Adipocytokines may delay pubertal maturation of human Sertoli cells.

Reproduction is an important target of obesity complications, including adverse effects on spermatogenesis and steroidogenesis. Adipocytokines are key mediators in various complications of obesity. Our aim was to study the potential of adipocytokines to affect Sertoli cell function, which is crucial for spermatogenesis, and possibly link these findings to the observed attenuation of spermatogenesis in obese males. Testicular biopsies were obtained from healthy donors. Highly purified adult human Sertoli cells (HSCs) were isolated by fluorescence-activated cell sorting. Cells were cultured and exposed to different concentrations of adipocytokines (10-1000ngmL-1 ) for 2-7 days. Expression of selected Sertoli cell genes was quantified by quantitative polymerase chain reaction. Long-term treatment (7 days) of HSCs with higher concentrations of chemerin, irisin, nicotinamide phosphoribosyltransferase (Nampt), resistin and progranulin significantly suppressed FSH receptor expression (by 79%, 83%, 64%, 71% and 26% respectively; P P invitro , may negatively affect Sertoli cell maturation and retain these cells in a more prepubertal stage. This could negatively affect testis function and add to fertility problems in obese adults.

Insulin-Like Peptide 5 Interacts with Sex Hormones and Metabolic Parameters in a Gender and Adiposity Dependent Manner in Humans.

Insulin-like peptide 5 (INSL5) is a gut hormone produced by L-cells in the colorectal epithelium and may play a role in the regulation of metabolic processes. The biological role of INSL5 is poorly investigated and nothing is known about the role of this hormone in obese and lean humans. Two cohorts were analyzed in the study. In the first cohort (n=76) the relationship between serum levels of INSL5 and different metabolic and hormonal parameters in obese and lean men and women were investigated. In the second cohort 14 male subjects underwent bariatric surgery. Circulating levels of INSL5 were then measured before and after interventions.We report for the first time that circulating INSL5 interacts with multiple metabolic and hormonal variables in lean and obese men and women and is affected by bariatric surgery. Serum levels of INSL5 negatively correlated with testosterone and blood lipids but positively with cortisol in obese men. In contrast to males, obese women had a strong negative correlation of plasma levels of INSL5 with C-reactive protein (CRP). We observed that adipose tissue loss after bariatric surgery significantly reduced serum levels of INSL5 in obese men with and without Type 2 Diabetes Mellitus (T2DM) that was associated with the restoration of circulating levels of testosterone. All together, our data demonstrated that INSL5 may interact with some metabolic parameters in obese humans and this process is dependent of gender and obesity state.

Endocrine disruptors and Leydig cell function.

During the past decades, a large body of information concerning the effects of endocrine disrupting compounds (EDCs) on animals and humans has been accumulated. EDCs are of synthetic or natural origin and certain groups are known to disrupt the action of androgens and to impair the development of the male reproductive tract and external genitalia. The present overview describes the effects of the different classes of EDCs, such as pesticides, phthalates, dioxins, and phytoestrogens, including newly synthesized resveratrol analogs on steroidogenesis in Leydig cells. The potential impact of these compounds on androgen production by Leydig cells during fetal development and in the adult age is discussed. In addition, the possible role of EDCs in connection with the increasing frequency of abnormalities in reproductive development in animals and humans is discussed.

Steroidogenic potential of human fetal kidney at early gestational age.

Steroidogenic potential of the human fetal kidney (hFK) at the end of first trimester is poorly investigated. Little is known about the ontogeny of steroidogenic enzymes and activities of steroidogenic pathways in the hFK at early pregnancy. Our aim was to explore steroidogenesis and the expression of steroidogenic enzymes in the hFK at gestational weeks (GW) 9-12. Steroids in the hFK were analyzed by gas chromatography/coupled to tandem mass spectrometry. The expression of steroidogenic enzymes in the hFK at GW 9-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We observed that the hFK produced substantial amount of steroids of the Δ5 and Δ4 pathways and several steroid precursors in the biosynthesis of DHT via the backdoor pathway but not DHT itself. The levels of steroids and expression of relevant steroidogenic enzymes (e.g., CYP17A1, HSD3B1, HSD3B2, CYP11B1 and AKR1C4) we significantly higher in the hFK at GW11-12 compared to GW9. We also found the expression of sex steroid receptors (e.g., AR, ERα and ERß) in the hFK at GW9-12. No sex-dependent differences in the levels of all identified steroids and expression of steroidogenic enzymes in the hFK from male and female fetuses were found. Altogether, our data indicate that the hFK at early pregnancy is steroidogenic organ with potential to synthesize multiple steroids that may play an important role in the formation and development of this organ in humans.

Ontogenesis of human fetal testicular steroidogenesis at early gestational age.

The onset of steroidogenesis in human fetal testes (HFT) during the first trimester is poorly investigated. One important unresolved question is the ontogeny of steroidogenic enzymes and formation of steroidogenic pathways in the HFT at early pregnancy. Our aim was to explore steroidogenesis, the expression of steroidogenic enzymes and their maturation in the HFT at gestational weeks (GW) 8-12. Steroids in the HFT were analyzed by gas chromatography/coupled to tandem mass spectrometry. The expression of steroidogenic enzymes in the HFT at GW8-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We demonstrated that the HFT at GW8-9 produced low level of testosterone via the Δ4 pathway and progesterone was the major steroid found in the testicular tissue. In contrast, more mature Leydig cells from the HFT at GW11-12 synthesized high levels of androgens via the Δ5 pathway. We also observed a significant upregulation of the expression of StAR, CYP11A1, CYP17A1 and its accessory proteins, P450 oxidoreductase (POR) and cytochrome b5 in the HFT at GW11-12 compared to GW8-9. Altogether, our data suggest that that human fetal Leydig cells differentiate rapidly at the end of the first trimester by acquiring capacity to express high levels of steroidogenic enzymes and switch from the Δ4 to the Δ5 pathways to synthesize high levels of androgens due to maturation of the CYP17-POR-b5 complex.

The human genital tubercle is steroidogenic organ at early pregnancy.

It is generally accepted that androgens produced by fetal Leydig cells (FLC) control proper masculinization of the male external genitalia. Here, we hypothesized that the human genital tubercle (GT) has potential to synthesize androgens independently of FLC at early pregnancy. We observed that human GT of both genders have capacity to synthesize steroids of the Δ4, Δ5 and alternative pathway of DHT synthesis including the androgen itself. The presence of steroids in the GT was associated with the expression of corresponding steroidogenic enzymes. Levels of steroids and the expression of steroidogenic enzymes were similar in the GT from male and female fetuses. In contrast to the GT, the human fetal testis synthesized DHT from testosterone but not via the alternative pathway. Our findings strongly suggest that the human GT at early pregnancy can synthesize DHT via the alternative pathway, which may play an important role in organogenesis of the urethra.

The influence of di-(2-ethylhexyl) phthalate on steroidogenesis by the ovarian granulosa cells of immature female rats.

Phthalate esters are known to exert harmful effects on mammalian reproduction and fertility, but their potential adverse effects on the hormonal functions of the ovary have not yet been elucidated in detail. Here, we investigated the effects of di-(2-ethylhexyl) phthalate (DEHP) on the hypothalamic-pituitary-gonadal axis of young developing female rats, as well as on ex vivo steroidogenesis by granulosa cells (GCs) and secretion of LH by gonadotropes. Exposure of 20-day-old female rats to 500 mg DEHP by oral gavage once daily for 10 days reduced their serum levels of progesterone and estradiol, while tending to enhance levels of LH. Furthermore, primary cultures of GCs isolated from these rats exhibited an attenuated capacity to produce progesterone in response to stimulation by LH and FSH, as well as a lower degree of transport of endogenous cholesterol into mitochondria. Moreover, the ability of primary cultures of pituitary cells isolated from DEHP-treated rats to produce and secrete LH in response to GnRH was significantly enhanced. In addition, 2-ethylhexanoic acid, a metabolite of DEHP, significantly potentiated GnRH-stimulated production of LH by cultures of pituitary cells isolated from untreated 20-day-old female rats. Together, these data indicate that DEHP exerts dual effects on the pituitary-gonadal axis, stimulating the hormonal function of the pituitary and, at the same time, by inhibiting steroidogenesis by GCs.

Androgenic potential of human fetal adrenals at the end of the first trimester.

The onset of steroidogenesis in human fetal adrenal glands (HFA) during the first trimester is poorly investigated. An unresolved question is the capacity of the HFA to produce potent androgen DHT via conventional and/or the backdoor pathway(s) at the end of first trimester, when androgen-responsive organs are developed. Our aim was to explore steroidogenesis and the expression of steroidogenic enzymes and transcription factors in HFA at gestational weeks (GW) 9-12 with focus on their androgenic potential. Steroids in the HFA were analyzed by gas chromatography/mass spectrometry. The expression of steroidogenic enzymes and transcription factors in the HFA at GW9-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We demonstrated that during GW9-12 HFA produced steroids of the ∆5, ∆4 and the backdoor pathways of the biosynthesis of DHT, though the latter was limited to production of 17α-OH-dihydroprogesterone, androsterone and androstanedione without further conversion to DHT. The only androgens identified in the HFA were testosterone and androsterone, a precursor in the biosynthesis of DHT. We also observed higher levels of CYP17A1 but low expression of 3ßHSD2 at GW11-12 in the HFA. Elevated levels of CYP17A1 were associated with an increased expression of SF-1 and GATA-6. Altogether, our data demonstrate that of those steroids analyzed, the only potent androgen directly produced by the HFA at GW9-12 was testosterone. The onset of steroidogenesis in the HFA is a complex process that is regulated by the coordinated action of related transcription factors.

Induction of steroidogenesis in immature rat Leydig cells by interleukin-1alpha is dependent on extracellular signal-regulated kinases.

Interleukin-1alpha (IL-1alpha) plays an important role in the regulation of immune responses as well as in non-inflammatory events in different types of cells. Here we have investigated the involvement of the extracellular signal-regulated kinase (ERK) cascade in IL-1alpha-induced steroidogenesis by primary cultures of immature rat Leydig cells. Our findings indicate that protein kinase C functions as an upstream component of signal transduction from the IL-1 receptor type I (IL-1RI) to the ERK cascade. It was observed that IL-1alpha upregulated both steroidogenic acute regulatory (StAR) protein expression and its phosphorylation when compared with controls. Selective inhibition of these mitogen-activated protein kinases (MAPKs) by UO126 enhanced both the expression and phosphorylation of the StAR protein, but suppressed androgen production by the immature Leydig cells as well as dissipating the mitochondrial electrochemical potential (Psim) in these cells. The evidence that water-soluble cholesterol but not 22R-hydroxycholesterol-stimulated steroidogenesis was inhibited by UO126 suggested that an intact Psim across the inner mitochondrial membrane is required for cholesterol translocation and is positively regulated by the ERK cascade. We propose that activation of ERKs by IL-1alpha plays a dual role in the regulation of steroidogenesis in immature Leydig cells: these MAPKs downregulate StAR expression and phosphorylation, while at the same time they support an intact Psim across the inner mitochondrial membrane, thereby promoting translocation of cholesterol into the mitochondria of the Leydig cell.

Amifostine protects against early but not late toxic effects of doxorubicin in infant rats.

The improved prognosis and increased expected lifetime among long-term survivors of childhood malignancies have made these patients especially sensitiveto the late toxicity of cancer therapy and prone to secondary malignancies. Recently, new strategies aiming to protect against cancer treatment toxicity have been developed, including the drug amifostine (Ethyol), which is suggested to protect normal tissues from the toxic effects of radiation and cytotoxic agents. In the present study, the possible protective effect of amifostine against toxicity induced by a single injection of doxorubicin (3 mg/kg) in immature rats was evaluated. Specifically, we evaluated the protection against long-term toxicity and the effects of amifostine on growing immature tissues. Amifostine (50-200 mg/kg) given 15 min before doxorubicin had a significant protective effect against doxorubicin-induced early alopecia in young rats. Significant protection against cataract formation was obtained by the use of low-dose amifostine (50 mg/kg). However, amifostine did not protect young rats against the late toxic effect of doxoubicin on linear growth, body weight, plasma leptin levels, and heart or testicular tissue. Worrisome, and in contrast to earlier studies in adult rats, an increased doxorubicin toxicity actually was observed and mortality was increased when the higher doses of amifostine (100-200 mg/kg) were used. The present results suggest that more data from growing immature animal models are needed to analyze the safety of amifostine treatment and its mechanisms of action before wider clinical use of this drug in pediatric cancer patients is recommended.

Prepubertal onset of obesity negatively impacts on testicular steroidogenesis in rats.

Obesity is a global health problem and impacts negatively on levels of testosterone and quality of sperm production. At present little is known about mechanisms that attenuate testicular function in obese males. Our study characterized testicular steroidogenesis and explored levels of relevant paracrine and hormonal factors in rats with short- and long-term obesity. We have found that obesity state increased serum levels of estradiol and leptin in both groups of obese rats and inhibited the expression of StAR and Cyp11a1 associated with low levels of intratesticular testosterone in rats with long-term obesity. Further, long-term obesity reduced the number of Leydig cells, increased the testicular levels of the proinflammatory adipocytokine TNFα and the number of testicular macrophages. All together, our data indicate that long-term obesity may cause chronic inflammation in the testis and negatively impacts on Leydig cell steroidogenesis.

Influence of long-term dietary administration of procymidone, a fungicide with anti-androgenic effects, or the phytoestrogen genistein to rats on the pituitary-gonadal axis and Leydig cell steroidogenesis.

Procymidone is a fungicide with anti-androgenic properties, widely used to protect fruits from fungal infection. Thereby it contaminates fruit products prepared for human consumption. Genistein-containing soy products are increasingly used as food additives with health-promoting properties. Therefore we examined the effects of long-term dietary administration (3 months) of the anti-androgen procymidone (26.4 mg/animal per day) or the phytoestrogen genistein (21.1 mg/animal per day) to rats on the pituitary-gonadal axis in vivo, as well as on Leydig cell steroidogenesis and on spermatogenesis ex vivo. The procymidone-containing diet elevated serum levels of LH and testosterone and, furthermore, Leydig cells isolated from procymidone-treated animals displayed an enhanced capacity for producing testosterone in response to stimulation by hCG or dibutyryl cAMP, as well as elevated expression of steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage (P450 scc) and cytochrome P450 17alpha (P450c17). In contrast, the rate of DNA synthesis during stages VIII and IX of spermatogenesis in segments of seminiferous tubules isolated from genistein-treated rats was decreased without accompanying changes in the serum level of either LH or testosterone. Nonetheless, genistein did suppress the ex vivo steroidogenic response of Leydig cells to hCG or dibutyryl cAMP by down-regulating their expression of P450 scc. Considered together, our present findings demonstrate that long-term dietary administration of procymidone or genistein to rats exerts different effects on the pituitary-gonadal axis in vivo and on Leydig cell steroidogenesis ex vivo. Possibly as a result of disruption of hormonal feedback control due to its anti-androgenic action, procymidone activates this endocrine axis, thereby causing hyper-gonadotropic activation of testicular steroidogenesis. In contrast, genistein influences spermatogenesis and significantly inhibits Leydig cell steroidogenesis ex vivo without altering the serum level of either LH or testosterone.

Molecular cloning and expression of a functionally different alternative splice variant of prointerleukin-1alpha from the rat testis.

We report here the characterization of an alternative splice variant of prointerleukin-1alpha (proIL-1alpha), constitutively expressed by the normal adult rat testis. In addition to the classical 32K proIL-1alpha (32proIL-1alpha) messenger RNA, the testis produced a shorter variant encoding a putative protein of 24K (24proIL-1alpha). In situ hybridization demonstrated constitutive expression of the splice transcript in the seminiferous tubules. This alternative complementary DNA lacked the fifth exon, harboring the calpain cleavage site essential for generation of mature 17K IL-1alpha. This was verified by calpain treatment, producing the expected cleavage products of recombinant 32proIL-1alpha, but not of 24proIL-1alpha. Similarly, expression in COS-7 cells demonstrated processing of 32proIL-1alpha to the mature 17K form and secretion, whereas 24proIL-1alpha remained unprocessed. Both 32proIL-1alpha and 24proIL-1alpha showed a dose-dependent stimulatory effect in a thymocyte proliferation assay, although at lower potency than mature 17K IL-1alpha. In contrast, when tested on hCG-stimulated Leydig cells in vitro, a dose-dependent inhibition of testosterone production was obtained with mature 17K IL-1alpha and at a lower potency with 32proIL-1alpha, whereas 24proIL-1alpha was inactive. In conclusion, the three IL-1 bioactive proteins described here contribute to IL-1 protein heterogeneity and may serve as constitutive paracrine mediators in the testis.

Extracellular signal-regulated kinases are involved in the acute activation of steroidogenesis in immature rat Leydig cells by human chorionic gonadotropin.

We studied the involvement of the ERK cascade in human chorionic gonadotropin (hCG)-induced steroidogenesis by primary cultures of immature rat Leydig cells. Our findings indicate that protein kinase A and protein kinase C function as upstream kinases in connection with transduction of the signal from the gonadotropin receptor to the ERK cascade. These MAPKs enhance the stimulatory effects of hCG on the de novo synthesis of the steroidogenic acute regulatory protein and the activity of protein phosphatase 2A, which are associated with increased androgen production by the Leydig cell. Specific inhibition of ERK1/2 by Uo126 suppressed all of these cellular responses to hCG. In contrast, steroidogenesis from 22OHC (a cell-permeable form of cholesterol) is not inhibited by Uo126, suggesting that cholesterol delivery to mitochondria is being affected by this compound. We propose that the ERK cascade is an important part of the signal transduction pathway involved in the rapid hormonal responses of Leydig cells to trophic hormones. In hCG-activated Leydig cells, these MAPKs may play a role in controlling the biosynthesis of the steroidogenic acute regulatory protein as well as regulating protein phosphatase 2A activity, thereby governing cholesterol transport across the mitochondrial membrane.

Interleukin-1alpha stimulates steroidogenic acute regulatory protein expression via p38 MAP kinase in immature rat Leydig cells.

We have investigated the involvement of the steroidogenic acute regulatory (StAR) protein in interleukin-1alpha (IL-1alpha)-induced steroidogenesis in immature (40-day-old) and adult Leydig cells in vitro. Further, IL-1alpha-mediated signaling pathway(s) controlling StAR expression in immature Leydig cells were also studied. IL-1alpha stimulated both androgen production and StAR protein expression in a dose- and time-dependent manner in immature but not adult Leydig cells. These effects of IL-1alpha were prevented by pretreatment of the cells with the specific inhibitors of the p38 MAP kinase, SB203580 and PD169316, suggesting that this kinase is an important part of IL-1alpha signaling in the immature Leydig cell. The present results suggest that IL-1alpha, which is constitutively produced by the rat testis from postnatal day 25, is an important paracrine regulator of postnatal Leydig cell maturation. Regulation of StAR protein expression is one of the possible mechanisms by which IL-1alpha contributes to the differentiation of immature Leydig cells into adult cells.

Age-dependent stimulation of Leydig cell steroidogenesis by interleukin-1 isoforms.

Different isoforms of testicular interleukin-1 (IL-1) were analysed to determine whether there were differences in the ability to modulate rat Leydig cell steroidogenesis in vitro. Rat 17K IL-1alpha and IL-1beta, 32K IL-1alpha precursor (32proIL-1alpha) and a 24K splice variant (24proIL-1alpha) stimulated testosterone production by Leydig cells from 40- but not 80-day-old rats. The potency of the isoforms was IL-1alpha>IL-1beta>32proIL-1alpha>24proIL-1alpha, IL-1alpha being 50-fold more potent than IL-1beta. IL-1 receptor antagonist reversed the effects and IL-1 receptor type I mRNA was expressed by the responding Leydig cells, indicating a receptor mediated action. Inhibition of PKA and Ca(2+) channels abolished IL-1-induced steroidogenesis, while inhibition of PKC had no significant effect. Except for 24proIL-1alpha which was stimulatory, all IL-1 isoforms suppressed hCG-driven testosterone production. This inhibitory effect was abolished by androstendione, suggesting that P450c17 was suppressed by IL-1. Our results indicate that IL-1 plays a paracrine role in the regulation of Leydig cell steroidogenesis.

The paracrine role played by interleukin-1 alpha in the testis.

Interleukin-1alpha (IL-1alpha) plays an important role(s) in the regulation of immune and inflammatory responses. The testis is an immunologically privileged organ and the variety of effects exerted by IL-1alpha on this organ have yet to be explored in detail. The aim of the present review is to describe our current view of the paracrine role played by IL-1alpha in testicular physiology. Testicular IL-1alpha is expressed during development, primarily in Sertoli cells, appearing in rats for the first time 20 days after birth. This cytokine is microheterogeneous, consisting of three molecular species with molecular weights of 45, 24 and 17 KDa. The 17 KDa form represents mature IL-1alpha, while the 24-KDa IL-1alpha has been shown by our research group to be an alternately spliced form of the 45-KDa pro-IL-1alpha. IL-1alpha was observed to stimulate the proliferation of immature Sertoli cells with higher efficacy than FSH. IL-1alpha was also found to exert mitogenic effects both on isolated peritubular cells and germ cells. Furthermore, isoforms of IL-1alpha were seen to stimulate basal testosterone production in immature Leydig cells, but not in the corresponding adult cells. This effect involved induction of the steroidogenic acute regulatory (StAR) protein and positively regulation by p38 MAPK. Recently, we have observed positive interactions between IL-1alpha and hormones of the GH/IGF-I system that lead to enhanced androgen production by the Leydig cell. In conclusion, our findings suggest that isoforms of IL-1alpha may serve as paracrine mediators, alone or in concert with other factors, that support proper testicular cell functioning and, thereby, reproduction and fertility.

Androgen and estrogen stimulation of ornithine decarboxylase activity in mouse kidney.

In the present work, the activity of mouse renal ornithine decarboxylase (ODC) from CBA female mice was used as a biological marker to detect (anti)androgenic activity of different groups of endocrine disruptors and steroids. Daily injections of testosterone or dihydrotestosterone (DHT) into 60 day old female mice for 4 days increased renal ODC activity in a dose-dependent manner that reached up to 100-fold (testosterone) or 250-fold (DHT) above the baseline when the highest dose, 200 microg/mouse, was used. Administration of flutamide concurrently with testosterone (75 microg/mouse) caused a potent decrease of ODC induction in a dose-dependent manner, suppressing the enzyme activity at the doses of 0.1 and 0.5 mg/mouse by about 88 and 95%, respectively. In contrast, estradiol at the doses of 0.5 and 1 mg/mouse induced a significant stimulation of renal ODC activity in a dose-dependent manner when it was given alone or in combination with testosterone. Using a sensitive increase in ODC activity in response to androgens as an end point, we did not detect an antiandrogenic effect of several antiandrogens, such as cyproterone acetate, spironolactone, p,p'DDE and vinclozolin. Also, none of these antiandrogens were able to change the basal level of renal ODC activity, with the exception of cyproterone acetate that at a dose of 0.1 mg/mouse stimulated ODC activity. The data obtained suggest that mouse renal ODC from CBA females is not strictly androgen-specific and cannot be used for estimation of antiandrogenic effects of compounds having an affinity to different types of receptors.

Molecular identity, expression and functional analysis of interleukin-1alpha and its isoforms in rat testis.

Interleukin-1alpha (IL-1alpha) is a proinflammatory cytokine that has also been found to act as a paracrine mediator involved in the regulation of testicular functions. The present review provides an overview of the role of IL-1alpha in testicular physiology. Bioactive IL-1alpha isolated from adult rat testis was found to consist of three distinct immunoreactive protein species with apparent sizes of 45, 24 and 19 kDa. These isoforms showed bioactivity in a thymocyte proliferation and steroidogenesis assays with different biopotencies. The background of the molecular heterogeneity and processing, secretion and regulation of the isoforms of testicular IL-1alpha are discussed. All three isoforms have been found to be secreted into the testis tubular lumen and interstitial space. We have provided evidence that IL-1alpha is a paracrine factor that may be of importance in, e.g., the regulation of Leydig cell steroidogenesis. Pathophysiologically, testicular IL-1alpha may contribute to testicular relapse of acute lymphocytic leukemia in boys.

[Genetic control of the activity of microsomal enzymes of steroidogenesis in Leydig cells of inbred strains of mice]. / Geneticheskii kontrol' aktivnostei mikrosomal'nykh fermentov steroidogeneza v kletkakh Leidiga inbrednykh linii myshei.

Four activities of key microsomal steroidogenic enzymes (3 beta-hydroxysteroid dehydrogenase-isomerase, microsomal cytochrome P45017 beta, and 17-ketosteroid reductase) were compared in Leydig cells of six inbred strains of mice: A/He, CBA/Lac, C57BL/6J, DD, YT, and PT. The activities of the enzymes were found to vary considerably from one strain to another, depending on the genotype. Analysis of testosterone biosynthesis revealed a decreased activity of 17-ketosteroid reductase as compared to the activities of other microsomal steroidogenic enzymes. The metabolism of pregnenolone and progesterone demonstrated features dependent on the genotype and caused by genotypic differences in the activities of microsomal steroidogenic enzymes. These activities demonstrated a correlative interstrain variation indicative of their coordinated genetic control. The correlative variation may be related to a major gene effect on the activities of microsomal steroidogenic enzymes.