Low-protein diet disrupts the crosstalk between the PKA and PKC signaling pathways in isolated pancreatic islets.

Protein restriction in the early stages of life can result in several changes in pancreatic function. These alterations include documented reductions in insulin secretion and in cytoplasmic calcium concentration [Ca(2+)]i. However, the mechanisms underlying these changes have not been completely elucidated and may result, in part, from alterations in signaling pathways that potentiate insulin secretion in the presence of glucose. Our findings suggest that protein restriction disrupts the insulin secretory synergism between Cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and Ca(2+)-dependent protein kinase C (PKC) in isolated islets. Western blot analysis demonstrated reduced levels of both phospho-cAMP response element-binding protein (phospho-CREB) at Ser-133 and substrates phosphorylated by PKCs (Phospho-(Ser) PKC substrate), suggesting that PKA and PKC activity was impaired in islets from rats fed a low-protein diet (LP). cAMP levels and global Ca(2+) entry were also reduced in LP islets. In summary, our findings showed that protein restriction altered the crosstalk between PKA and PKC signaling pathways, resulting in the alteration of secretory synergism in isolated islets.

Chronic treatment with sildenafil stimulates Leydig cell and testosterone secretion.

The phosphodiesterase type 5 (PDE5) inhibitor, Sildenafil, is a novel, oral treatment approach for pulmonary hypertension. As Leydig cells present PDE5, this study was conducted to investigate the effects of the chronic treatment with Sildenafil (25 mg/kg) on male Swiss Webster mice steroidogenesis. After a 4-week long experimental design, Leydig cells were analysed by morphological and immunocytochemical procedures. Serum testosterone was assayed by radioimmunoassay. Leydig cells presented noteworthy ultrastructural alterations, such as a vesicular smooth endoplasmic reticulum, large vacuoles scattered through the cytoplasm, enlarged mitochondria with discontinue cristaes and whorle membranes with vesicles at the periphery, which are typical characteristics of an activated steroid-secreting cell. Important immunocytochemical labelling for steroidogenic acute regulatory protein, cytochrome P450 side-chain cleavage enzyme and testosterone were detected in isolated Leydig cells. In addition, Sildenafil-treated mice showed significant increased levels of total testosterone. The results obtained in the present study are consistent with the hypothesis that the accumulation of cyclic guanosine monophosphate by PDE5 inhibition could be involved in the androgen biosynthesis stimulation. Important clinical implications of hormonal disorders should be taken into account for patients with pulmonary hypertension.

Follicle development and luteal cell morphology altered by phosphodiesterase-5 inhibitor.

Vardenafil citrate is a potent vasodilator used in the treatment of patients with erectile dysfunction. Its mechanism of action is based on the selective inhibition of phosphodiesterase-5 (PDE5), specific to guanosine 3',5'-cyclic monophosphate (cGMP). Recently, chronic treatment with Vardenafil has been successfully used in cases of pulmonary hypertension and, despite being used in high doses for long periods, little is known about its effects on other systems. In the present study, female mice were treated daily with 5 mg/kg Vardenafil for 4 weeks, after which the ovaries were collected for morphological analyses and sera were collected for biochemical assays. This study found that treatment with Vardenafil decreased HDL serum levels and the number of antral follicles as well as induced lesser lipid content in luteal cells, suggesting that high levels of cGMP may affect follicle development.

Inhibitory action of in vitro ethanol and acetaldehyde exposure on LHRH and phorbol ester stimulated testerone secretion by rat testicular interstitial cells

Ha sido demostrado que el etanol y el acetaldehido inhiben la esteroidogenesis testicular. No obstante el (los) mecanismo(s) de trasnducción de señal envuelto en su acción no fue aún elucidado. Hemos examinado el posible envolvimiento de la proteína quinasa sensible a fosfolípidos y dependiente de calcio (proteína quinasa C, PK-C) en el mecanismo intracelular de acción de etanol y de acetaldehido, estimulando la producción de testosterona en células intersticiales de testículo de rata con LHRH y éster de forbolPDBu, los cuales activan la PK-C a nivel de receptor (LHRH) y pos-receptor (PDBu). El etanol(2000 mg por ciento) inhibió la producción de testosterona estimulada con 10**-7 M LHRH y 200 nM PDBu en 81 + 4,7 por ciento y 60 + 20.4 por ciento respectivamente. El acetaldehido (20 mg por ciento) redujo la cantidad de testosterona producida por 10**-7 M LHRH y 200 nM PDBu en 59,4 + 1,2 por ciento, respectivamente. El nivel basal de testosterona no fue modificado por el etanol pero si fue reducido por el acetaldehido. No obstante, la prueba funcional de la viabilidad celular a través de la preincubación de las células con las referidas dosis de etanol y acetaldehido no disminuyó la capacidad de respuesta a una subsecuente estimulación con LHRH, demostrando que la viabilidad celular no fue afectada por la incubación con esas substancias. Los resultados aquí presentados sugieren que la exposición directa al etanol y acetaldehido redujo la habilidad de las células intersticiales del testículo de rata de responder a la estimulación de la vía mediada por la PK-C

Inhibitory action of in vitro ethanol and acetaldehyde exposure on LHRH and phorbol ester stimulated testerone secretion by rat testicular interstitial cells

Ha sido demostrado que el etanol y el acetaldehido inhiben la esteroidogenesis testicular. No obstante el (los) mecanismo(s) de trasnducción de señal envuelto en su acción no fue aún elucidado. Hemos examinado el posible envolvimiento de la proteína quinasa sensible a fosfolípidos y dependiente de calcio (proteína quinasa C, PK-C) en el mecanismo intracelular de acción de etanol y de acetaldehido, estimulando la producción de testosterona en células intersticiales de testículo de rata con LHRH y éster de forbolPDBu, los cuales activan la PK-C a nivel de receptor (LHRH) y pos-receptor (PDBu). El etanol(2000 mg por ciento) inhibió la producción de testosterona estimulada con 10**-7 M LHRH y 200 nM PDBu en 81 + 4,7 por ciento y 60 + 20.4 por ciento respectivamente. El acetaldehido (20 mg por ciento) redujo la cantidad de testosterona producida por 10**-7 M LHRH y 200 nM PDBu en 59,4 + 1,2 por ciento, respectivamente. El nivel basal de testosterona no fue modificado por el etanol pero si fue reducido por el acetaldehido. No obstante, la prueba funcional de la viabilidad celular a través de la preincubación de las células con las referidas dosis de etanol y acetaldehido no disminuyó la capacidad de respuesta a una subsecuente estimulación con LHRH, demostrando que la viabilidad celular no fue afectada por la incubación con esas substancias. Los resultados aquí presentados sugieren que la exposición directa al etanol y acetaldehido redujo la habilidad de las células intersticiales del testículo de rata de responder a la estimulación de la vía mediada por la PK-C (AU)

Fluoride and phosphatidylserine induced inhibition of cytosolic insulin-degrading activity

Cytosol (C) (100,000 x g/60 min, supernatant) from liver, brain and testis (Wistar male rats) are shown to contain insulin degrading activity (C-IDA). The regulation of C-IDA in these fractions by ligands that activate G protein and PKC were examined C-IDA from liver, brain and testis was inhibited 76%; 64% and 50% by 50 mM F- respectively. Chromatography of C fraction from liver on Sephadex G-50 in presence of 1 M (NH4)2SO4 and 20% (v/v) glycerol (experimental condition to remove guanine nucleotides from G proteins) decreased in about 3-fold aluminum fluoride effect on C-IDA. Mg++ (from 5mM to 10 mM) enhanced fluoride effects by inhibiting fully C-IDA. Phosphatidylserine in presence of ATP completely inhibited C-IDA; this inhibition was 31.3% mediated by a phosphorylation reaction. It is concluded that cytosol from different tissues contain proteins capable to associate ligands as aluminum fluoride and PS to regulate C-IDA. It is proposed a mechanism of protein-protein interaction to modulate C-IDA (Au)

Fluoride and phosphatidylserine induced inhibition of cytosolic insulin-degrading activity

Cytosol (C) (100,000 x g/60 min, supernatant) from liver, brain and testis (Wistar male rats) are shown to contain insulin degrading activity (C-IDA). The regulation of C-IDA in these fractions by ligands that activate G protein and PKC were examined C-IDA from liver, brain and testis was inhibited 76%; 64% and 50% by 50 mM F- respectively. Chromatography of C fraction from liver on Sephadex G-50 in presence of 1 M (NH4)2SO4 and 20% (v/v) glycerol (experimental condition to remove guanine nucleotides from G proteins) decreased in about 3-fold aluminum fluoride effect on C-IDA. Mg++ (from 5mM to 10 mM) enhanced fluoride effects by inhibiting fully C-IDA. Phosphatidylserine in presence of ATP completely inhibited C-IDA; this inhibition was 31.3% mediated by a phosphorylation reaction. It is concluded that cytosol from different tissues contain proteins capable to associate ligands as aluminum fluoride and PS to regulate C-IDA. It is proposed a mechanism of protein-protein interaction to modulate C-IDA