosterone delivery to breast cancer cells based on HSA NPs.

The steroid 17α-methyltestosterone (MT) inhibits ovarian function and is often used to induce sex reversal artificially in vertebrates. In the present study, different concentrations of MT were added as dietary supplementation, and the effects on sex ratio, growth, and gonadal development were examined. After 40 days, the sex ratio (male:female) in each group increased at different degrees with 50 (1.36:1), 100 (1.57:1), and 200 (2.61:1) mg/kg MT, and neo-males with testis–ovary coexistence were observed in the 200 mg/kg MT group. Furthermore, 50 and 100 mg/kg MT could induce female reversion in neo-males. Histologically, the development of the testes in experimental groups was slower, but the ovaries of the experimental and control groups had similar developmental rates. The expression levels of DMRT11E, Foxl2, and SoxE1 in males at 200 mg/kg MT were 8.65-, 3.75-, and 3.45-fold greater than those of the control group. In crustaceans, sex reversal through vertebrate sex hormones can be observed. Neo-males (sex-reversed female prawns) were maintained by exogenous androgen, and over-reliance led to slow testis growth, small body size, and low growth rate, but sperm was still produced. In female prawns, MT inhibited ovary development and promoted growth.

Simple Summary: Although gonochoristic fish express one sex or the other in adulthood, some adults display traits similar to those of their opposite sex, suggesting plasticity similar to hermaphrodites. To fully test this potential in the eastern mosquitofish, a gonochorist, two adult stages of females were fed 17α-Methyltestosterone incorporated feed (0–200 mg/kg diet) for 50 days. The hormone (particularly at 50 mg/kg diet) stimulated the formation of complex male copulatory structures and testicular tissue as well as upregulated expression of Anti-Müllerian Hormone gene and altered the behavior from females to males. Collectively, we infer that Gambusia holbrooki retains potential for sex reversal at adulthood, similar to what occurs naturally in hermaphroditic fish and can provide an ideal system to investigate these mechanisms in a regulated fashion. 17α-Methyltestosterone (MT) is a synthetic steroid that has been widely used to masculinize many fish species when administered early during larval development, however, reports on its efficacy on adults is limited. To this end, this study investigated the efficacy of MT in the masculinization of the eastern mosquitofish (G. holbrooki) at two adult stages (maiden and repeat gravid females). The treated females were fed control or respective MT incorporated feed (0–200 mg/kg diet) for 50 days. Effects of the hormone on secondary sexual characteristics, internal gonad morphology, expression of the Anti-Müllerian Hormone (amh) gene and sexual behavior of the treated females were investigated. The results showed that MT at the dose of 50 mg/kg feed stimulated secondary sexual character development, upregulated expression of amh, formation of testicular tissue and a shift in the behavior similar to those of normal males, prominently so in treated maiden gravid females. Post-treatment, long-term observations indicated that only two masculinized females reverted back to being females and gave birth to young. Induction of masculinizing effects in most individuals suggests that the sexual phenotype of this species appears to be highly plastic with potential to sex reverse at adulthood. This in combination with its small size and short reproductive cycle could provide an ideal system to explore the mechanisms of sequential hermaphroditism in fish and contribute to genetic control of this pest fish. [ABSTRACT FROM AUTHOR]

The purpose of this study was to evaluate the influence of different 17a-methyltestosterone injection frequencies at optimal doses to improve the quality of spermatophore and reproductive performance of captive male broodstock of tiger shrimp Penaeus monodon. Broodstocks of tiger shrimp weighing 51-65 g for males and 86-161 g for females were collected from ponds and stored in a 3,000 m3 concrete tank. Male broodstocks were injected with 17a-methyltestosterone at the optimized protocol at 300 ng/100 g body weight at different injection frequencies, three times, four times, and five times at seven-day intervals. As a control, male broodstocks were injected with a saline solution to determine the spermatophore quality. Evaluation of the reproductive performance of the captive male was observed in the captive female broodstock tiger shrimp using artificial insemination. The weight of spermatophore (0.041 g), gonadosomatic index (7.71%), the total number of spermatozoa (64 × 106 cells/mL) significantly increased, and the percentage of abnormal spermatozoa significantly decreased by 73% as well as hatching rate increased to 43.14% at the injection frequency of four times compared to the other treatments. In conclusion, injection of 17a-methyltestosterone at the dosage of 300 ng/100 g body weight at the frequency of giving four times was effective enough to improve the reproductive performance of captive tiger shrimp. Therefore, the method proposed can be a strategy to produce better larvae and increase the income of the aquaculture business. [ABSTRACT FROM AUTHOR]

Objective: To examine sex reversal both by oral and by immersion using 17α-methyltestosterone on the methyltestosterone residual concentration and the organ histopathology of tilapia fish. Methods: This study used oral and immersion treatment methods for sex reversal of tilapia fish and used normal fish as the control and each treatment was repeated 4 times. 17α-methyltestosterone at dosages of 60 mg/kg feed and 0.5 mg/L were used for oral and immersion methods, respectively. In the first step, tilapia fry were reared at 100 L aquaria, with a density of 1 fish/L for 2 months. In the next step, male tilapias were reared at happa (net cage) of (2×1×1) m3 size in the controlled pond, with a density of 30 fish/happa for 3 months. The methyltestosterone residual concentrations were analyzed by one-way analysis of variance and Duncan’s multiple range tests, while organ histopathology was analyzed by descriptive method. Results: Residual concentrations in the serum of methyltestosterone- treated fish were significantly lower than that in normal fish, especially in 4- and 5-month-old tilapias with averages of less than 5 μg/L, while in normal fish was more than 5 μg/L. In the flesh, methyltestosterone residual concentrations showed relatively no significant differences between the oral and immersion treatment groups and methyltestosterone-treated fish remained lower compared to normal fish, except in 5-month-old tilapia. Methyltestosterone-treated tilapia exhibited histopathological changes on gill, liver, kidneys, and intestine organs. Conclusions: Sex reversal either by oral or by immersion has methyltestosterone residual concentration, but does not exceed the limits (5 μg/L or 5 μg/kg) of synthetic steroid on the fish body, although methyltestosterone causes histopathological changes on gill, liver, kidneys, and intestine.

This study aimed to investigate the effects of 17α-Methyltestosterone (MT) on hepatic lipid metabolism in Gobiocypris rarus. G. rarus was exposed to varying concentrations of MT (0, 25, 50, and 100 ng/L) for durations of 7, 14, and 21 d. Biochemical and transcriptomic analyses were conducted using methods, such as ELISA, RT-qPCR, Western Blotting, and RNA-seq, to decipher the key signals and molecular mechanisms triggered by MT in vivo. The results revealed that MT induced hepatomegaly in G. rarus and markedly increased the hepatic steatosis index (HSI). After 14 d of exposure, significant increase in PPARγ mRNA expression was observed, whereas after 21 d, PPARα mRNA expression was significantly reduced. The expression pattern of SREBP1C mRNA initially decreased before increasing, mirroring the trend observed for SREBP1C protein expression. Furthermore, MT increased the levels of key lipid synthesis enzymes, including HSL, CPT1, GPAT, and FAS, thereby fostering lipid accumulation. RNA-seq analysis revealed that MT modulated hepatic bile acid metabolism via the PPAR pathway, consequently influencing cholesterol and lipid metabolism. Considering the differential metabolic pathways of MT across genders, it is postulated that MT may undergo aromatization to estrogen within G. rarus, thereby exerting estrogenic effects. These findings provide crucial experimental insights into the detrimental effects of MT in aquatic settings, underscoring its implications for safeguarding aquatic organisms and human health., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2024. Published by Elsevier Inc.)

The sex determination system of largemouth bass ( Micropterus salmoides , LMB) is XX/XY; however, the underlying molecular mechanisms involved in early sex differentiation, gonadal development, and exogenous hormone-induced sex reversal remain unknown. In this study, LMB at 15 days post-hatching (dph) were fed diets containing 20 mg/kg of 17α-methyltestosterone (17α-MT) or 30 mg/kg of 17β-estradiol (17β-E 2 ) for 60 days, respectively. Serum steroid levels, histological observations of the gonads, and identification of sex-specific markers were employed to screen the gonads of 60-day-old normal female fish (XX-F), normal male fish (XY-M), 17β-E 2 induced pseudo-female fish (XY-F), and 17α-MT-induced pseudo-male fish (XX-M) for transcriptome sequencing in order to uncover genes and pathway involved in the process of sexual reversal. The results from histology and serum sex steroid hormone analysis showed that both 17α-MT and 17β-E2 were capable of inducing sex reversal of LMB at 15 dph. Transcriptome results revealed a total of 2,753 genes exhibiting differential expression, and the expression pattern of these genes in the gonads of XX-M or XY-F resembled that of normal females or males. The male sex-biased genes that are upregulated in XX-M and downregulated in XY-F are referred to as key genes for male reversal, while the female sex-biased genes that are upregulated in XY-F and downregulated in XX-M are referred to as key genes for female reversal. Finally, 12 differentially expressed genes (DEGs) related to male sex reversal were screened, including star2 , cyp17a , cyp11b1 , dmrt1 , amh , sox9a , katnal1 , spata4 , spata6l , spata7 , spata18 and foxl3 . 2 DEGs ( foxl2a and cyp19a1b ) were found to be associated with female sex reversal. The changes in these genes collectively influence the direction of sex differentiation of LMB. Among them, star2 , dmrt1 and cyp19a1b with significantly altered expression levels may play potentially crucial role in the process of gender reversal. The expression patterns of 21 randomly selected genes were verified using qRT-PCR which confirmed the reliability and accuracy of the RNA-seq results. These findings not only enhance our understanding of the molecular basis underlying sex reversal but also provide crucial data support for future breeding research on unisexual LMB., Competing Interests: LH was employed by Guangdong Province Liangshi Aquaculture Seed Industry. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhang, Tian, Li, Du, Lei, Zhu, Han and Song.)

Copyright of Asian Journals of Ecotoxicology is the property of Gai Kan Bian Wei Hui and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Elucidating the global molecular changes that occur during aromatase inhibitor (AI)– or 17α-methyltestosterone (MT)–induced masculinization and estradiol-17β (E2)–induced feminization is critical to understanding the roles that endocrine and genetic factors play in regulating the process of sex differentiation in fish. Here, fugu larvae were treated with AI (letrozole), MT, or E2 from 25 to 80 days after hatching (dah), and gonadal transcriptomic analysis at 80 dah was performed. The expression of dmrt1 , gsdf , foxl2 , and other key genes (star , hsd3b1 , cyp11c1, cyp19a1a , etc.) involved in the steroid hormone biosynthesis pathway were found be altered. The expression of dmrt1 , gsdf , cyp19a1a , and foxl2 was further verified by quantitative polymerase chain reaction. In the control group, the expression of dmrt1 and gsdf was significantly higher in XY larvae than in XX larvae, while the expression of foxl2 and cyp19a1a was significantly higher in XX larvae than in XY larvae (P <.05). AI treatment suppressed the expression of foxl2 and cyp19a1a , and induced the expression of dmrt1 and gsdf in XX larvae. MT treatment suppressed the expression of foxl2 , cyp19a1a , dmrt1 , and gsdf in XX larvae. E2 treatment suppressed the expression of dmrt1 and gsdf , but did not restore the expression of foxl2 and cyp19a1a in XY larvae. The shared response following AI, MT, and E2 treatment suggested that these genes are essential for sex differentiation. This finding offers some insight into AI or MT-induced masculinization, and E2-induced femininization in fugu. [ABSTRACT FROM AUTHOR]

Steroid hormones are essential for the normal reproductive activity in females and males, such as early sex differentiation, gonad development and sex reversal, which works by binding with the steroid receptors. In the present study, the full-length of estrogen receptors (ERα, ERβ1 and ERβ2) and androgen receptors (ARα and ARβ) cDNAs were isolated from the mandarin fish (Siniperca chuatsi). Real time quantitative PCR (RT-qPCR) revealed that ERα and ERβ2 expressed mainly in the liver of females and males, and ERβ1 mostly distributed in ovary and testis. Additionally, the high expressions of ARα and ARβ in both females and males were detected in gonads and livers respectively. During the early sex differentiation stage, the expressions of ERs and ARs in female were higher than in male during 5–30 day after hatching (dah), and reached the peak level at 30 dah. In males, the expressions of ERβ1, ERβ2, and ARβ increased largely during 30 dah to 60 dah, and revealed significantly higher levels than in females during 60–180 dah. In female, the expression of ERβ2 in ovary increased sharply at 180 dah. Moreover, the result of gonadal histology revealed that the spermatocytes were observed at 45 dah and the spermatozoon could be found largely at 60 dah in males and oil droplets appeared for the first time in the oocytes at 180 dah. The expression levels of three ERs and ARβ were up-regulated and the ARα was down-regulated during female to male sex reversal induced by 17α-methyltestosterone (MT). Furthermore, after MT treatment for 30 and 35 days, the expressions of ERβ1, ERβ2 and ARβ were significantly up-regulated. These results suggest that ERβ1, ERβ2 and ARβ may play critical roles in spermatogenesis and masculinization, while ERβ2 and ERβ1 may be essential for vitellogenesis and ovary development in female.

17α-Methyltestosterone (MT) is a synthetic androgen. The objective of this study was to explore the effects of exogenous MT on the growth and gonadal development of female rare minnow Gobiocypris rarus. Female G. rarus groups were exposed to 25–100 ng/L of MT for 7 days. After exposure for 7 days, the total weight and body length were significantly decreased in the 50-ng/L MT groups. The major oocytes in the ovaries of the control group were vitellogenic oocytes (Voc) and cortical alveolus stage oocytes (Coc). In the MT exposure groups, some fish had mature ovaries with a relatively lower proportion of mature oocytes, and the diameter of the perinucleolar oocytes (Poc) was decreased compared with those of the control group. Ovarian VTG, FSH, LH, 11-KT, E2, and T were significantly increased after exposure to 50 ng/L of MT for 7 days. Unigenes (73,449), 24 known mature microRNAs (miRNAs), and 897 novel miRNAs in the gonads of G. rarus were found using high-throughput sequencing. Six mature miRNAs (miR-19, miR-183, miR-203, miR-204, miR-205, and miR-96) as well as six differentially expressed genes (fabp3, mfap4, abca1, foxo3, tgfb1, and zfp36l1) that may be associated with ovarian development and innate immune response were assayed using qPCR. Furthermore, the miR-183 cluster and miR-203 were differentially expressed in MT-exposed ovaries of the different G. rarus groups. This study provides some information about the role of miRNA–mRNA pairs in the regulation of ovarian development and innate immune system, which will facilitate future studies of the miRNA–RNA-associated regulation of teleost reproduction.

17α-Methyltestosterone (MT) is a synthetic androgen. The objective of this study was to explore the effects of exogenous MT on the growth and gonadal development of female rare minnow Gobiocypris rarus. Female G. rarus groups were exposed to 25–100 ng/L of MT for 7 days. After exposure for 7 days, the total weight and body length were significantly decreased in the 50-ng/L MT groups. The major oocytes in the ovaries of the control group were vitellogenic oocytes (Voc) and cortical alveolus stage oocytes (Coc). In the MT exposure groups, some fish had mature ovaries with a relatively lower proportion of mature oocytes, and the diameter of the perinucleolar oocytes (Poc) was decreased compared with those of the control group. Ovarian VTG, FSH, LH, 11-KT, E2, and T were significantly increased after exposure to 50 ng/L of MT for 7 days. Unigenes (73,449), 24 known mature microRNAs (miRNAs), and 897 novel miRNAs in the gonads of G. rarus were found using high-throughput sequencing. Six mature miRNAs (miR-19, miR-183, miR-203, miR-204, miR-205, and miR-96) as well as six differentially expressed genes (fabp3 , mfap4 , abca1 , foxo3 , tgfb1 , and zfp36l1) that may be associated with ovarian development and innate immune response were assayed using qPCR. Furthermore, the miR-183 cluster and miR-203 were differentially expressed in MT-exposed ovaries of the different G. rarus groups. This study provides some information about the role of miRNA–mRNA pairs in the regulation of ovarian development and innate immune system, which will facilitate future studies of the miRNA–RNA-associated regulation of teleost reproduction. [ABSTRACT FROM AUTHOR]

17α-methyltestosterone (MT) is an exogenous anabolic steroid hormone used widely in aquafarming. This study aimed to emphasize the environmental hazards of MT originated from a continuously operating masculinization pond with an intensive water recirculation system in Samut Prakan, Thailand. Risk quotient (RQ) and concentration factor (CF) were used to examine the ecological risk of MT in the pond environment. The results revealed relatively high MT concentrations in the water and sediments. These findings concurred the benefit of masculinization ponds with a water recirculation system in the way that it can effectively save the water and reduce the MT dosage that needs to be added into the feed. However, this could bring serious environmental concerns since releasing water containing MT residues and androgenic active metabolites through water overflowing from the masculinization pond to the environment during the wet season is inevitable. This is the first report indicating that CF relates to the androgenic potential activity and can be used to explain the ecological risk of MT contaminated water discharged from a full water recirculating earthen masculinization pond. This study provides important information indicating that CF values in a range of 7 to 16 are high enough to cause adversely affect the reproductive performance of tested fish (i.e., Probarbus jullieni, Barbonymus gonionotus, and Anabas testudineus). [ABSTRACT FROM AUTHOR]

• AKR1Cs display 5α-reductase activities towards oxymesterone, and the catalytic activities of AKR1C9 and AKR1C4 are higher than those of AKR1C1, AKR1C2 and AKR1C3. • 5α-reductase activities of AKR1Cs towards oxymesterone were higher than towards 17α-methyltestosterone, likely due to the presence of the C4‑hydroxyl group. • In contrast to 4-hydroxytestosterone, AKR1Cs has no 3α/β-HSD activity towards oxymesterone, likely due to the presence of the C17α-methyl group. • Oxymesterone was interconverted with 17α-methyltestosterone by both dehydroxylation and hydroxylation reactions catalyzed by AKR1Cs. The function of aldo-keto reductases (AKRs) is highly variable and is controlled by various groups within the steroid structure. The representative AKRs are AKR1C1-AKR1C4 from humans and AKR1C9 from rats (AKR1Cs). In this study, we focused on how the C4‑hydroxyl in the A-ring and C17α-methyl in the d-ring of oxymesterone (OXY) affect the catalytic function of AKR1Cs. We found that OXY bound to all AKR1Cs with greater affinity for AKR1C9 and AKR1C2, according to both autodocking predictions and binding experiments using microscale thermophoresis. Conversion and enzyme kinetic studies showed that AKR1C4 and AKR1C9 catalyzed approximately 90 % of OXY and that their Kcat and Kcat/Km values were higher than other AKR1Cs. Moreover, both purified AKR1C enzymes and AKR1C-transfected cells reduced OXY to 17α-methyl-4ξ,17β-dihydroxy-androstane-3-one with 5α-reductase activity, that was promoted by C4‑hydroxyl. Nevertheless, AKR1Cs lost the 3α/3β-hydroxysteroid dehydrogenase activity for OXY due to C17α-methyl and did not form 17α-methyl-3ξ, 4ξ, 17β-trihydroxy-androstane. [Display omitted] [ABSTRACT FROM AUTHOR]

The present study aimed to develop novel masculinization agents for enhancing sex-reversal in red tilapia (Oreochromis sp.) by immersion administration using 17α-methyltestosterone-loaded nanostructured lipid carriers (MT-NLC). Moreover, the surface charge of the MT-NLC was modified with chitosan (CS-MT-NLC) to improve their mucoadhesive property. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to characterize the morphology of the nanoparticles. Furthermore, we assessed the toxicity associated with immersion-induced masculinization in red tilapia. Subsequently, we examined alterations in gill morphology, determined the percentage of masculinization, analyzed their length-weight relationship, and investigated the expression of genes related to sex differentiation. MT-NLC had an average size, polydispersity index, and negatively charged surface of 140.8 ± 2.6 nm, <0.2, and −26.9 ± 0.3 mV, respectively, with higher stability at 4 °C and 25 °C compared to CS-MT-NLC. The release mechanisms of NLC and CS-NLC were regulated by diffusion processes, which were characterized using Avrami's equation. In contrast, the release mechanism of non-encapsulated MT (MT-ET) was governed by burst release kinetic. Furthermore, tests assessing mucin binding confirmed that CS-MT-NLC, with a positive charge, exhibited superior mucoadhesive properties compared to MT-NLC, with a negative charge. Importantly, CS-MT-NLC immersion was found to be toxic to red tilapia fry; therefore, it has been discontinued for further experiments. The immersions of both MT-NLC and MT-ET were non-toxic to red tilapia fry, with an LC 50 exceeding 800 μg/L, and the morphological appearance of the gills after treatment was normal. The MT-NLC-400 and MT-ET-800 groups exhibited elevated expression levels of dmrt1 and sox9 compared to cyp19a1a and soxl2. These higher expression levels corresponded to significantly higher percentages of males (98.3% and 95.7%) in both the MT-NLC and MT-ET groups compared to the control group (54.7%). The MT-NLC and MT-ET groups displayed a negative allometric growth behavior in red tilapia fry, with a relative condition factor > 1. Adopting MT-NLC-400 immersion treatment for masculinizing red tilapia could serve as a beneficial strategy for commercial tilapia farms to reduce the required dosage and the time needed to treat fish. • Novel masculinization agents for enhancing sex-reversal efficiency in red tilapia by immersion using17α-alpha-methyltestosterone-loaded nanostructured lipid carriers (MT-NLC). • MT-NLC showed140.8 nm avg. size, <0.2 polydispersity index, and −26.9mV surface charge, with improved stability at cool and ambient temps. • Red tilapia fry exhibited no toxicity to MT-NLC and MT-ethanol (MT-ET) treatments by immersion, with LC 50 >800 μg/L and normal gill morphology. • MT-NLC-400 obtained 98.3% males with heightened expressions of dmrt1 and sox9 , compared to 54.7% males in control. • Red tilapia farming can use MT-NLC-400 immersion to reduce the dose and duration of MT treatment for masculinization. [ABSTRACT FROM AUTHOR]

The objective of this study was to reveal the molecular mechanisms involved in the exogenous androgen and aromatase inhibitor‐induced gonadal masculinization of all‐female triploid rainbow trout (RBT) populations. 17α‐methyltestosterone (MT) and letrozole (LET) were used for treatment. The female triploid (XXX) rainbow trout is an effective model to study the molecular mechanism of gonadal masculinization, owing to its tendency towards masculinization under natural conditions. The MT and LET were supplemented with a diet in ethanol at 1.5 mg/kg and 50 mg/kg of food. The treatments did not influence ovary development in female triploid RBT in the early stage (56–80 days post fertilization, i.e. dpf). However, LET could more successfully induce the morphological masculinization of gonads when compared with MT, from 94 dpf to 377 dpf. Gene expression in gonads was analysed after MT and LET treatment. The results showed that expression of male‐specific genes (sdy, dmrt1, sox9 and amh) increased more significantly in the LET group than in the control and MT group, and expression of female‐specific genes (cyp19a1a and foxl2) in LET group tended to decrease significantly. At 377 dpf, significant inhibition of female‐specific genes was shown in the LET group, and male‐specific gene expression increased to levels almost similar to that in diploid male samples. Our preliminary results suggest that the genes involved in gonadal development are pivotal factors in fish sex reversal, and aromatase inhibitor is a more effective inducer of masculinization when compared with androgen. [ABSTRACT FROM AUTHOR]

Objective(s): This study aims to enhance 17 a-methyltestosterone loaded human serum albumin nanoparticles (MT-HSA NPs) bioavailability through a desolvation technique. Dopamine (DA) molecules were conjugated on the surface of MT-HSA NPs and have the potential to act as tiny proper ligands in a unique treatment system to cope with cancer in which drug will be transmitted to the cancer area. Herein, we used HSA as an adaptable carrier of anticancer agents for methyltestosterone transport to the tumor site via DA D1-D5 receptors. In the present study, sonication of MT-HSA solution was carried out before the desolvation procedure to increase the drug loading and entrapment efficiency. Materials and Methods: Various parameters were optimized to characterize NPs including morphology, size, zeta potential, polydispersity index, drug release profile, and entrapment efficiency. Results: Under the optimum conditions of HSA and drug (1:41), at pH 9, results demonstrate sizes of 69 nm and 82 nm for MT-HSA and MT-HSA-DA NPs respectively. For MT-HSA NPs, the polydispersity index was found to be 0.3 and the average drug loading and encapsulation efficiency were 14% and 91% respectively. Anticancer activity and the release of drug was investigated through MCF-7 breast cancer cell line. Results show that targeted NPs are more effective than non-targeted NPs. Conclusion: According to these studies, the therapeutic effects against various diseases such as cancers increase through cellular targeting property of a biocompatible drug delivery system. This is the first report for methyltestosterone delivery to breast cancer cells based on HSA NPs. [ABSTRACT FROM AUTHOR]

17α-Methyltestosterone (MT) is a synthetic steroid that has been widely used to masculinize many fish species when administered early during larval development, however, reports on its efficacy on adults is limited. To this end, this study investigated the efficacy of MT in the masculinization of the eastern mosquitofish (G. holbrooki) at two adult stages (maiden and repeat gravid females). The treated females were fed control or respective MT incorporated feed (0–200 mg/kg diet) for 50 days. Effects of the hormone on secondary sexual characteristics, internal gonad morphology, expression of the Anti-Müllerian Hormone (amh) gene and sexual behavior of the treated females were investigated. The results showed that MT at the dose of 50 mg/kg feed stimulated secondary sexual character development, upregulated expression of amh, formation of testicular tissue and a shift in the behavior similar to those of normal males, prominently so in treated maiden gravid females. Post-treatment, long-term observations indicated that only two masculinized females reverted back to being females and gave birth to young. Induction of masculinizing effects in most individuals suggests that the sexual phenotype of this species appears to be highly plastic with potential to sex reverse at adulthood. This in combination with its small size and short reproductive cycle could provide an ideal system to explore the mechanisms of sequential hermaphroditism in fish and contribute to genetic control of this pest fish.

Sustainable development of Vaname shrimp aquaculture in the future depends on several things, one of which is the availability of the male parent in a sufficient quantity. Regarding this problem, sex reversal can be performed as the solution. Sex reversal is a technique of sex genetic differentiation to change sexes artificially from males genetically into phenotype females or vice versa. This research was intended to obtain the maximal mono-sex (single sex) male of Vaname shrimp using 17α-methyltestosterone (MT) hormone through sex reversal technology to the limited sex change in Vaname shrimp, in which to date the Vaname male parent gained is still dependent on natural selection. This research used a descriptive and experimental method. The experimental research design used was a completely randomized design (RAL – Rancangan Acak Lengkap). The treatment given was immersing the shrimp larvae in 17α-methyltestosterone (MT) for 8 hours at different doses of 0 mg/L, 5 mg/L, 8 mg/L, and 12 mg/L. The immersion of the post-larvae (PL1) of Vaname shrimp using the 17α-methyltestosterone hormone gave a significant result (P < 0.05) on all of the treatments and control in which the highest percentage of male Vaname shrimp was obtained from the treatments with 8 mg/L and 12 mg/L doses, respectively reaching 64% and 66.7%. As for the weight growth, the immersion of the post-larvae (LP1) growth of Vaname shrimp using 17α-methyltestosterone hormone provided a very significant result (P < 0.05) on all the treatments and control in which the highest weight growth (weight gain) and Specific Growth Rate of Body Weight (SGR BW) were shown by the treatments with 8 mg/L and 12 mg/L doses, respectively reaching 1.5415 grams and 1.5871 grams.

In recent years, the increasing use of environmental endocrine disruptors has caused serious environmental pollution and hurt aquatic organisms. It is still risky for aquatic species and humans exposed to 17α-methyltestosterone (MT), however, the harmful effect of MT on fish is still poorly understood. The main purpose of this study is to evaluate the influence of MT on Pseudorasbora parva at multi-levels. We analyzed gonadal histology, the sex steroid hormones, steroidogenic genes expression, and transcriptome profiling of gonads in response to MT in adult P. parva. Through this study, we found MT could inhibit the gonadal development of P. parva , and the growth and development of fish could be delayed by exposure to MT at 200 ng/L. MT could produce disruption effects on fish from multiple pathways, while its interference to the HPGL axis happens primarily through the steroidogenic pathway, e.g., disturbing the expression of crucial genes and sex steroids synthesis. Besides, we constructed 4 RNAseq libraries and obtained 7758 and 11,543 DEGs in females and males, respectively. Interestingly, we found MT had more obvious disruption effects on males than the females, mainly reflected in the immune system. Interestingly, we found three common pathways in both sexes after MT exposure, i.e., cell adhesion molecules, cytokine-cytokine receptor interaction, and neuroactive ligand-receptor interaction. These results confirm the suitability of P. parva as a model fish for aquatic toxicological study and provide us a multidimensional sight for the disruption effects of MT on fish. • MT could inhibit the gonadal development of P. parva. • MT could interference to the HPGL axis happens primarily through the steroidogenic pathway. • MT had more obvious disruption effects on males than the females. [ABSTRACT FROM AUTHOR]

Sex change was induced in Epinephelus marginatus juveniles using a nonsteroidal aromatase inhibitor (AI), a synthetic androgen (17α-methyltestosterone; MT), and a combination of both (MT + AI) in a 90-day experiment. A detailed remodeling of the gonads, the plasma level of gonadal steroids, and immunostaining of pituitary follicle-stimulating hormone (FSH), luteinizing hormone (LH), and somatolactin (SL) cells were analyzed. Sex inversion reached the final spermatogenesis stages using MT, while AI triggered spermatogenesis, but reaching only the spermatid stage. Estradiol (E2) levels did not change in fish treated with AI but decreased throughout the experimental period in animals treated with MT and MT + AI. Testosterone (T) levels increased in animals treated with MT during the first 60 days (and combined with AI in the first 30 days), decreasing in all experimental groups at 90 days, while AI-treated animals had increased plasma 11-ketotestosterone (11-KT) levels after 90 days. In control fish, FSH- and SL-producing cells (ir-FSH and ir-SL) were restricted to pars intermedia (PI) of the adenohypophysis. Pituitary ir-FSH cells were decreased at the end of the experimental period in all treatments compared with the CT animals. LH-producing cells (ir-LH) were present in proximal pars distalis (PPD) and pars intermedia (PI) of adenohypophysis and did not change after the experimental period. The decreased number of ir-FSH cells at the end of the experiment in all treatments could be related to the negative feedback loop triggered by the increase in natural and/or synthetic androgens. [ABSTRACT FROM AUTHOR]

Sexual phenotype is highly plastic in teleosts and controlled by a variety of species‐dependent factors. In aquaculture, hormone manipulations applied at different life stages are used to control sex ratios and exploit desirable traits such as ornate coloration in male ornamental species. This study investigated the effects of dietary 17α‐methyltestosterone (MT) administration on adult female Rosy Barbs Pethia conchonius and Dwarf Gourami Trichogaster lalius and Dwarf Gouramis. A panel of judges evaluated male secondary sex coloration weekly during and after prolonged MT feeding. Growth parameters, survival, and gonadosomatic index (GSI) were assessed at the end of the feeding regime. In both species, none of the MT diet concentrations affected survival, and all resulted in coloration changes, highlighting both the efficacy and safety of MT administration. In Dwarf Gouramis, MT diets caused pronounced color changes and reduced their GSIs at all dietary concentrations. In Rosy Barbs, MT diets induced the expression of male coloration but generally did not reach the color intensity of naturally occurring males and did not result in differences in GSIs. This study suggests that postdifferentiation androgenic treatment may be a viable option to induce secondary male sex coloration in ornamental species. [ABSTRACT FROM AUTHOR]

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Summary: This study investigated the effects of 17α‐methyltestosterone (17α‐MT) on the physiological properties of gynogenetic topmouth culter (Culter alburnus). Five 17α‐MT concentration (0, 10, 25, 40, and 60 μg/g) groups were set up in Experiment 1, and three treatment and recovery methods of the above five concentrations were studied in Experiment 2. In Experiment 3, fish were fed with increasing concentrations of 17α‐MT (from 40 μg/g to 140 μg/g) in an outdoor pond. In Experiment 1, growth was promoted during the initial 60 days in the 10 μg/g group, whereas inhibition was first observed at 20 days after treatment in the 60 μg/g group. Increased concentration (40–60 μg/g) of 17α‐MT significantly inhibited the expression of FOXL2 and CYP19a gene but significantly activated the expression of AMH and DMRT1 gene. The levels of superoxide dismutase, estradiol, testosterone, growth hormone, and tetraiodothyronine in the 40 and 60 μg/g groups were significantly higher than those in the other concentration groups at 40 days after treatment. In Experiment 2, almost all indexes detected besides the expression of reproduction‐related genes in the 17α‐MT treatment groups were recovered to those in the control groups. All gonads observed in Experiments 1 and 2 remained in a primordial state after treatment with different concentrations of 17α‐MT. In Experiment 3, 100% of the detected gonads developed into testes at 126 and 460 days post‐fertilization after three‐month outdoor 17α‐MT induction. These results suggest that 17α‐MT affects the physiological processes related to sexual differentiation in topmouth culter and that feeding 17α‐MT with increasing dose in an outdoor pond provides an effective protocol for producing neo‐male topmouth culter. [ABSTRACT FROM AUTHOR]

The effects of 17α-Methyltestosterone (MT) on sex reversal of Red tilapia (Oreochromis niloticus x Oreochromis mossambicus) in the free-swimming stage by immersion and feeding techniques were studied at the Department of Fisheries, Faculty of Agricultural Technology, Rajamangala University of Technology Thanyaburi, Pathumthani Province from April to July 2017. The experimental design was a factorial experiment in CRD which studied two factors: 1) sex reversal by immersion treatment of 17α-MT (0, 150 and 300 μg/l) for 3 hours (T1, T2, and T3) and 2) sex reversal by feeding treatment of 17α-MT (40, 60 and 80 mg/kg) for 28 days (F1, F2 and F3). Results from these studies indicated that during the free-swimming stage, the immersion treatment of 17α-MT in 300 μg/l for 3 hours (T3) had the highest growth rate in terms of weight, total length (28 days), ADG, FCR and survival rate (0.731 g, 3.340 cm, 0.024 g/d, 0.627 and 95.443 % respectively). The immersion treatment of 17α-MT in 150 μg/l for 3 hours (T2) had a medial growth rate among the other factors. The immersion treatment of 17α-MT in 0 μg/l for 3 hours (T1) had a minimal growth rate but had the highest sex ratio of males (83.889 %). The effects on sex reversal by feeding treatment of 17α-MT in 60 mg/kg for 28 days (F2) had the highest growth rate in terms of weight, ADG and FCR (0.644 g, 0.022 g/d and 0.714 respectively). The effects on sex reversal by the feeding treatment of 17α-MT in 80 mg/kg for 28 days (F3) had a medial growth rate and the highest survival rate (95.037 %) and sex ratio of males (85.667 %3T). The effects on sex reversal by feeding treatment of 17α-MT in 40 mg/kg for 28 days (FR1R) had a good growth rate but the other factors were minimal3T. [ABSTRACT FROM AUTHOR]

Abstract Androgens are a recognized class of endocrine disrupting compounds with the ability to impact reproductive status in aquatic organisms. The current study utilized in vitro exposure of mummichog (Fundulus heteroclitus) testis tissue to either the aromatizable androgen 17α-methyltestosterone (MT) or the non-aromatizable androgen 5α-dihydrotestosterone (DHT) over the course of 24 h to determine if there were differential effects on steroidogenic gene expression. Testis tissue was exposed to androgen concentrations of 10−12 M, 10−9 M and 10−6 M for 6, 12, 18 or 24 h, after which a suite of steroidogenic genes, including steroidogenic acute regulatory protein, 3β-hydroxysteroid dehydrogenase (3βhsd) and cytochrome P450 17A1 (cyp17a1), were quantified using real-time polymerase chain reaction. Both androgens affected steroidogenic gene expression, with most alterations occurring at the 24-hour time point. The gene with the highest fold-change, and shortest interval to expression alteration, was 3βhsd for both androgens. Potential differences between the two model androgens were observed in increased expression of cyp17a1 and 11β-hydroxysteroid dehydrogenase (11βhsd), which were only altered after exposure to DHT and in expression levels of cytochrome P450 11A1 (cyp11a1), which was upregulated by MT but not altered by DHT. Results from this study show both androgens interact at the gonadal level of the hypothalamus-pituitary-gonadal axis and may possess some distinct gene expression impacts. These data strengthen the current research initiatives of establishing in vitro test systems that allow toxic potential of untested chemicals to be predicted from molecular perturbations. [ABSTRACT FROM AUTHOR]

In this study, we have developed bridge heterologous ELISA for the detection of 17α- Methyltestosterone by incorporating aromatic spacers between 17α-Methyltestosterone-3-Carboxymethyloxime and Horseradish peroxidase label through N-hydroxysuccinimide mediated carbodiimide reaction method. The immunogen 17α-Methyltestosterone-3-Carboxymethyloxime-Bovine serum albumin used to generate the antibody was also prepared by the N-hydroxysuccinimide mediated carbodiimide reaction without using any spacer. We have studied the impact of bridge/aromatic spacers on functional parameters i.e. sensitivity, affinity and ED 50 of the bridge heterologous assay and compared it with homologous assay. The five combinations of bridge heterologous assay using 17α-Methyl testosterone-3-CMO-BSA antiserum and 17α-MT-3-CMO-4,4'-Diaminodiphenyl sulphide-HRP, 17α MT-3-CMO-4,4'-Oxydianiline-HRP, 17α-MT-3-CMO-Benzidine-HRP, 17α- MT-3-CMO-p-Phenylenediamine-HRP and 17α-MT-3-CMO-Dapson-HRP enzyme conjugates were evaluated. Out of these five combinations, the combination 17α-MT-3-CMO-BSA with 17α-MT-3-CMO-Benzidine-HRP showed the best results. Sensitivity, affinity and ED 50 were improved and found to be 0.02 ng/mL, 0.086 × 10−8 L/mol and 2.95 ng/mL than homologous assay where Sensitivity, affinity and ED 50 were 0.11 ng/mL, 0.02 × 10−8 L/mol and 5.78 ng/mL respectively. The cross-reactivity for this bridge heterologous assay combination was seen with only 4 steroids (6-hydrotestosterone- 6%, Testosterone-5.14%, Danazol-0.9% and Nandrolone-0.85%) instead of eight steroids (6-hydrotestosterone-43.75%, Testosterone-38.3%, Danazol-25.14%, Androstenediol-19.16%, Nandrolone-19%, Metandienone-5%, Androstenedione-3.52%, and 17α dimethyltestosterone-2%) as in homologous assay out of 59 structurally related steroids. Thus, the results of this study conclude that the incorporation of aromatic spacer (bridge) in enzyme conjugate has a crucial role in improving sensitivity, specificity, ED 50 and affinity of the developed assay. The assay was then studied for parameters such as recovery (97.4%–108.6%), precision (Inter and Intra-assay coefficient of variation <10%), correlation coefficient (R2 = 0.96) by comparing with the commercial kit and validated by measuring levels of 17α- methyltestosterone in rat serum after administering them. [Display omitted] • In this study, Five aromatic spacers; p-Phenylenediamine (PPD), 4,4'-Oxydianiline (ODA), 4,4'-Diaminodiphenyl sulphide (DPS), Benzidine (B), Dapson (D) for the first time have been used in the development of 17 alpha methyl testosterone ELISA. • Out of five combinations, the combination of antiserum 17α MT-3-CMO-BSA with enzyme conjugate 17α MT-3-CMO-Benzidine-HRP showed the best results. • This assay was studied further for analytical variables like Recovery, and Precision (Inter and Intra-assay coefficient of variation). • Correlation (R2 = 0.96) and validation of the assay was done by comparing with commercially available kit. • The study concludes that incorporation of aromatic spacer (bridge) in enzyme conjugate have fundamental role in improving sensitivity, specificity, ED 50 and affinity of the developed assay. [ABSTRACT FROM AUTHOR]