Lamia Fathallah

An aldose reductase inhibitor reverses early diabetes-induced changes in peripheral nerve function, metabolism, and antioxidative defense

Aldose reductase inhibitors (ARIs) prevent peripheral nerve dysfunction and morphological abnormalities in diabetic animal models. However, some experimental intervention studies and clinical trials of ARIs on diabetic neuropathy appeared disappointing because of either 1) their inadequate design and, in particular, insufficient correction of the sorbitol pathway activity or 2) the inability to reverse established functional and metabolic deficits of diabetic neuropathy by AR inhibition in general. We evaluated whether diabetes‐induced changes in nerve function, metabolism, and antioxidative defense are corrected by the dose of ARI (sorbinil, 65 mg/kg/d in the diet), resulting in complete inhibition of increased sorbitol pathway activity. The groups included control rats and streptozotocin‐diabetic rats treated with/without ARI for 2 weeks after 4 weeks of untreated diabetes. ARI treatment corrected diabetes‐induced nerve functional changes; that is, decrease in endoneurial nutritive blood flow, motor and sensory nerve conduction velocities, and metabolic abnormalities (i.e., mitochondrial and cytosolic NAD+/NADH redox imbalances and energy deficiency). ARI restored nerve concentrations of two major non‐enzymatic antioxidants, reduced glutathione (GSH) and ascorbate, and completely arrested diabetes‐induced lipid peroxidation. In conclusion, treatment with adequate doses of ARIs (that is, doses that completely inhibit increased sorbitol pathway activity) is an effective approach for reversal of, at least, early diabetic neuropathy.

Antioxidants attenuate early up regulation of retinal vascular endothelial growth factor in streptozotocin-diabetic rats

Abstract.Aims/hypothesis: A strong positive correlation has been found between lipid peroxidation product and vascular endothelial growth factor concentrations in the vitreous of patients with proliferative diabetic retinopathy. To establish a causal relation between diabetes-associated enhanced oxidative stress and vascular endothelial growth factor production, we evaluated two antioxidants, dl-α-lipoic acid and taurine, on retinal vascular endothelial growth factor protein and mRNA expression and on parameters of oxidative stress in streptozotocin-diabetic rats. Methods: Our experiments were on control rats and streptozotocin-diabetic rats with a 6-week duration of diabetes, treated with or without dl-α-lipoic acid (100 mg · kg–1· d–1, i. p.) or taurine (1 % in the diet) starting from induction of diabetes. Vascular endothelial growth factor protein in retinal homogenates was assessed by sandwich ELISA with an affinity-purified polyclonal antibody and vascular endothelial growth factor mRNA by ribonuclease protection assay. Retinal lipid peroxidation products i. e. malondialdehyde plus 4-hydroxyalkenals were quantified with n-methyl-2-phenylindole. Retinal reduced and oxidized glutathione, ascorbate, dehydroascorbate, and sorbitol pathway intermediates were measured spectrofluorometrically, and taurine by reverse-phase HPLC. Results: Vascular endothelial growth factor protein concentration (means ± SD) was increased in diabetic rats compared with control rats (33 ± 7 vs 19 ± 5 pg/mg total protein, p < 0.01) This increase was attenuated by taurine (26 ± 8, p < 0.05) and prevented by dl-α-lipoic acid (21 ± 4, p < 0.01). Vascular endothelial growth factor mRNA abundance was reduced by 1.4-fold in diabetic rats compared with control rats and this decrease was attenuated but not completely prevented by both antioxidants. Malondialdehyde plus 4-hydroxyalkenal concentration was increased in diabetic rats compared with control rats, and both antioxidants arrested accumulation of lipid peroxidation products. Taurine, reduced glutathione, oxidized glutathione, ascorbate, dehydroascorbate and sorbitol pathway intermediate concentrations as well as oxidized glutathione/reduced glutathione and dehydroascorbate/ascorbate ratios were similar in control and diabetic rats treated with or without taurine. Conclusion/interpretation: Oxidative stress is directly involved in up regulation of vascular endothelial growth factor protein in the retina during early diabetes. [Diabetologia (2001) 44: 1102–1110]

Early changes in lipid peroxidation and antioxidative defense in diabetic rat retina: effect of dl-α-lipoic acid

This study was designed to (1) evaluate retinal lipid peroxidation in early diabetes by the method specific for free malondialdehyde and 4-hydroxyalkenals, (2) identify impaired antioxidative defense mechanisms and (3) assess if enhanced retinal oxidative stress in diabetes is prevented by the potent antioxidant, DL-alpha-lipoic acid. The groups included control and streptozotocin-diabetic rats treated with or without DL-alpha-lipoic acid (100 mg kg(-1) day(-1), i.p., for 6 weeks). All parameters were measured in individual retinae. 4-Hydroxyalkenal concentration was increased in diabetic rats (2.63+/-0.60 vs. 1.44+/-0.30 nmol/mg soluble protein in controls, P<0.01), and this increase was prevented by DL-alpha-lipoic acid (1.20+/-0.88, P<0.01 vs. untreated diabetic group). Malondialdehyde, reduced glutathione (GSH) and oxidized glutathione (GSSG) concentrations were similar among the groups. Superoxide dismutase, glutathione peroxidase (GSHPx), glutathione reductase (GSSGRed) and glutathione transferase (GSHTrans) activities were decreased in diabetic rats vs. controls. Quinone reductase was upregulated in diabetic rats, whereas catalase and cytoplasmic NADH oxidase activities were unchanged. DL-alpha-Lipoic acid prevented changes in superoxide dismutase and quinone reductase activities induced by diabetes without affecting the enzymes of glutathione metabolism. In conclusion, accumulation of 4-hydroxyalkenals is an early marker of oxidative stress in the diabetic retina. Increased lipid peroxidation occurs in the absence of GSH depletion, and is prevented by DL-alpha-lipoic acid.

Early oxidative stress in the diabetic kidney: Effect of DL-α-lipoic acid

Oxidative stress is implicated in the pathogenesis of diabetic nephropathy. The attempts to identify early markers of diabetes-induced renal oxidative injury resulted in contradictory findings. We characterized early oxidative stress in renal cortex of diabetic rats, and evaluated whether it can be prevented by the potent antioxidant, DL-α-lipoic acid. The experiments were performed on control rats and streptozotocin-diabetic rats treated with/without DL-α-lipoic acid (100 mg/kg i.p., for 3 weeks from induction of diabetes). Malondialdehyde plus 4-hydroxyalkenal concentration was increased in diabetic rats vs. controls (p < .01) and this increase was partially prevented by DL-α-lipoic acid. F2 isoprostane concentrations (measured by GCMS) expressed per either mg protein or arachidonic acid content were not different in control and diabetic rats but were decreased several-fold with DL-α-lipoic acid treatment. Both GSH and ascorbate (AA) levels were decreased and GSSG/GSH and dehydroascorbate/AA ratios increased in diabetic rats vs. controls (p < .01 for all comparisons), and these changes were completely or partially (AA) prevented by DL-α-lipoic acid. Superoxide dismutase, glutathione peroxidase, glutathione reductase, glutathione transferase, and NADH oxidase, but not catalase, were upregulated in diabetic rats vs. controls, and these activities, except glutathione peroxidase, were decreased by DL-α-lipoic acid. In conclusion, enhanced oxidative stress is present in rat renal cortex in early diabetes, and is prevented by DL-α-lipoic acid.

Evaluation of a sorbitol dehydrogenase inhibitor on diabetic peripheral nerve metabolism: a prevention study

Aims/hypothesis. Studies of the role of sorbitol dehydrogenase in nerve functional deficits induced by diabetes reported contradictory results. We evaluated whether sorbitol dehydrogenase inhibition reduces metabolic abnormalities and enhances oxidative stress characteristic of experimental diabetic neuropathy. Methods. Control and streptozotocin-diabetic rats were treated with or without sorbitol dehydrogenase inhibitor (SDI)-157 (100 mg · kg–1· day–1, in the drinking water, for 3 weeks). Sciatic nerve free mitochondrial (cristae and matrix) and cytosolic NAD+: NADH ratios were calculated from the β-hydroxybutyrate, glutamate and lactate dehydrogenase systems. Concentrations of metabolites, e. g. sorbitol pathway intermediates and variables of energy state were measured in individual nerves spectrofluorometrically by enzymatic procedures. Results. The flux through sorbitol dehydrogenase (manifested by nerve fructose concentrations) was inhibited by 53 % and 74 % in control and diabetic rats treated with SDI compared with untreated control and diabetic groups. Free NAD+:NADH ratios in mitochondrial cristae, matrix and cytosol were decreased in diabetic rats compared with controls and reduction in either of the three variables was not prevented by sorbitol dehydrogenase inhibitor. Phosphocreatine concentrations and phosphocreatine:creatine ratios were decreased in diabetic rats compared with controls and were further reduced by the inhibitor. Malondialdehyde plus 4-hydroxyalkenals concentration was increased and reduced gluthathione concentration was reduced in diabetic rats compared with the control group, and changes in both variables were further exacerbated by sorbitol dehydrogenase inhibitor. Neither NAD-redox and energy states nor lipid aldehyde and reduced gluthathione concentrations were affected by treatment with the inhibitor in control rats. Conclusion/interpretation. Inhibition of sorbitol dehydrogenase does not offer an effective approach for prevention of oxidation and metabolic imbalances in the peripheral nerve that is induced by diabetes and is adverse rather than beneficial. [Diabetologia (1999) 42: 1187–1194]

Evaluation of α1-adrenoceptor antagonist on diabetes-induced changes in peripheral nerve function, metabolism, and antioxidative defense

The role for nerve blood flow (NBF) vs. other factors in motor nerve conduction (MNC) slowing in short‐term diabetes was assessed by evaluating α1‐adrenoceptor antagonist prazosin on NBF, MNC, as well as metabolic imbalances and oxidative stress in the neural tissue. Control and diabetic rats were treated with or without prazosin (5 mg*kg−1·d−1 for 3 wk). NBF was measured by hydrogen clearance. Both endoneurial vascular conductance and MNC velocity were decreased in diabetic rats vs. controls, and this decrease was prevented by prazosin. Free NAD+: NADH ratios in mitochondrial cristae, matrix, and cytosol assessed by metabolite indicator method, as well as phosphocreatine levels and phosphocreatine/creatine ratios, were decreased in diabetic rats, and this reduction was ameliorated by prazosin. Neither diabetes‐induced accumulation of two major glycation agents, glucose and fructose, as well as sorbitol and total malondialdehyde plus 4‐hydroxyalkenals nor depletion of myo‐inositol, GSH, and taurine or decrease in (Na/K)‐ATP‐ase activity were affected by prazosin. In conclusion, decreased NBF, but not metabolic imbalances or oxidative stress in the neural tissue, is a key mechanism of MNC slowing in short‐term diabetes. Further experiments are needed to estimate whether preservation of NBF is sufficient for prevention of nerve dysfunction and morphological abnormalities in long‐standing diabetes or whether the aforementioned metabolic imbalances closely associated with impaired neurotropism are of greater importance in advanced than in early diabetic neuropathy.–Obrosova, I. G., Van Huysen, C., Fathallah, L., Cao, X., Stevens, M. J., Greene, D. A. Evaluation of α1‐adrenoceptor antagonist on diabetes‐induced changes in peripheral nerve function, metabolism, and antioxidative defense. FASEB J. 14, 1548–1558 (2000)

Evaluation of an aldose reductase inhibitor on lens metabolism, ATPases and antioxidative defense in streptozotocin-diabetic rats: an intervention study.

Aims/hypothesis. Aldose reductase inhibitors (ARIs) prevent biochemical abnormalities associated with diabetic complications. We evaluated whether a short-term intervention with an adequate dose of ARI, introduced at the very early, precataractous stage, reversed diabetes-induced metabolic imbalances, down-regulation of ATPases and oxidative stress in the lens. Methods. The groups included mature control and streptozotocin-diabetic rats treated with or without ARI sorbinil (65 mg · kg–1· day–1, in the diet, for 2 weeks after 4 weeks of untreated diabetes). Free cytosolic NAD+:NADH and NADP+:NADPH ratios were calculated from the lactate dehydrogenase and malic enzyme systems. Concentrations of metabolites and adenine nucleotides, Na+/K+-ATPase, H+-ATPase and Ca++-independent Mg++-ATPase activities and variables of oxidative stress were measured in individual lenses. Results. Sorbinil treatment essentially corrected diabetes-induced sorbitol and fructose accumulation, myo-inositol depletion, decrease in free cytosolic NAD+:NADH ratio and energy deficiency. Malondialdehyde accumulation, reduced glutathione depletion and the increase in oxidized glutathione:reduced glutathione ratio were partially corrected. Free cytosolic NADP+:NADPH ratio and 4-hydroxyalkenal concentrations were similarly increased in diabetic rats treated with or without ARI. Sorbinil did not counteract diabetes-induced down-regulation of the three ATPase activities. Conclusion/interpretation. All biochemical changes assessed in our study are known to be prevented by ARIs. Despite the essential normalization of the sorbitol pathway activity, only part of them were, however, reversed by the ARI treatment introduced at the very early, i. e. precataractous, stage of diabetes. Therefore, intervention studies can easily underestimate the importance of aldose reductase in the pathogenesis of diabetic complications and should be interpreted with caution. [Diabetologia (2000) 43: 1048–1055]

Taurine counteracts oxidative stress and nerve growth factor deficit in early experimental diabetic neuropathy

Oxidative stress has a key role in the pathogenesis of diabetic complications. We have previously reported that taurine (T), which is known to counteract oxidative stress in tissues (lens, kidney, retina) of diabetic rats, attenuates nerve blood flow and conduction deficits in early experimental diabetic neuropathy (EDN). The purpose of this study was to evaluate whether dietary T supplementation counteracts oxidative stress and the nerve growth factor (NGF) deficit in the diabetic peripheral nerve. The experiments were performed in control rats and streptozotocin-diabetic rats fed standard or 1% T-supplemented diets for 6 weeks. All measurements were performed in the sciatic nerve. Malondialdehyde (MDA) plus 4-hydroxyalkenals (4-HA) were quantified with N-methyl-2-phenylindole. GSH, GSSG, dehydroascorbate (DHAA), and ascorbate (AA) were assayed spectrofluorometrically, T by reverse-phase HPLC, and NGF by ELISA. MDA plus 4-HA concentration (mean +/− SEM) was increased in diabetic rats (0.127 +/−0.006 vs 0.053 +/−0.003 mu mol/g in controls, P<0.01), and this increase was partially prevented by T (0.0960.004, P<0.01 vs untreated diabetic group). GSH levels were similarly decreased in diabetic rats treated with or without taurine vs controls. GSSG levels were similar in control and diabetic rats but were lower in diabetic rats treated with T (P<0.05 vs controls). AA levels were decreased in diabetic rats (0.133+0.015 vs 0.219 +/−0.023 mu mol/g in controls, P<0.05), and this deficit was prevented by T. DHAA/AA ratio was increased in diabetic rats vs controls (P<0.05), and this increase was prevented by T. T levels were decreased in diabetic rats (2.7 +/−0.16 vs 3.8 +/−0.1 mu mol/g in controls, P<0.05) and were repleted by T supplementation (4.20.3). NGF levels were decreased in diabetic rats (2.35 +/−0.20 vs 3.57 +/−0.20 ng/g in controls, P<0.01), and this decrease was attenuated by T treatment (3.160.28, P<0.05 vs diabetic group). In conclusion, T counteracts oxidative stress and the NGF deficit in early EDN. Antioxidant effects of T in peripheral nerve are, at least in part, mediated through the ascorbate system of antioxidative defense. The findings are consistent with the important role for oxidative stress in impaired neurotrophic support in EDN.