Genomic cloning, expression, and single nucleotide polymorphism association analysis of the insulin-like androgenic gland hormone gene in the oriental river prawn (Macrobrachium nipponense).

Increasing evidence suggests that the insulin-like androgenic gland hormone (IAG) gene plays an important role in male sexual differentiation, metabolism, and growth in crustaceans. In the present study, we isolated the full-length genome sequence of IAG by genome walking based on the cDNA sequence in Macrobrachium nipponense. Four novel single nucleotide polymorphisms (SNPs) were studied, including 509G>T, 529G>T, 590A>T in intron 1, and 2226A>G in intron 2. The association of genetic variation with growth traits [body length (BL) and body weight (BW)] was analyzed. Individuals with GG geno- type at locus 2226A>G maintained higher mean BL (P < 0.01) and BW (P < 0.05) than AA and GA individuals. These results suggest that IAG SNPs may be useful molecular markers for selecting growth traits in M. nipponense.

Recent patents on new steroid agents targeting the steroidogenesis for endocrine cancer treatments.

Cancer is a leading cause of death in the population and despite the significant technological advances that have been made over the last years, there is a great need for new and better treatments with fewer side effects. Among the various types, hormone-dependent cancers are stimulated by the presence of certain steroidal hormones such as androgens and estrogens, which act through a nuclear receptor. The use of small molecules to block the biosynthesis (steroidogenesis) or the action of hormones (androgens or estrogens) is a therapeutic approach that has yielded interesting results and whose development continues. This review article emphasizes the patents and patent applications published over the last five years. It deals exclusively with steroid compounds developed as inhibitors of key enzymes (17α-hydroxylase/17,20-lyase, steroid sulfatase, 5α-reductases, aromatase and 17ß-hydroxysteroid dehydrogenases) involved in the steroidogenesis and identified as therapeutic targets. Such inhibitors could be used as a drug to reduce the concentration of androgens or estrogens and, consequently, for treating hormone-dependent diseases such as prostate cancer, breast cancer and endometriosis.

Ovarian hormone withdrawal in prepubertal developmental stage does not prevent thymic involution in rats.

The study was undertaken to assess the effects of ovarian hormone withdrawal in prepubertal age on thymopoiesis in 2- (young) and 11-month-old (middle-aged) rats. In ovariectomized (Ox) rats, irrespective of age, thymic weight and cellularity were greater than in age-matched controls, but the values of both parameters exhibited the age-related decline. In addition, although thymopoietic efficiency was increased in both groups of Ox rats when compared with age-matched controls, thymopoiesis exhibited the age-related decline mirrored in the lower numbers of both CD4+ and CD8+ recent thymic emigrants in peripheral blood. This reflected the prethymic changes affecting bone marrow progenitor generation/entry and the thymic alterations encompassing the impaired progenitor progression through early pre-T-cell receptor developmental stages (defined by CD45RC/CD2 expression) and, possibly, a more pronounced decrease in the proliferation of the most mature thymocytes. Apart from the changes at thymocyte level, in Ox rats the age-related alterations in thymic stroma (substantiated in a prominent loss of thymic epithelial cells) were registered. Ovariectomy-induced changes in thymic lymphoid and epithelial component, most probably, influenced each other leading to the increase in thymic expression of interleukin-6 and interleukin-7 mRNAs along with time after ovariectomy. Collectively, the study showed that the withdrawal of ovarian hormones in prepubertal age increases the efficiency of thymopoiesis in young adult rats, but does not prevent decline in thymopoiesis occurring with age.

Mitochondria and mammalian reproduction.

Mitochondria are cellular organelles with crucial roles in ATP synthesis, metabolic integration, reactive oxygen species (ROS) synthesis and management, the regulation of apoptosis (namely via the intrinsic pathway), among many others. Additionally, mitochondria in different organs or cell types may have distinct properties that can decisively influence functional analysis. In terms of the importance of mitochondria in mammalian reproduction, and although there are species-specific differences, these aspects involve both energetic considerations for gametogenesis and fertilization, control of apoptosis to ensure the proper production of viable gametes, and ROS signaling, as well as other emerging aspects. Crucially, mitochondria are the starting point for steroid hormone biosynthesis, given that the conversion of cholesterol to pregnenolone (a common precursor for all steroid hormones) takes place via the activity of the cytochrome P450 side-chain cleavage enzyme (P450scc) on the inner mitochondrial membrane. Furthermore, mitochondrial activity in reproduction has to be considered in accordance with the very distinct strategies for gamete production in the male and female. These include distinct gonad morpho-physiologies, different types of steroids that are more prevalent (testosterone, estrogens, progesterone), and, importantly, the very particular timings of gametogenesis. While spermatogenesis is complete and continuous since puberty, producing a seemingly inexhaustible pool of gametes in a fixed environment; oogenesis involves the episodic production of very few gametes in an environment that changes cyclically. These aspects have always to be taken into account when considering the roles of any common element in mammalian reproduction.

Molecular characterization and expression analysis of an insulin-like gene from the androgenic gland of the oriental river prawn, Macrobrachium nipponense.

The androgenic gland (AG), a male-specific endocrine organ in crustacean, is responsible for the maintenance of male characteristics and gender differentiation. In this study, an AG-specific gene, the Macrobrachium nipponesne insulin-like androgenic gland factor (MnIAG) was isolated from a transcriptome library of M. nipponesne and its full-length cDNA sequences were obtained by RACE method. The cDNA was 1,547 bp in length and encoded a precursor protein of 175 amino acids. The deduced precursor protein consisted of a signal peptide, B chain, C peptide and an A chain, which exhibited the same structural organization as that of previously identified insulin-like androgenic gland in crustacean. The mature peptide of the MnIAG owned two additional conserved cysteine residues, which were also found in the Palaemonidae species reported. Results of the tissue distribution and in situ hybridization showed the MnIAG expressed exclusively in androgenic gland. The quantitative RT-PCR results demonstrated that the MnIAG transcript was present at blastula stage and later developmental stages with low levels, which suggested that the primordial cells of the AG might form at these stages.

Bilateral eyestalk ablation of the blue swimmer crab, Portunus pelagicus, produces hypertrophy of the androgenic gland and an increase of cells producing insulin-like androgenic gland hormone.

The androgenic glands (AG) of male decapod crustaceans produce insulin-like androgenic gland (IAG) hormone that controls male sex differentiation, growth and behavior. Functions of the AG are inhibited by gonad-inhibiting hormone originating from X-organ-sinus gland complex in the eyestalk. The AG, and its interaction with the eyestalk, had not been studied in the blue swimmer crab, Portunus pelagicus, so we investigated the AG structure, and then changes of the AG and IAG-producing cells following eyestalk ablation. The AG of P. pelagicus is a small endrocrine organ ensheathed in a connective tissue and attached to the distal part of spermatic duct and ejaculatory bulb. The gland is composed of several lobules, each containing two major cell types. Type I cells are located near the periphery of each lobule, and distinguished as small globular cells of 5-7 µm in diameter, with nuclei containing mostly heterochromatin. Type II cells are 13-15 µm in diameter, with nuclei containing mostly euchromatin and prominent nucleoli. Both cell types were immunoreactive with anti-IAG. Following bilateral eyestalk ablation, the AG underwent hypertrophy, and at day 8 had increased approximately 3-fold in size. The percentage of type I cells had increased more than twice compared with controls, while type II cells showed a corresponding decrease.

Neurosteroids, immunosteroids, and the Balkanization of endocrinology.

Traditionally, the production and regulation of steroid hormones has been viewed as a multi-organ process involving the hypothalamic-pituitary-gonadal (HPG) axis for sex steroids and the hypothalamic-pituitary-adrenal (HPA) axis for glucocorticoids. However, active steroids can also be synthesized locally in target tissues, either from circulating inactive precursors or de novo from cholesterol. Here, we review recent work demonstrating local steroid synthesis, with an emphasis on steroids synthesized in the brain (neurosteroids) and steroids synthesized in the immune system (immunosteroids). Furthermore, recent evidence suggests that other components of the HPG axis (luteinizing hormone and gonadotropin-releasing hormone) and HPA axis (adrenocorticotropic hormone and corticotropin-releasing hormone) are expressed locally in target tissues, potentially providing a mechanism for local regulation of neurosteroid and immunosteroid synthesis. The balance between systemic and local steroid signals depends critically on life history stage, species adaptations, and the costs of systemic signals. During particular life history stages, there can be a shift from systemic to local steroid signals. We propose that the shift to local synthesis and regulation of steroids within target tissues represents a "Balkanization" of the endocrine system, whereby individual tissues and organs may become capable of autonomously synthesizing and modulating local steroid signals, perhaps independently of the HPG and HPA axes.

Effect of bisphenol A on steroid hormone production in rat ovarian theca-interstitial and granulosa cells.

Many studies have shown that 2,2-bis 4-hydroxyphenyl propane (BPA), an estrogenic chemical, affects the reproductive health of wildlife and possibly of humans. In this study, we investigated the effects of BPA on steroid hormone production in rat ovarian theca-interstitial cells (T-I cells) and granulosa cells. In T-I cells, BPA increased testosterone synthesis and mRNA expression of 17-alpha hydroxylase (P450c17), cholesterol side chain cleavage enzyme (P450scc) and steroidogenic acute regulatory protein (StAR) at concentrations of 10(-7) to 10(-4)M after a 72 h incubation period. Treatment of granulosa cells with BPA at concentrations of 10(-7) to 10(-5)M caused an increase in progesterone levels and P450scc mRNA expression, with an unexpected decrease at 10(-4)M. BPA (10(-7) to 10(-5)M) tended to elevate the expression of StAR mRNA with a significant increase at 10(-4)M concentration. A significant concentration-dependent inhibitory effect of BPA (10(-6) to 10(-4)M) on estradiol levels and the expression of P450arom mRNA was observed. These results suggest that BPA may interrupt ovarian steroidogenesis by altering the steroidogenic enzymes.

Cobalt chloride, a hypoxia-mimicking agent, modulates redox status and functional parameters of cultured swine granulosa cells.

Hypoxia occurs physiologically during ovarian follicle growth; this deprivation represents a triggering stimulus for the production of vascular endothelial growth factor (VEGF) by proliferating granulosa cells, which are mostly responsible for the growth of the follicle. Moreover, the steroidogenic activity of these cells ensures a receptive environment for the implantation and development of the early embryo. The present paper reports the adaptive response of swine granulosa cells to cobalt chloride (CoCl(2), a chemical hypoxia-mimicking agent. The effects of the treatment were evaluated on cell proliferation, steroidogenesis and VEGF production. In addition, because mithocondrial reactive oxygen species (ROS) are possibly involved in O(2) sensing, ROS levels and scavenging enzyme activity were investigated. In the present study, CoCl 2 had no effect on progesterone production, although it significantly reduced oestradiol synthesis. The addition of CoCl(2) to granulosa cell culture stimulated VEGF production and the generation of hydrogen peroxide. Chemical hypoxia had different effects on scavenger enzyme activities: the activity of superoxide dismutase was enhanced, that of peroxidase reduced and catalase activity was unaffected. The net result is a 'pro-oxidant' state, which appears to be possibly involved in the stimulation of VEGF production, thus inducing follicular angiogenesis.