Female fertility requires normal ovarian follicular growth and ovulation. The nuclear receptor liver receptor homolog 1 has been implicated in processes as diverse as bile acid metabolism, steroidogenesis, and cell proliferation. In the ovary, Lrh1 is expressed exclusively in granulosa and luteal cells. Using somatic targeted mutagenesis, we show that mice lacking Lrh1 in granulosa cells are sterile, due to anovulation. The preovulatory stimulus fails to elicit cumulus expansion, luteinization, and follicular rupture in these mice. Multiple defects, including severely reduced transactivation of the Lrh1 target gene, nitric oxide synthase 3, leads to increased intrafollicular estradiol levels in the absence of Lrh1. This further causes dysfunction of prostaglandin and hyaluronic acid cascades and interrupts cumulus expansion. Lack of Lrh1 also interferes with progesterone synthesis because of failure of normal expression of the Lrh1 targets, steroidogenic acute regulatory protein and cytochrome P450 side-chain cleavage. In addition, expression of extracellular matrix proteases essential for ovulation is compromised. These results demonstrate that Lrh1 is a regulator of multiple mechanisms essential for maturation of ovarian follicles and for ovulation. Lrh1 is therefore a key modulator of female fertility and a potential target for contraception. Much of mammalian female infertility can be attributed to dysfunction in ovarian folliculogenesis and ovulation. Both processes are tightly controlled by pituitary gonadotropins and locally produced factors, including steroid hormones and growth factors, which act in a coordinated fashion. In the ovary, the two main female hormones that drive these processes-i.e., estradiol-17␤ and progesterone-have overlapping but clearly distinct functions. Progesterone is required for successful ovulation, as deletion of progesterone receptor (Pgr) disrupts ovulation without affecting follicular growth or luteinization (Robker et al. 2000). Estradiol-17␤ is critical for both follicular growth and ovulation, as mice null for cytochrome P450 aromatase (Cyp19) (Fisher et al. 1998) have arrested follicular growth, while enhanced estradiol action has been linked to ovulatory defects (JablonkaShariff and Olson 1998;Gershon et al. 2007). Liver receptor homolog 1 (Lrh1, official gene name: Nr5a2), a member of the NR5A subfamily, is highly expressed in the granulosa cells of follicles and in the corpus luteum (CL) Zhao et al. 2007). In the present study, we explored the role of Lrh1 in ovarian follicular development using somatic targeted mutagenesis and show that Lrh1 is a critical regulator of multiple mechanisms essential for maturation of ovarian follicles and for ovulation. Results and Discussion Lrh1 gc−/− mice are sterileWe generated granulosa-specific mutants (Lrh1 (Fig. 1B). When subjected to a 6-mo breeding trial, both Lrh1 gc+/+ and Amhr2 Cre/+ control females proved fertile, with expected frequency of parturition and litter sizes, while no litters were born to Lrh1 gc−/− females ...

Cholesterol is the obligate precursor to adrenal steroids but is cytotoxic at high concentrations. Here, we show the role of the liver X receptors (LXRα and LXRβ) in preventing accumulation of free cholesterol in mouse adrenal glands by controlling expression of genes involved in all aspects of cholesterol utilization, including the steroidogenic acute regulatory protein, StAR, a novel LXR target. Under chronic dietary stress, adrenal glands from Lxrαβ −/− mice accumulated free cholesterol. In contrast, wild-type animals maintained cholesterol homeostasis through basal expression of genes involved in cholesterol efflux and storage (ABC transporter A1 [ABCA1], apoE, SREBP-1c) while preventing steroidogenic gene (StAR) expression. Upon treatment with an LXR agonist that mimics activation by oxysterols, expression of these target genes was increased. Basally, Lxrαβ -/-mice exhibited a marked decrease in ABCA1 and a derepression of StAR expression, causing a net decrease in cholesterol efflux and an increase in steroidogenesis. These changes occurred under conditions that prevented the acute stress response and resulted in a phenotype more specific to the loss of LXRα, including hypercorticosteronemia, cholesterol ester accumulation, and adrenomegaly. These results imply LXRα provides a safety valve to limit free cholesterol levels as a basal protective mechanism in the adrenal gland, where cholesterol is under constant flux. IntroductionThe adrenal cortex is responsible for synthesizing glucocorticoid hormones that are essential for survival under stress. This endocrine pathway is acutely regulated by the hypothalamicpituitary-adrenal axis in response to stress through the release of ACTH from the anterior pituitary. ACTH signals the adrenal gland to increase the expression of a cascade of enzymes required for the conversion of cholesterol into biologically active glucocorticoids. The initial and rate-limiting step in this cascade is mediated by the steroidogenic acute regulatory protein (StAR) that transfers cholesterol from the outer to the inner mitochondrial membrane (1, 2). Inside the mitochondria, cytochrome P450 11A1 (CYP11A1) cleaves the cholesterol side chain to form pregnenolone (3), which can be further converted by a series of enzymes (e.g., type I 3β-hydroxysteroid dehydrogenase/D 5 -D 4 -isomerase) to all steroid hormones produced by the adrenal cortex. Because the stress response is intended to be of limited duration, tight regulation of this system is maintained by the negative feedback of circulating glucocorticoids on the hypothalamus and pituitary that decreases ACTH secretion and ultimately turn off glucocorticoid production (4, 5).

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Oxysterol nuclear receptors liver X receptor (LXR)alpha and LXRbeta are known to regulate lipid homeostasis in cells exposed to high amounts of cholesterol and/or fatty acids. In order to elucidate the specific and redundant roles of the LXRs in the testis, we explored the reproductive phenotypes of mice deficient of LXRalpha, LXRbeta, and both, of which only the lxralpha;beta-/- mice are infertile by 5 months of age. We demonstrate that LXRalpha-deficient mice had lower levels of testicular testosterone that correlated with a higher apoptotic rate of the germ cells. LXRbeta-deficient mice showed increased lipid accumulation in the Sertoli cells and a lower proliferation rate of the germ cells. In lxralpha;beta-/- mice, fatty acid metabolism was affected through a decrease of srebp1c and increase in scd1 mRNA expression. The retinoid acid signaling pathway was also altered in lxralpha;beta-/- mice, with a higher accumulation of all-trans retinoid receptor alpha, all-trans retinoid receptor beta, and retinoic aldehyde dehydrogenase-2 mRNA. Combination of these alterations might explain the deleterious phenotype of infertility observed only in lxralpha;beta-/- mice, even though lipid homeostasis seemed to be first altered. Wild-type mice treated with a specific LXR agonist showed an increase of testosterone production involving both LXR isoforms. Altogether, these data identify new roles of each LXR, collaborating to maintain both integrity and functions of the testis.

The small heterodimer partner (SHP) is an atypical nuclear receptor known mainly for its role in bile acid homeostasis in the enterohepatic tract. We explore here the role of SHP in the testis. SHP is expressed in the interstitial compartment of the adult testes, which contain the Leydig cells. SHP there inhibits the expression of steroidogenic genes, on the one hand by inhibiting the expression of the nuclear receptors steroidogenic factor-1 and liver receptor homolog-1 (lrh-1), and on the other hand by directly repressing the transcriptional activity of LRH-1. Consequently, in SHP knockout mice, testicular testosterone synthesis is increased independently of the hypothalamus-pituitary axis. Independent of its action on androgen synthesis, SHP also determines the timing of germ cell differentiation by controlling testicular retinoic acid metabolism. Through the inhibition of the transcriptional activity of retinoic acid receptors, SHP controls the expression of stimulated by retinoic acid gene 8 (stra8)-a gene that is indispensable for germ cell meiosis and differentiation. Together, our data demonstrate new roles for SHP in testicular androgen and retinoic acid metabolism, making SHP a testicular gatekeeper of the timing of male sexual maturation.[Keywords: Farnesoid X receptor; germ cell differentiation; liver receptor homolog-1; retinoic acid receptor; steroidogenesis; testis] Supplemental material is available at http://www.genesdev.org.

Bile acids (BAs) are signaling molecules that are involved in many physiological functions, such as glucose and energy metabolism. These effects are mediated through activation of the nuclear and membrane receptors, farnesoid X receptor (FXR-a) and TGR5 (G-protein-coupled bile acid receptor 1; GPBAR1). Although both receptors are expressed within the testes, the potential effect of BAs on testis physiology and male fertility has not been explored thus far. Here, we demonstrate that mice fed a diet supplemented with cholic acid have reduced fertility subsequent to testicular defects. Initially, germ cell sloughing and rupture of the blood-testis barrier occur and are correlated with decreased protein accumulation of connexin-43 (Cx43) and N-cadherin, whereas at later stages, apoptosis of spermatids is observed. These abnormalities are associated with increased intratesticular BA levels in general and deoxycholic acid, a TGR5 agonist, in particular. We demonstrate here that Tgr5 is expressed within the germ cell lineage, where it represses Cx43 expression through regulation of the transcriptional repressor, T-box transcription factor 2 gene. Consistent with this finding, mice deficient for Tgr5 are protected against the deleterious testicular effects of BA exposure. Conclusions: These data identify the testis as a new target of BAs and emphasize TGR5 as a critical element in testicular pathophysiology. This work may open new perspectives on the potential effect of BAs on testis physiology during liver dysfunction. (HEPATOLOGY 2014;60:1054-1065 T he incidence of infertility is constantly increasing and affects up to 25% of couples. Approximately 50% of the cases comprise disorders of the male reproductive system. They have frequently been associated with an inadequate number and quality of male germ cells (e.g., spermatozoa). 1,2 Alterations of testicular physiology play an important role in reduced sperm number, leading to infertility. The major functions of the testes include synthesis of the male sex hormone (testosterone) and production of gametes. Spermatogenesis takes place within the seminiferous tubules in association with the Sertoli cells (SCs), which provide structural support for developing germ cells through cell-cell interactions involving proteins such as

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Studies in rodents have shown that male sexual function can be disrupted by fetal or neonatal administration of compounds that alter endocrine homeostasis, such as the synthetic nonsteroidal estrogen diethylstilbestrol (DES). Although the molecular basis for this effect remains unknown, estrogen receptors likely play a critical role in mediating DES-induced infertility. Recently, we showed that the orphan nuclear receptor small heterodimer partner (Nr0b2), which is both a target gene and a transcriptional repressor of estrogen receptors, controls testicular function by regulating germ cell entry into meiosis and testosterone synthesis. We therefore hypothesized that some of the harmful effects of DES on testes could be mediated through Nr0b2. Here, we present data demonstrating that Nr0b2 deficiency protected mice against the negative effects of DES on testis development and function. During postnatal development, Nr0b2-null mice were resistant to DES-mediated inhibition of germ cell differentiation, which may be the result of interference by Nr0b2 with retinoid signals that control meiosis. Adult Nr0b2-null male mice were also protected against the effects of DES; however, we suggest that this phenomenon was due to the removal of the repressive effects of Nr0b2 on steroidogenesis.Together, these data demonstrate that Nr0b2 plays a critical role in the pathophysiological changes induced by DES in the mouse testis.