Showing papers on "Steroid biosynthesis published in 1991"

Diazepam binding inhibitor and its processing products stimulate mitochondrial steroid biosynthesis via an interaction with mitochondrial benzodiazepine receptors.

Abstract: A recognition site for benzodiazepines structurally different from that linked to various γ-aminobutyric acid A (GABAA) receptor subtypes is located on the outer mitochondrial membranes of steroidogenic cells. This protein has been signified to be important in the regulation of steroid biosynthesis. Because of its location it is designated herein as the mitochondrial benzodiazepine receptor (MBR). A putative endogenous ligand for MBR is the peptide diazepam binding inhibitor (DBI), previously shown to displace drugs from MBR and to be expressed and stored in steroidogenic cells rich in MBR. The two model systems used to study steroidogenic regulation by DBI were the Y-l adrenocortical and MA-10 Leydig cell lines previously shown to be applicable in studies of mitochondrial steroidogenesis. Both cell lines contain DBI as well as DBI processing products, including the DBI fragments that on reverse phase HPLC coelute with the naturally occurring triakontatetraneuropeptide [TTN; DBI-(17–50)] and octadecaneuro...

Hormone-stimulated Steroidogenesis Is Coupled to Mitochondrial Benzodiazepine Receptors

Abstract: The mitochondrial (peripheral-type) benzodiazepine receptor (MBR) is a drug binding site associated with outer mitochondrial membranes which is coupled to intramitochondrial cholesterol transport, the rate-determining step of steroid biosynthesis. To examine the relationship between MBR function and steroid synthesis regulated by polypeptide hormones, the Y-1 adrenocortical and MA-10 Leydig cell lines were used as model systems responsive to adrenocorticotropin and human choriogonadotropin, respectively. Flunitrazepam, a benzodiazepine which binds to MBR with high nanomolar affinity, inhibited the steroidogenic activity of these hormones, or the activation by 1 mM dibutyryl CAMP, in both cell lines by 30-60% with an IC60 of 500-1000 nM. Scatchard analysis in both cell lines revealed one class of specific binding sites for [3H] flunitrazepam verified as being MBR by displacement studies with a series of MBR ligands. The potencies of these ligands to compete against the antagonism of hormone-stimulated steroidogenesis by flunitrazepam correlated significantly with their abilities to compete against [3H]flunitrazepam binding to MBR (r = 0.99). An inhibition in pregnenolone formation was also observed in isolated mitochondrial preparations characterized as a reduction of cholesterol transport to inner mitochondrial membranes. These observations provide unequivocal evidence that the antagonistic action of flunitrazepam is mediated through its interaction with MBR demonstrating that these drug recognition sites are coupled to steroid biosynthesis activated by tropic hormones.

Identification and purification of a human Sertoli cell-secreted protein (hSCSP-80) stimulating Leydig cell steroid biosynthesis.

Abstract: Human Sertoli cells in vitro secrete a factor that stimulates steroid biosynthesis in purified human and rat Leydig cells as well as in the MA-10 mouse tumor Leydig cell line. MA-10 cells were used as a bioassay system to follow the characterization and purification of the active principle in the conditioned medium of human Sertoli cells. The Leydig cell stimulatory factor is a thermo-labile and trypsin-sensitive protein retained onto 10,000 mol wt (MW) cut-off filters. The following scheme was used to purify the active protein: concentration by ammonium sulfate (80%) precipitation, followed by dialysis using molecularporous membrane tubing of MW cut-off 12,000-14,000, heparin-agarose, Concanavalin-A-Sepharose, and immobilized reactive textile dye affinity chromatography. Yellow 86 and green 19 dyes immobilized on agarose matrix were used. This procedure resulted in the rapid (less than 24-h) purification of a 79,000 +/- 6,082 (n = 3; under denaturating conditions) MW protein which stimulated Leydig cell steroid biosynthesis 25-fold at picomolar concentrations. The MW of the biologically active protein was further confirmed to be around 80,000 by gel filtration chromatography. This 80,000 MW human Sertoli cell-secreted protein (hSCSP-80) was shown to be different from human transferrin, human serum albumin, and rat testibumin. hSCSP-80, by modulating Leydig cell steroid biosynthesis, may play a significant role in the regulation of testicular function.

A renin-like enzyme in rat luteal tissue: evidence of local synthesis and its regulation during pregnancy.

Abstract: Changes in the concentration of a renin-like enzyme were studied in androgenized rats in which a single luteal phase was induced by the administration of chorionic gonadotrophin. A significant increase in the luteal renin-like enzyme (RLE) concentration was found between the youngest corpora lutea (48 h old) and the oldest one studied (6 days old). The luteal RLE content varied independently of changes in plasma renin concentration. These results suggest that this enzyme was produced locally. The lack of correlation between the luteal RLE and plasma prolactin supports our previous observation that the changes in luteal renin concentration appear not to be prolactin-dependent. Furthermore, the suckling-associated hormones appear not to be related with the regulation of luteal RLE concentration, since the values were not modified in androgenized maternal rats which were suckling when compared with the controls. Changes in luteal renin concentration were also studied during pregnancy. A significant increase was found a few hours after a fertile mating which reached a peak on day 1 of pregnancy, followed by a rapid decrease to low levels throughout the remainder of the pregnancy. Because the renin-angiotensin facilitates angiogenesis, luteal renin may act as an angiogenic factor, stimulating blood vessel growth in the corpora lutea. An alternative hypothesis is that the increase in RLE could be a trigger for calcium flux redistribution and steroid biosynthesis.

Optical probe for the cytochrome P-450 cholesterol side chain cleavage enzyme

Abstract: An optical probe enables the study of enzyme activity by absorbance spectroscopy or by sensitive fluorescence methods. In particular, the probe provides the ability to monitor the activity of cytochrome P-450 scc enzyme, the rate limiting enzyme for steroid biosynthesis. Located on the inner mitochondrial membrane, P-450 scc catalyzes the conversion of cholesterol to pregnenolone and isocapraldehyde by sequential oxidations of the cholesterol side chain. The fluorogenic probe includes a cholesterol-like steroid linked to a chromophore through a linking group. The chromophore is selected to have little optical response when linked to the steroid substrate and an enhanced optical response when cleaved from the substrate and linking group. Thus, a fluorescent anion that can be optically detected is generated by the side-chain cleavage reaction during steroidogenesis.

Agonist-induced desensitization of adenylyl cyclase in Y1 adrenocortical tumor cells.

Abstract: Y1 adrenocortical tumor cells (Y1DS) and Y1 mutants resistant to ACTH-induced desensitization of adenylyl cyclase (Y1DR) were transfected with a gene encoding the mouse beta 2-adrenergic receptor (beta 2-AR). Transfectants expressed beta 2-ARs that were able to stimulate adenylyl cyclase activity and steroid biosynthesis. These transfectants were used to explore the basis for the DR mutation in Y1 cells. We demonstrate that beta-adrenergic agonists desensitize the adenylyl cyclase system in transfected Y1DS cells whereas transfected Y1DR cells are resistant to desensitization by beta-adrenergic agonists. The fate of the beta 2-ARs during desensitization was evaluated by photoaffinity labelling with [125I]iodocyanopindolol diazerine. Desensitization of Y1DS transfectants was accompanied by a modest loss in receptor density that was insufficient to account for the complete loss of responsiveness to beta-adrenergic agonists. The extent of receptor loss induced by beta-adrenergic agonists in Y1DR transfectants exceeded that in the Y1DS transfectants indicating that the mutation which protects Y1DR cells from agonist-induced desensitization is prior to receptor down-regulation in the desensitization pathway. From these results we infer that ACTH and isoproterenol desensitize adenylyl cyclase by a common pathway and that receptor loss is not a major component of the desensitization process in these cells.

Reversal of female sexual phenotype in poultry

Abstract: JLT1aY 18024Y TITLE OF THE INVENTION REVERSAL OF FEMALE SEXUAL PHENOTYPE IN POULTRY Fertilized poultry embryos are treated with steroid biosynthesis inhibitors or antagonists which prevents the conversion of androgens to estrogens. By blocking the production of estrogens the genotypic female is converted into a phenotypic male. The phenotypic conversion of females to males gives the treated birds the advantage of male growth characteristics. A single administration of a steroid biosynthesis inhibitor or antagonist prior to about day 9 of embryonic incubation results in an irreversible change the sexual phenotype.

Animal growth promotion with aromatase inhibitors

Abstract: Female prenatal, neonatal and postnatal animals are treated with compositions containing steroid biosynthesis inhibitors or antagonists which prevents the conversion of androgens to estrogens. The compositions are useful for improving growth and feed efficiency.