Outcome of acute renal injury in diabetic mice with experimental endotoxemia: role of hypoxia-inducible factor-1 α.

The role of diabetic nephropathy in the outcome of acute renal injury (AKI) is not well defined. Herein we evaluate the outcome of lipopolysaccharide- (LPS-) induced AKI in streptozotocin-induced diabetes, as well as the potential role of Hypoxia Inducible Factor (HIF-1 α ) in this condition. Although 6 h after LPS injection all mice developed a decrease in renal function, proteinuric diabetic mice showed a better recovery of this parameter throughout the study (72 h). Both HIF-1 α and vascular endothelium growth factor (VEGF) were found to be upregulated in diabetic mice. After LPS injection, all animals showed an upregulation of these factors, although it was higher in the diabetic group. Glycated albumin (GA) was found to upregulate HIF-1 α in HK-2 cells, which resulted in increased production of VEGF. Interestingly, LPS cooperated with GA to induce HIF-1 α upregulation. In conclusion, diabetic mice display a better recovery of AKI after experimental endotoxemia. Moreover, these animals showed an increased expression of both HIF-1 α and VEGF that was reproduced by incubating renal cells with GA. Since VEGF is considered a survival factor for tubular cells, our findings suggest that diabetes displays HIF-1 α upregulation that might function as a "precondition state" offering protection from endotoxic AKI.

Enhancement of TNF-alpha-induced apoptosis by immobilized arginine-glycine-aspartate: involvement of a tyrosine kinase-dependent, MAP kinase-independent mechanism.

Extracellular matrix facilitates anchorage-dependent cell survival via interaction of its arginine-glycine-aspartate (RGD) motif with integrins. In this report, we describe an unexpected, apoptosis-promoting the effect of immobilized RGD (iRGD) on tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis. Mesangial cells cultured on RGD-coated plates showed enhanced susceptibility to TNF-alpha-induced apoptosis. iRGD alone did not affect cell survival. In contrast, iRGD did not facilitate but inhibited apoptosis induced by H(2)O(2). Mitogen-activated protein (MAP) kinases and tyrosine kinases are important mediators for the RGD-integrin signaling. Pretreatment with MAP kinase kinase inhibitor PD098059, c-Jun N-terminal kinase (JNK)-c-Jun/AP-1 inhibitor curcumin or p38 MAP kinase inhibitor SB203580 did not attenuate the apoptosis-promoting effect of iRGD. Consistently, transfection with dominant-negative mutants of extracellular signal-regulated kinases, JNK or p38 MAP kinase did not inhibit the effect of iRGD. In contrast, protein tyrosine kinase inhibitors, genistein, and herbimycin A, abrogated the apoptosis-promoting effect of iRGD. Of note, TNF-alpha-induced apoptosis on uncoated plates was not attenuated by tyrosine kinase inhibitors. These data provide the first evidence that iRGD accelerates certain apoptosis. We identified that the effect was mediated by the tyrosine kinase-dependent, MAP kinase-independent mechanism.

Selective involvement of superoxide anion, but not downstream compounds hydrogen peroxide and peroxynitrite, in tumor necrosis factor-alpha-induced apoptosis of rat mesangial cells.

Tumor necrosis factor-alpha (TNF-alpha) induces reactive oxygen species (ROS) that serve as second messengers for intracellular signaling. Currently, precise roles of individual ROS in the actions of TNF-alpha remain to be elucidated. In this report, we investigated the roles of superoxide anion (O-(2)), hydrogen peroxide (H(2)O(2)), and peroxynitrite (ONOO(-)) in TNF-alpha-triggered apoptosis of mesangial cells. Mesangial cells stimulated by TNF-alpha produced O-(2) and underwent apoptosis. The apoptosis was inhibited by transfection with manganese superoxide dismutase or treatment with a pharmacological scavenger of O-(2), Tiron. In contrast, although exogenous H(2)O(2) induced apoptosis, TNF-alpha-triggered apoptosis was not affected either by transfection with catalase cDNA or by treatment with catalase protein or glutathione ethyl ester. Similarly, although ONOO(-) precursor SIN-1 induced apoptosis, treatment with a scavenger of ONOO(-), uric acid, or an inhibitor of nitric oxide synthesis, N(G)-nitro-L-argininemethyl ester hydrochloride, did not affect the TNF-alpha-triggered apoptosis. Like TNF-alpha-induced apoptosis, treatment with a O-(2)-releasing agent, pyrogallol, induced typical apoptosis even in the concurrent presence of scavengers for H(2)O(2) and ONOO(-). These results suggested that, in mesangial cells, TNF-alpha induces apoptosis through selective ROS. O-(2), but not H(2)O(2) or ONOO(-), was identified as the crucial mediator for the TNF-alpha-initiated, apoptotic pathway.

Suppression of apoptosis by all-trans-retinoic acid. Dual intervention in the c-Jun n-terminal kinase-AP-1 pathway.

Retinoic acid induces apoptosis of various cells, whereas little is known about its anti-apoptotic potential. In this report, we describe an anti-apoptotic property of all-trans-retinoic acid (t-RA) in mammalian cells. Mesangial cells exposed to hydrogen peroxide (H2O2) exhibited shrinkage of the cytoplasm, membrane blebbing, condensation of nuclei, and DNA fragmentation. Pretreatment with t-RA attenuated the morphologic and biochemical hallmarks of apoptosis. t-RA also inhibited apoptosis of mesangial cells triggered by pyrrolidine dithiocarbamate, whereas it did not prevent tumor necrosis factor-alpha-induced apoptosis. The anti-apoptotic effect against H2O2 was similarly observed in NRK49F fibroblasts, but not in Madin-Darby canine kidney epithelial cells and ECV304 endothelial cells. Mesangial cells exposed to H2O2 undergo apoptosis via the activator protein 1 (AP-1)-dependent pathway. We found that t-RA abrogated the H2O2-induced expression of c-fos/c-jun and activation of AP-1. Furthermore, t-RA inhibited H2O2-triggered activation of c-Jun N-terminal kinase (JNK), and dominant-negative inhibition of JNK attenuated the H2O2-induced apoptosis. These data disclosed the novel potential of retinoic acid as an inhibitor of apoptosis. The anti-apoptotic action of t-RA was ascribed, at least in part, to dual suppression of the cell death pathway mediated by JNK and AP-1.

Prevention by tretinoin (all-trans-retinoic acid) of age-related renal changes.

The present work shows that dietary tretinoin (all-trans-retinoic acid) supplementation may be a useful manoeuvre for the prevention of age-related renal changes. 18-month-old male Fischer 344 rats fed during three months with standard chow plus tretinoin (1 mg/kg/day) did not exhibit any adverse effect in terms of bodyweight, urinary volume, renal handling of sodium and both hematological and blood chemistry parameters. Although the diet did not reduce age-related proteinuria nor renal lipid peroxidation, glomerular filtration rate and renal cortex protein content were, respectively, 30% higher and 30% lower than in age-matched control rats. These results suggest that dietary tretinoin supplementation may be a useful manoeuvre to slow the progression of age-related renal changes. Since glomerular H2O2 production increases during renal aging in rats, we studied the effect of tretinoin on the biology of cultured glomerular rat mesangial cells exposed to H2O2. Preincubation with tretinoin abolished cell proliferation or cell death induced, respectively, by low and high concentrations of H2O2. These results suggest that the modulation of the cellular actions of H2O2 may be relevant in the mechanisms through which tretinoin prevents age-related renal changes.

A possible role for platelet-activating factor in the hydrogen peroxide-induced TXB2 and PGE2 glomerular synthesis.

Hydrogen peroxide stimulates both prostanoid and platelet-activating factor (PAF) biosynthesis in cultured rat mesangial cells and isolated rat glomeruli. The present experiments were designed to try to establish some relationship between prostanoid and PAF synthesis in these renal structures, in the presence of hydrogen peroxide. Cells and glomeruli were incubated with hydrogen peroxide under different experimental conditions, and thromboxane B2 (TXB2), the stable metabolite of thromboxane A2 (TXA2), and prostaglandin E2 (PGE2) concentrations were measured in the supernatants of the cells or glomeruli. Moreover, H2O2-dependent PAF synthesis was measured by high performance liquid chromatography (HPLC) ([3H]acetate incorporation) and radioimmunoassay. H2O2 induced increased TXB2 and PGE2 production in cultured rat mesangial cells and isolated rat glomeruli. This effect was blocked by incubation in the presence of a PAF-receptor antagonist, BN-52021. This antagonist has no intrinsic effect either in basal prostanoid synthesis or in arachidonic acid-stimulated glomerular TXB2 synthesis. Alprazolam, another PAF antagonist, nonchemically related to BN-52021, also completely blocked the H2O2-induced production of TXB2 by isolated rat glomeruli. Moreover, H2O2 was also able to induce an increased [3H]acetate incorporation into a fraction comigrating with a PAF standard in HPLC in isolated glomeruli, and this effect was dependent on the H2O2 concentration tested. Moreover, H2O2 was also able to induce an increased [3H]acetate incorporation and increased synthesis of radioimmunoassayable PAF in cultured mesangial cells. These results suggest that the increased synthesis of PGE2 and TXB2 induced by H2O2 could be dependent on platelet-activating factor production.