Incubation as a driver of maternal effects: Temperature influences levels of yolk maternally derived 5α-dihydrotestosterone.

In birds, maternal hormones deposited into eggs in response to environmental stimuli can impact offspring phenotype. Although less studied, environmental conditions can also influence females' incubation behavior, which might play a role in regulating embryo exposure to maternal hormones through changes in incubation temperature that affect the activity of the enzymes responsible for converting testosterone (T) to 5α-dihydrotestosterone (DHT) or estradiol. Here, we tested the hypothesis that the initial T content of the yolk and incubation temperature determine exposure to T metabolites during early embryo development. In the Japanese quail (Coturnix japonica), we experimentally manipulated yolk T and incubation temperature (38° C versus 36° C) and analyzed DHT and estradiol titers on day four of incubation. We found that eggs with experimentally increased T and those incubated at 36° C showed higher DHT concentration in egg yolk (with no synergistic effect of the two treatments). Estradiol titers were not affected by T manipulation or incubation temperature. Our study suggests that incubation temperature influences DHT titers and may act as an understudied source of maternal influence on offspring phenotype.

The Similarities and Differences between the Effects of Testosterone and DHEA on the Innate and Adaptive Immune Response.

Androgens are steroids that modulate various processes in the body, ranging from reproduction, metabolism, and even immune response. The main androgens are testosterone, dihydrotestosterone (DHT) and dehydroepiandrosterone (DHEA). These steroids modulate the development and function of immune response cells. Androgens are generally attributed to immunosuppressive effects; however, this is not always the case. Variations in the concentrations of these hormones induce differences in the innate, humoral, and cell-mediated immune response, which is concentration dependent. The androgens at the highest concentration in the organism that bind to the androgen receptor (AR) are DHEA and testosterone. Therefore, in this work, we review the effects of DHEA and testosterone on the immune response. The main findings of this review are that DHEA and testosterone induce similar but also opposite effects on the immune response. Both steroids promote the activation of regulatory T cells, which suppresses the Th17-type response. However, while testosterone suppresses the inflammatory response, DHEA promotes it, and this modulation is important for understanding the involvement of androgens in infectious (bacterial, viral and parasitic) and autoimmune diseases, as well as in the sexual dimorphism that occurs in these diseases.

Molecular genetic analysis of AKR1C2-4 and HSD17B6 genes in subjects 46,XY with hypospadias.

BACKGROUND: The formation of the male urethra depends to enzyme-mediated testosterone (T) conversion into 5α-dihydrotestosterone (DHT). Two metabolic pathways could be operating in the fetal testis to synthesize androgens: 1) the "classic" route (T→DHT) mediated by SRD5A2 and 2) a "backdoor" pathway in which DHT is synthesized by aldo-keto reductase family 1, member C2 (AKR1C2), AKR1C3, and AKR1C4 enzymes without formation of a T intermediate. OBJECTIVE: We studied four genes of the "backdoor" pathway in karyotypic males with hypospadias to ascertain whether gene defects in AKRs impair urethral DHT formation that result in hypospadias. DESIGN AND PATIENTS: The coding regions of the AKR1C2-4 and HSD17B6 genes were analyzed by PCR-SSCP and sequencing in a cohort of 25 Mexican patients (0.3-9 year-old-children) with 46,XY-hypospadias. Chi-squared tests was performed to evaluate the distribution of genotypes, alleles, and the Hardy-Weinberg (H-W) equilibrium. The effect of the genetic variants was investigated by in silico studies. RESULTS: Screening studies revealed distinct genotypic patterns at different exons of AKR1C2-4 whereas HSD17B6 presented a wild-type sequence. The DNA analyses detected two synonymous variants (c.327C>T, c.666T>C/unreported) in AKR1C2. The AKR1C3 had two variants (c.15C>G, c.230A>G), two unreported variants (c.538T>C, c.596G>A), and one silent variant (c.312G>A). Two variants (c.434C>G, c.931C>G) were identified in AKR1C4. All variants were in H-W equilibrium without structural changes. DISCUSSION: Hypospadias have been associated with defects that alter androgen biosynthesis in the human fetal testis, specifically 5α-DHT. We selected four candidate genes involved in the "backdoor" pathway for the formation of 5α-DHT. Molecular assays of the AKR1C2, AKR1C3, and AKR1C4 genes revealed a total of nine genetic single nucleotide variants. Several variants in the AKR1C genes have been associated with a variety of human pathologies. However, our studies suggest that active steroid biosynthesis via AKR1C might not be involved in hypospadias. Additionally, genetic research suggests a low involvement in the "backdoor" 5α-DHT pathway during human sexual development, specifically, the differentiation of male external genitalia. CONCLUSION: These results indicate that substitutions in AKR1C2-4 are polymorphisms and all genetic variants lacks deleterious significant association with hypospadias. The data suggest that inactivating mutations in the AKR1C2-4 and HSD17B6 genes are an infrequent cause of hypospadias, which might weaken the contribution of the "backdoor" pathway to embryonic urethral masculinization.

Hamster SRD5A3 lacks steroid 5α-reductase activity in vitro.

According to current knowledge, two steroid 5α-reductases, designated type 1 (SRD5A1) and type 2 (SRD5A2), are present in all species examined to date. These isozymes play a central role in steroid hormone physiology by catalyzing the reduction of 3-keto-4-ene-steroids into more active 5α-reduced derivatives, including the conversion of testosterone (T) to dihydrotestosterone (DHT). A third 5α-reductase (SRD5A3, -type 3), which is overexpressed in hormone-refractory prostate cancer cells, has been identified; however, its enzymatic characteristics are practically unknown. Here, we isolated a cDNA encoding hamster Srd5a3 (hSrd5a3) and performed functional metabolic assays to investigate its biochemical properties. The cloned cDNA encodes a 330 amino acid protein that is 87% identical to the homologous protein in mice and 78% to that in humans. However, hSrd5a3 exhibits low sequence homology with its counterparts hSrd5a1 (19%) and hSrd5a2 (17%). A fusion protein consisting of hSrd5a3 and green fluorescent protein provided evidence for cytoplasmic localization in transfected mammalian cells. Real-time PCR analysis revealed that, Srd5a3 mRNA was present in nearly all hamster tissues, with high expression in the cerebellum, Harderian gland and testis. Functional assays expressing hSrd5a3 cDNA in HEK-293 cells revealed that this isozyme is unable to reduce T into DHT. Further expression assays confirmed that similar to testosterone, progesterone, androstenedione and corticosterone are not reduced by hSrd5a3 or human SRD5A3. Together, these results indicate that hSrd5a3 lacks the catalytic activity to transform 3-keto-4-ene-compounds; therefore 5α-reductase type 3 may not be involved in 5α-reduction of steroids.

Effects of 5α-dihydrotestosterone on expression of genes related to steroidogenesis and spermatogenesis during the sex determination and differentiation periods of the pejerrey, Odontesthes bonariensis.

Sex steroid hormones are important players in the control of sex differentiation by regulating gonadal development in teleosts. Although estrogens are clearly associated with the ovarian differentiation in teleosts, the effects of androgens on early gonadal development are still a matter of debate. Traditionally, 11-ketotestosterone (11-KT) is considered the major androgen in fish; however, 5α-dihydrotestosterone (5α-DHT), the most potent androgen in tetrapods, was recently found in fish testis and plasma, but its physiological role is still unknown. In this context, the expression of genes associated with steroidogenesis and spermatogenesis, body growth and sex differentiation were assessed in Odontesthes bonariensis larvae fed with food supplemented with two doses of 5α-DHT (0.1 and 10µg/g of food) from hatching to 6weeks of age. At the lowest dose, 5α-DHT treated larvae showed an estrogenic gene expression pattern, with low hsd11b2 and high cyp19a1a and er2 expression levels with no differences in sex ratio. At the highest dose, 5α-DHT produced a male-shifted sex ratio and the larvae exhibited a gene expression profile characteristic of an advancement of spermatogenesis, with inhibition of amh and stimulation of ndrg3. No differences were observed in somatic growth. These results suggest that in this species, 5α-DHT could have a role on sex differentiation and its effects can differ according to the dose.

Hormonal changes and increased anxiety-like behavior in a perimenopause-animal model induced by 4-vinylcyclohexene diepoxide (VCD) in female rats.

Perimenopause, a transition period that precedes menopause, is characterized by neuroendocrine, metabolic and behavioral changes, and is associated with increased vulnerability to affective disorders. The decrease in ovarian follicles during perimenopause contributes to a dynamic and complex hormonal milieu that is not yet well characterized. In rodents, 4-vinylcyclohexene diepoxide (VCD) induces a gradual depletion of ovarian follicles, modeling the transition to menopause in women. This study was aimed to investigate, in VCD-treated rats, the hormonal status and the behavior in the elevated plus-maze (EPM), a widely used test to assess anxiety-like behavior. From the postnatal day 28, rats were treated with VCD or vehicle for 15 days. At 80±5 days after the beginning of treatment the experiments were performed at proestrus and diestrus. In the first experiment rats were decapitated, ovary was collected and blood samples were taken for estradiol, progesterone, follicle stimulant hormone (FSH), testosterone, dihydrotestosterone (DHT) and corticosterone measurements. In the second experiment, rats were subjected to the EPM for 5 min, and behavioral categories recorded. Administration of VCD induced follicular depletion as well as an increase of the number of atretic follicles demonstrating the treatment efficacy. The transitional follicular depletion was accompanied by lower progesterone, testosterone and DHT with no changes in the FSH, estradiol and corticosterone plasma levels. On the EPM, rats showed decreased open arm exploration and increased risk assessment behavior, indicating increased anxiety. These findings show that administration of VCD to induce ovarian failure results in endocrine and anxiety-related changes that are similar to the symptoms exhibited by women during menopause transition. Thus, this model seems to be promising in the study of perimenopause-related changes.

Hormonal correlates of male life history stages in wild white-faced capuchin monkeys (Cebus capucinus).

Much attention has been paid to hormonal variation in relation to male dominance status and reproductive seasonality, but we know relatively little about how hormones vary across life history stages. Here we examine fecal testosterone (fT), dihydrotestosterone (fDHT), and glucocorticoid (fGC) profiles across male life history stages in wild white-faced capuchins (Cebus capucinus). Study subjects included 37 males residing in three habituated social groups in the Área de Conservacíon Guanacaste, Costa Rica. Male life history stages included infant (0 to <12months; N=3), early juvenile (1 to <3years; N=10), late juvenile (3 to <6years; N=9), subadult (6 to <10years; N=8), subordinate adult (⩾10years; N=3), and alpha adult (⩾10years; N=4, including one recently deposed alpha). Life history stage was a significant predictor of fT; levels were low throughout the infant and juvenile phases, doubled in subadult and subordinate adults, and were highest for alpha males. Life history stage was not a significant predictor of fDHT, fDHT:fT, or fGC levels. Puberty in white-faced capuchins appears to begin in earnest during the subadult male phase, indicated by the first significant rise in fT. Given their high fT levels and exaggerated secondary sexual characteristics, we argue that alpha adult males represent a distinctive life history stage not experienced by all male capuchins. This study is the first to physiologically validate observable male life history stages using patterns of hormone excretion in wild Neotropical primates, with evidence for a strong association between fT levels and life history stage.

The antidepressant-like effect of ethynyl estradiol is mediated by both serotonergic and noradrenergic systems in the forced swimming test.

17α-Ethynyl-estradiol (EE2, a synthetic steroidal estrogen) induces antidepressant-like effects in the forced swimming test (FST) similar to those induced by 5-HT and noradrenaline reuptake inhibitors (dual antidepressants). However, the precise mechanism of action of EE2 has not been studied. In the present study, the participation of estrogen receptors (ERs) and the serotonergic and the noradrenergic presynaptic sites in the antidepressant-like action of EE2 was evaluated in the FST. The effects of the ER antagonist ICI 182,780 (10 µg/rat; i.c.v.), the serotonergic and noradrenergic terminal destruction with 5,7-dihydroxytryptamine (5,7-DHT; 200 µg/rat, i.c.v.), and N-(2-chloro-ethyl)-N-ethyl-2-bromobenzylamine (DSP4; 10mg/kg, i.p.) were studied in ovariectomized rats treated with EE2 and subjected to the FST. In addition, the participation of α2-adrenergic receptors in the antidepressant-like action of EE2 was explored using the selective α2-receptor antagonist idazoxan (0.25, 0.5 and 1.0mg/kg, i.p.). EE2 induced an antidepressant-like action characterized by a decrease in immobility behavior with a concomitant increase in swimming and climbing behaviors. The ER antagonist, 5,7-DHT, DSP4, and idazoxan blocked the effects of EE2 on the immobility behavior, whereas ICI 182,780 and 5,7-DHT affected swimming behavior. The noradrenergic compound DSP4 altered climbing behavior, while Idazoxan inhibited the increase of swimming and climbing behaviors induced by EE2. Our results suggest that the antidepressant-like action of EE2 implies a complex mechanism of action on monoaminergic systems and estrogen receptors.