Biomedical subjects

M Culty

Publications and source records attributed to M Culty.

24 records · Page 2Linked to original sources

PubMed1984

Stimulation of phosphatidylinositol turnover by acetylcholine, angiotensin II and ACTH in bovine adrenal fasciculata cells.

The effect of acetylcholine, angiotensin II and adrenocorticotropin (ACTH) on phosphatidylinositol (PI) metabolism was examined using bovine adrenocortical fasciculata cell suspensions. The three agents, which acutely stimulate glucocorticoid production by these cells, were all able to increase [32P]Pi incorporation into cellular PI. However, whereas the relative steroidogenic potency (at maximally active concentrations) was ACTH greater than or equal to angiotensin II greater than acetylcholine, the effect on PI labeling was in the order angiotensin II greater than acetylcholine greater than ACTH. The dose-response curves for steroidogenesis and that for PI labeling were superimposable in the case of angiotensin II (ED50 = 1 X 10(-8) M) and of acetylcholine (ED50 = 5 X 10(-7) M), while the two responses were dissociated under graded ACTH challenge. Both steroidogenic response and increased PI labeling elicited by angiotensin II and acetylcholine were respectively inhibited by (Sar1-Ala8)-angiotensin II and muscarinic antagonists. Time-course study showed that in the case of angiotensin II and acetylcholine, the sequence of events was: increased phosphatidic acid labeling, increased PI labeling, activated steroidogenesis. By sharp contrast, under ACTH stimulation, increased steroidogenesis was detected well before activation of PI metabolism. These data suggest that in bovine adrenocortical fasciculata cell, steroidogenesis may be activated by two different pathways. The first one would act mainly through cyclic AMP-dependent intracellular events and is usually accepted in the mechanism of action of ACTH. The other, cyclic AMP-independent pathway, as in the case of angiotensin II and acetylcholine actions, may involve phospholipid-mediated intracellular processes.

Acetylcholine↗

PubMed1984

Steroidogenic properties of phorbol ester and a Ca2+ ionophore in bovine adrenocortical cell suspensions.

When added independently to bovine adrenocortical fasciculata cell suspensions, 12 tetradecanoyl-phorbol-13 acetate (TPA) and the Ca2+ ionophore A23187 activated net cortisol production in a time and dose-dependent manner during one hour incubation. When added together (each at 1 microM concentration), the drugs appeared synergistic and mimicked the steroidogenic effect of suboptimal concentration of angiotensin II or acetylcholine on these cells, with no detectable variation of cellular cyclic nucleotide levels. In addition, the drug mixture markedly enhanced the steroidogenic effect of acetylcholine. These observations suggest that Ca2+-activated, phospholipid dependent protein kinase, which is present in adrenal cortex, might be considered as a possible target in the mechanism of action of steroidogenic agents such as angiotensin and acetylcholine, acting in adrenocortical cell through cyclic AMP independent processes.

Adrenal Cortex↗

PubMedDate not supplied

In vitro studies on the role of the peripheral-type benzodiazepine receptor in steroidogenesis.

In vitro studies using isolated cells, mitochondria and submitochondrial fractions demonstrated that in steroid synthesizing cells, the peripheral-type benzodiazepine receptor (PBR) is an outer mitochondrial membrane protein, preferentially located in the outer/inner membrane contact sites, involved in the regulation of cholesterol transport from the outer to the inner mitochondrial membrane, the rate-determining step in steroid biosynthesis. Mitochondrial PBR ligand binding characteristics and topography are sensitive to hormone treatment suggesting a role of PBR in the regulation of hormone-mediated steroidogenesis. Targeted disruption of the PBR gene in Leydig cells in vitro resulted in the arrest of cholesterol transport into mitochondria and steroid formation; transfection of the mutant cells with a PBR cDNA rescued steroidogenesis demonstrating an obligatory role for PBR in cholesterol transport. Molecular modeling of PBR suggested that it might function as a channel for cholesterol. This hypothesis was tested in a bacterial system devoid of PBR and cholesterol. Cholesterol uptake and transport by these cells was induced upon PBR expression. Amino acid deletion followed by site-directed mutagenesis studies and expression of mutant PBRs demonstrated the presence in the cytoplasmic carboxy-terminus of the receptor of a cholesterol recognition/interaction amino acid consensus sequence. This amino acid sequence may help for recruiting the cholesterol coming from intracellular sites to the mitochondria.

Animals↗

PubMedDate not supplied

In vivo studies on the role of the peripheral benzodiazepine receptor (PBR) in steroidogenesis.

In various steroidogenic cell models, mitochondrial preparations and submitochondrial fractions, the expression of the mitochondrial 18 kDa peripheral-type benzodiazepine receptor (PBR) protein confers the ability to take up and release, upon ligand activation, cholesterol. Thus, cholesterol becomes available to P450scc on the inner mitochondrial membrane. These in vitro studies were validated by in vivo experiments. Treatment of rats with ginkgolide B (GKB), specifically reduced the ligand binding capacity, protein, and mRNA expression of the adrenocortical PBR and circulating glucocorticoid levels. Treatment with GKB also resulted in inhibition of PBR protein synthesis and corticosterone production by isolated adrenocortical cells in response to ACTH. The ontogeny of both PBR binding capacity and protein directly paralleled that of ACTH-inducible steroidogenesis in rat adrenal cells and in rats injected with ACTH. In addition, the previously described suppression of luteal progesterone synthesis in the pregnant rat by continuous in vivo administration of a gonadotropin-releasing hormone agonist may be due to decreased luteal PBR ligand binding and mRNA. These results suggest that (i) PBR is an absolute prerequisite for adrenocortical and luteal steroidogenesis, (ii) regulation of adrenal PBR expression may be used as a tool to control circulating glucocorticoid levels and (iii) the stress hypo-responsive period of neonatal rats may result from decreased adrenal cortical PBR expression.

Adrenal Cortex Hormones↗

PubMedDate not supplied

Drug-induced inhibition of the peripheral-type benzodiazepine receptor expression and cell proliferation in human breast cancer cells.

The peripheral-type benzodiazepine receptor (PBR) expression and localization correlate with human breast cancer cell proliferation and aggressive phenotype expression. The standardized extract of Ginkgo biloba leaves (EGb 761) and isolated ginkgolide B (GKB) were shown to decrease PBR mRNA expression in adrenal cells. We examined the effect of EGb 761 and GKB on PBR expression and cell proliferation in human breast cancer cells. EGb 761 and GKB decreased in a time- and dose-dependent manner PBR expression and cell proliferation in the highly aggressive, rich in PBR, human breast cancer cell line MDA-231 whereas they did not affect the proliferation of the non-aggressive human breast cancer cell line MCF-7, which contains very low PBR levels. This effect was reversible and not due to the antioxidant properties of the compounds tested. Using a human cDNA expression array we determined that EGb 761 treatment altered, in addition to PBR, the expression of 36 gene products involved in various pathways regulating cell proliferation. These in vitro data were further validated in an in vivo model where EGb 761 and GKB significantly inhibited the nuclear PBR expression and growth of MDA-231 cell xenografts in nude mice. Taken together, these data suggest that the manipulation of PBR expression could be used to control tumor growth and that EGb 761 and GKB, under the conditions used, exert cytostatic properties.

Animals↗

PubMedDate not supplied

Structure, function and regulation of the mitochondrial peripheral-type benzodiazepine receptor.

Steroid biosynthesis begins with the transfer of cholesterol from intracellular stores into mitochondria. Through in vitro and in vivo studies using various steroidogenic cell models and with the help of pharmacological, biochemical, morphological and molecular approaches we demonstrated that the peripheral-type benzodiazepine receptor (PBR) is an 18 kDa mitochondrial protein that interacts with other proteins in the outer mitochondrial membrane to form a multimeric complex. PBR is required for the binding, uptake and release, upon ligand activation, of the substrate cholesterol. Thus, cholesterol becomes available to the inner mitochondrial membrane P450scc where steroid biosynthesis begins. The presence of mitochondrial PBR is also critical in maintaining outer mitochondrial membrane stability and in preventing apoptosis. Considering these functions of PBR and the fact that PBR is a ubiquitous protein, it is suggested that this drug receptor may serve as a target to control various mitochondrial and cell functions and to protect against experimentally or pathologically induced mitochondrial and cell toxicity.

Animals↗