Characterization of oxidative stress in blood from diabetic vs. hypercholesterolaemic patients, using a novel synthesized marker

In the present study, we extend our novel concept of designing and using exogenous markers for the characterization of oxidative stress (OS) and OS-associated diseases. The aim was to use such a synthetic compound as a tool for studying OS in blood from diabetic and hypercholesterolaemic (Hc) patients. The marker used N-linoleoyl tyrosine (LT) was constructed from tyrosine and linoleic acid (LA); both components are known to be easily oxidized upon exposure to different types of reactive oxygen/nitrogen species (ROS/RNS), and to generate specific oxidized products, depending on the type of oxidants present in vivo. Using the LT probe, we showed that the ratios of oxidized LT to total LT (Ox-LT/LT) is significantly higher in blood samples obtained from diabetic patients, than in Hc patients or healthy control subjects. LC/MS analysis revealed that blood from diabetic patients oxidizes the marker with predominant formation of Ox-LT hydroperoxide (LT-OOH) and epoxide (epoxy-LT), where the LA moiety is oxidized to hydroperoxide and to epoxide, respectively. Analysis of oxysterol levels in these samples (GC/MS) revealed that the blood of both diabetic and Hc patients contained significantly more oxysterols than blood of control subjects. Consumption of pomegranate juice by diabetic patients for 3 months suppressed their blood capacity to oxidize the LT and similarly also reduced their blood oxysterol/total cholesterol ratio by 93%. The use of an exogenous marker to characterize OS in blood samples yields important information on the extent of OS, and can provide a fingerprint for the early identification of different pathological conditions associated with OS.     

Plasma angiogenesis and oxidative stress markers in patients with diabetic retinopathy

Abstract

Background: Neovascularization in the retina and hyperglycaemia-induced oxidative stress are implicated in the pathogenesis of diabetic retinopathy (DR). In this study, we hypothesized that the plasma angiogenic and oxidative stress markers associated with these derangements could aid in the screening of diabetic patients who are at an increased risk of developing retinopathy.

Methods: This study included normal (n?=?148), type2 diabetes without retinopathy (DNR; n?=?148), proliferative DR (PDR; n?=?74) and non-PDR (NPDR; n?=?148) subjects. Plasma concentrations of vascular endothelial growth factor-A (VEGF-A), hypoxia-inducible factor-1α (HIF-1α), matrix metalloproteinase-9 (MMP-9), pigment epithelium-derived factor (PEDF), nitric oxide (NO), soluble receptors for advanced glycation end products (sRAGE), malondialdehyde (MDA) and protein thiols were estimated.

Results: A statistically significant increase was observed in the plasma concentrations of pro-angiogenic factors and markers of oxidative stress in both retinopathy groups. By contrast, the concentrations of anti-angiogenic factors and antioxidants were decreased significantly in these groups. Receiver operating characteristic analysis indicated that the plasma thresholds of HIF-1α and PEDF can be suitable markers in case of NPDR. However, in PDR, HIF-1α, NO, MMP-9 and PEDF showed high sensitivity and specificity.

Conclusions: The factors associated with hypoxia, matrix degradation and angiogenic inhibition play a crucial role in predicting DR.     

Lipoic acid ameliorates oxidative stress and renal injury in alloxan diabetic rabbits

The therapeutic potential of lipoic acid (LA) in diabetes and diabetic nephropathy treatment was elucidated. Alloxan diabetic rabbits were treated daily for three weeks with either 10 or 50 mg of LA per kg body weight (i.p.). The following parameters were measured: 1) serum glucose, urea, creatinine and hydroxyl free radical (HFR) levels; 2) blood glutathione redox state; 3) urine albumin concentration; 4) hepatic and renal HFR levels, GSH/GSSG ratios, cysteine contents and the activities of the enzymes of glutathione metabolism; and 5) the activity of renal NADPH oxidase. Histological studies of kidneys were also performed. The treatment of diabetic rabbits with 50 mg of LA resulted in lethal hypoglycaemia in 50% of animals studied. Although the low dose of LA did not change serum glucose concentration, it decreased serum urea and creatinine concentrations, attenuated diabetes-induced decline in GSH/GSSG ratio and abolished hydroxyl free radicals accumulation in serum, liver and kidney cortex. LA did not change the activities of the enzymes of glutathione metabolism, but it elevated hepatic content of cysteine, which limits the rate of glutathione biosynthesis. Moreover, LA lowered urine albumin concentration and attenuated glomerulopathy characteristic of diabetes. However, it did not affect diabetes-stimulated activity of renal NADPH oxidase. In view of these data, it is concluded that low doses of LA might be useful for the therapy of diabetes and diabetic nephropathy. Beneficial action of LA seems to result mainly from direct scavenging of HFR and restoring glutathione redox state due to elevation of intracellular cysteine levels.     

Dexpanthenol reduces diabetic nephropathy and renal oxidative stress in rats

Hyperglycemia increases reactive oxygen species (ROS) and the resulting oxidative stress contributes to the development of diabetic complications. Dexpanthenol (Dxp) is the biological active form of pantothenic acid. We investigated whether Dxp administration could decrease oxidative stress as a way to treat renal complications of diabetes mellitus (DM). Thirty-two male Wistar albino rats were divided into four groups: control, Dxp, DM and DM + Dxp. Experimental diabetes was induced by a single dose of streptozotocin (STZ). After administration of STZ, the DM + Dxp group was administered 500 mg/kg Dxp intraperitoneally every day for 6 weeks. At the end of the study, blood glucose levels were measured and rats were sacrificed. Kidneys were embedded in paraffin, sectioned and stained with hematoxylin and eosin, and periodic acid-Schiff. The mean malondialdehyde levels, glutathione peroxidase, superoxide dismutase and catalase activities, and total antioxidant and total oxidant status also were measured. The control group was normal in histological appearance. We observed congestion, inflammation, glomerulosclerosis, tubular desquamation, loss of villi and hydropic degeneration in tubule cells in the DM group. Indicators of oxidative stress were elevated and antioxidant activity was reduced in the DM group compared to controls. In the DM + Dxp group, oxidative stress was decreased, antioxidant activity was increased and histopathological changes were reduced compared to the DM group. We found that Dxp exhibited ameliorative effects on STZ induced diabetic nephropathy by increasing antioxidant activity.     

Association between miRNAs expression and signaling pathways of oxidative stress in diabetic retinopathy

Diabetic retinopathy (DR) is a major cause of vision reduction in diabetic patients. Hyperglycemia is a known instigator for the development of DR, even though the role of oxidative stress pathways in the pathogenesis of DR is established. The studies indicate that microRNAs (miRNAs) are significant to the etiology of DR; changes in miRNAs expression levels may be associated with onset and progression of DR. In addition, miRNAs have emerged as a useful disease marker due to their availability and stability in detecting the severity of DR. The relationship between miRNAs expression levels and oxidative stress pathways has been investigated in several studies. The aim of this study is the examination of function and expression levels of target miRNAs in oxidative stress pathway and pathogenesis of diabetic retinopathy.     

Sirt3-dependent deacetylation of SOD2 plays a protective role against oxidative stress in oocytes from diabetic mice

Maternal diabetes has been demonstrated to adversely affect oocyte quality in mouse oocytes. However, the potential molecular mechanisms are poorly understood. Here, we established a type I diabetic mouse model and detected the increased reactive oxygen species (ROS) levels and decreased Sirt3 expression in oocytes from diabetic mice. Furthermore, we found that forced expression of Sirt3 in diabetic oocytes significantly attenuates such an excessive production of ROS. The acetylation status of lysine 68 of superoxide dismutase (SOD2K68) is dependent on Sirt3 in oocytes. In line with this, SOD2K68 acetylation levels were markedly increased in diabetic oocytes, and Sirt3 overexpression could effectively suppress this tendency. Importantly, the deacetylation-mimetic mutant SOD2K68R is capable of partly preventing the oxidative stress in oocytes from diabetic mice. In conclusion, our findings support a model where Sirt3 plays a protective role against oxidative stress in oocytes exposed to maternal diabetes through deacetylating SOD2K68.     

Mitochondrial dysfunction and oxidative stress in diabetic wound

Diabetic wounds nowadays have become a major health challenge with the changes of the disease spectrum. Mitochondria are closely associated with stubborn nonhealing diabetic wounds for their vital role in energy metabolism, redox homeostasis, and signal transduction. There is significant mitochondrial dysfunction and oxidative stress in diabetic wounds. However, the contribution of mitochondrial dysfunction in oxidative stress induced nonhealing diabetic wound is still not fully understood. In this review, we will briefly summarize the current knowledge of the reported signaling pathways and therapeutic strategies involved in mitochondrial dysfunction in diabetic wounds. The findings provide further understanding of strategies that focus on mitochondria in diabetic wound treatment.     

The effect ofGongronema latifolium extracts on serum lipid profile and oxidative stress in hepatocytes of diabetic rats

Diabetes is known to involve oxidative stress and changes in lipid metabolism. Many secondary plant metabolites have been shown to possess antioxidant activities, improving the effects of oxidative stress on diabetes. This study evaluated the effects of extracts from Gongronema latifolium leaves on antioxidant enzymes and lipid profile in a rat model of non insulin dependent diabetes mellitus (NIDDM). The results confirmed that the untreated diabetic rats were subjected to oxidative stress as indicated by significantly abnormal activities of their scavenging enzymes (low superoxide dismutase and glutathione peroxide activities), compared to treated diabetic rats, and in the extent of lipid peroxidation (high malondialdehyde levels) present in the hepatocytes. The ethanolic extract of G. latifolium leaves possessed antioxidant activity as shown by increased superoxide dismutase and glutathione peroxidase activities and decreases in malondialdehyde levels. High levels of triglycerides and total cholesterol, which are typical of the diabetic condition, were also found in our rat models of diabetes. The ethanolic extract also significantly decreased triglyceride levels and normalized total cholesterol concentration.     

Ulva rigida improves carbohydrate metabolism, hyperlipidemia and oxidative stress in streptozotocin-induced diabetic rats

This study was designed to investigate the effects of Ulva rigida, one of the green algae, on the lipid profile and oxidative–antioxidative systems in streptozotocin‐induced diabetic rats. Forty Wistar rats randomly divided into four groups: control (C), control + U. rigida extract (C + URE), diabetes (D) and diabetes + U. rigida extract (D + URE). U. rigida (2%) was administered in drinking water for 5 weeks after the induction of diabetes. U. rigida reduced the blood glucose, serum total cholesterol, triglyceride levels and plasma and tissue malondialdehyde (MDA) levels in the D + URE group. Insulin levels were significantly higher in the D + URE than those of the D group. Serum total cholesterol and tissue MDA levels were reduced in the C + URE group. Whole blood glutathione peroxidase and erythrocyte superoxide dismutase activities were higher in the D and C + URE groups compared with the C group. Paraoxonase and arylesterase activities were lower in the D group while U. rigida increased paraoxonase activities in C + URE and D + URE groups. This is the first study which showed U. rigida has antidiabetic and antihyperlipidemic effects and improves oxidative stress in diabetic rats. We conclude that U. rigida might have a potential use as a protective and/or therapeutic agent in diabetes mellitus. Copyright © 2011 John Wiley & Sons, Ltd.     

α-亚麻酸对糖尿病大鼠炎症介质和氧化应激的影响

目的:观察α-亚麻酸(ALA)对糖尿病大鼠体内炎症介质和氧化应激的影响,探讨ALA在糖尿病防治中的作用。方法:雄性SD大鼠高脂饮食喂养4周后,腹腔注射链脲佐菌素(STZ)30 mg/kg建立2型糖尿病(T2DM)模型。将大鼠随机分为3组(n=10):正常对照组、糖尿病模型组和ALA治疗组(500μg/kg.d)。4周后测定大鼠血清中肿瘤坏死因子(TNF-α)、可溶性P-选择素(sP-selectin)、可溶性细胞间黏附分子(sICAM-1)、一氧化氮(NO)、丙二醛(MDA)的含量以及超氧化物岐化酶(SOD)和过氧化氢酶(CAT)的活性。结果:与正常对照组相比,糖尿病大鼠血清中炎症介质TNF-α、sP-selectin和sICAM-1的含量增加,血清NO含量下降而MDA升高,同时抗氧化酶SOD和CAT的活性降低;ALA治疗可显著降低糖尿病大鼠血清中TNF-α、sP-selectin和sICAM-1的含量(与STZ+vehicle组相比,P<0.01),增加血清NO水平并减少MDA含量,升高抗氧化酶SOD和CAT的活性(与STZ+vehicle组相比,均P<0.05)。结论:ALA可显著降低糖尿病大鼠血清炎症介质的生成,减轻氧化应激水平,具有抗炎和抗氧化作用。提示ALA对糖尿病及糖尿病并发症的发生发展可能具有一定的防治作用。     

A histological,immunohistochemical and biochemical study of the effects of pomegranate peel extracts on gibberellic acid induced oxidative stress in adult rat testes

ABSTRACT

Gibberellins are commonly used plant growth regulators that exhibit deleterious effects on various animal tissues. We investigated the histological, immunohistochemical and biochemical effects of gibberellic acid (GA3) on rat testes as well as the possible protective role of pomegranate peel extract (PPE). We used 28 adult male rats divided into control, PPE treated, GS3 treated and GA3 + PPE treated groups. Testis specimens were analyzed for superoxide dismutase (SOD) and catalase (CAT) activity, and examined histologically. We also investigated the androgen receptor using immunohistochemistry. The GA3 treated group exhibited significantly decreased SOD and CAT levels and area percent of androgen receptor. Seminiferous tubules (ST) were widely separated and the germinal epithelium was separated from the basement membrane in some tubules. Areas of vacuolation, degenerated germ cells with pyknotic nuclei and large multinucleated cells were observed. Ultrastructurally, primary spermatocytes exhibited vacuolated cytoplasm, degenerated mitochondria and hyperchromatic nuclei. Degenerated early spermatids with a ruptured or hyperchromatic nucleus were found. Spermatozoa exhibited abnormalities of the head and tail. The cytoplasm of Sertoli and Leydig cells exhibited dilated smooth endoplasmic reticulum. A significant improvement of the biochemical, histological and immunohistochemical alterations was observed in the GA3 + PPE treated group compared to the GA3 treated group.     

糖、脂代谢及氧化应激与糖尿病肾病的相关性

糖尿病肾病是糖尿病的主要并发症之一。近年来,糖尿病肾病的发病率呈逐渐上升趋势。研究发现,糖、脂代谢与糖尿病肾病的发生密切相关,而氧化应激也在糖、脂代谢异常中起重要作用。本文综述了有关糖、脂代谢与糖尿病肾病相关性研究,以及硫氧还蛋白及硫氧还蛋白结合蛋白2在糖尿病肾病发生中的作用。     

Resveratrol‐enhanced autophagic flux ameliorates myocardial oxidative stress injury in diabetic mice

Autophagic dysfunction is observed in diabetes mellitus. Resveratrol has a beneficial effect on diabetic cardiomyopathy. Whether the resveratrol‐induced improvement in cardiac function in diabetes is via regulating autophagy remains unclear. We investigated the mechanisms underlying resveratrol‐mediated protection against heart failure in diabetic mice, with a focus on the role of sirtuin 1 (SIRT1) in regulating autophagic flux. Diabetic cardiomyopathy in mice was induced by streptozotocin (STZ). Long‐term resveratrol treatment improved cardiac function, ameliorated oxidative injury and reduced apoptosis in the diabetic mouse heart. Western blot analysis revealed that resveratrol decreased p62 protein expression and promoted SIRT1 activity and Rab7 expression. Inhibiting autophagic flux with bafilomycin A1 increased diabetic mouse mortality and attenuated resveratrol‐induced down‐regulation of p62, but not SIRT1 activity or Rab7 expression in diabetic mouse hearts. In cultured H9C2 cells, redundant or overactive H₂O₂ increased p62 and cleaved caspase 3 expression as well as acetylated forkhead box protein O1 (FOXO1) and inhibited SIRT1 expression. Sirtinol, SIRT1 and Rab7 siRNA impaired the resveratrol amelioration of dysfunctional autophagic flux and reduced apoptosis under oxidative conditions. Furthermore, resveratrol enhanced FOXO1 DNA binding at the Rab7 promoter region through a SIRT1‐dependent pathway. These results highlight the role of the SIRT1/FOXO1/Rab7 axis in the effect of resveratrol on autophagic flux in vivo and in vitro, which suggests a therapeutic strategy for diabetic cardiomyopathy.     

Exogenous N-linoleoyl tyrosine marker as a tool for the characterization of cellular oxidative stress in macrophages

Oxidative stress and its resultant products continue to attract investigators. Numerous endogenous substances have been suggested as potential markers for the identification of oxidative stress in tissues and organisms. In this study, we present a novel concept whereby an exogenous marker is designed and synthesized for the characterization of oxidative stress. The designed marker is constructed from tyrosine (Tyr) and linoleic acid (LA), which are attached covalently to form N-linoleoyl tyrosine (N-LT). Each of the two components (Tyr and LA) is known to be easily oxidized upon exposure to different types of reactive species. Combining the two allows their distinction from the endogenous Tyr and LA in the tested biological samples. The ability of the N-LT marker to characterize oxidative stress in macrophage cell lines was first studied using different types of ROS/RNS. N-LT was found to interact with macrophages, binding to the cell membrane. Upon treatment of J-774 A.1 macrophages with N-LT (40 μM) and with various oxidants; HOCl (0.2, 0.4 mM), copper ions (20 μM), SIN-1 (0.1, 1.0 mM), specific oxidized N-LT (Ox-N-LT) products were formed, depending on the type of oxidant used. Exposing cells to HOCl (0.2 mM) resulted in exclusive attack of the LA residue of N-LT, preferentially forming an adduct of HOCl to the LA double bond (N-L(HOCl)T, 4.3%). In contrast, when SIN-1 (0.1 mM) was applied as the oxidant, the Tyr moiety of N-LT was most reactive, yielding a nitration product of the Tyr aromatic ring (N-LT(NO₂), 1.8%). Similar N-LT oxidation in cell-free systems yielded a significantly higher content of Ox-N-LT (10.8% N-L(HOCl)T, 7% N-LT(NO₂)). The designed marker was then tested with peritoneal macrophages taken from atherosclerotic apolipoprotein-deficient (E⁰) mice showing specific and selective oxidation of N-LT to yield N-LT-hydroperoxide (1.9% N-L(OOH)T), at significantly higher levels than resulted from similar experiments using peritoneal macrophages harvested from control BalbC mice (0.0% N-L(OOH)T). In contrast, the differences in N-L(epoxy)T level between BalbC and E⁰ mice were not significant using both types of peritoneal macrophages (E⁰ and BalbC), suggesting that N-L(OOH)T is characteristic of the atherosclerotic state. Thus, we show that the designed marker is sufficiently sensitive to detect oxidative stress imposed on cells and cell-free systems and to react selectively with the various ROS/RNS induced. Such a marker may be useful for characterizing oxidative stress in general, and possibly also in oxidative-stress-associated diseases.     

The effect of consumption of purple grape juice in the gestational period on oxidative stress parameters in Wistar rat foetuses

This study aimed to evaluate the effects of purple grape juice consumption in pregnancy on oxidative stress parameters in Wistar rat fetuses. Twenty-four pregnant rats were divided into five groups: control group, indomethacin group (received a single dose of indomethacin in DG20), group grape juice DG14 (received an amount for 14 days/first and second gestational trim), group grape juice DG20 (received a dose throughout the gestational period), group grape juice two doses (received two doses, at morning and afternoon). On the 20th day of pregnancy (DG20), rats were anesthetized, and a cesarean section was performed to obtain the fetuses. A sample of liver, heart, and total brain of fetuses was collected for oxidative stress analyses. Values P<0.05 were considered significant. In fetuses’ heart, we observed that the grape juice two dose group decreased sulfhydryl and increased SOD. In the liver, the grape juice decreased TBARS and SOD. There was a decrease in carbonyl and sulfhydryl in the indomethacin and grape juice one dose groups in the brain. We conclude that indomethacin altered oxidative stress parameters only in the fetal brain, and grape juice was presented as an important modulator of antioxidant capacity when consumed in a dose.     

NADK3, a novel cytoplasmic source of NADPH, is required under conditions of oxidative stress and modulates abscisic acid responses in Arabidopsis

In plants, excess reactive oxygen species are toxic molecules induced under environmental stresses, including pathogen invasions and abiotic stresses. Many anti-oxidant defense systems have been reported to require NADPH as an important reducing energy equivalent. However, the sources of NADPH and the molecular mechanisms of maintaining cytoplasmic redox balance are unclear. Here, we report the biological function of a putative cytoplasmic NADH kinase (NADK3) in several abiotic stress responses in Arabidopsis. We found that cytoplasmic NADPH is provided mostly by the product of the NADK3 gene in Arabidopsis. Expression of he NADK3 gene is responsive to abscisic acid (ABA) and abiotic stress conditions, including methyl violgen (MV), high salinity and osmotic shock. An NADK3 null mutant showed hypersensitivity to oxidative stress in both seed germination and seedling growth. Seed germination of the mutant plants also showed increased sensitivity to ABA, salt and mannitol. Furthermore, stress-related target genes were identified as upregulated in the mutant by mannitol and MV. Our study indicates that this cytoplasmic NADH kinase, a key source of the cellular reductant NADPH, is required for various abiotic stress responses.     

Effect of acute vs chronic H2O2-induced oxidative stress on antioxidant enzyme activities

H₂O₂ can freely crosses membranes and in the presence of Fe²⁺ (or Cu⁺) it is prone to participate in Fenton reaction. This study evaluated the concentration and time-dependent effects of H₂O₂-induced oxidative stress on MnSOD, Se:GPx and catalase and on aconitase. Acute and chronic H₂O₂ treatments were able to induce oxidative stress in HeLa cells as they significantly decreased aconitase activity and also caused a very significant decrease on antioxidant enzyme activities. The inhibition of enzyme activities was time- and concentration-dependent. Chronic treatment with 5 µM H₂O₂/h after 24 h was able to decrease all enzyme activities almost at the same level as the acute treatment. Acute and chronic treatments on antioxidant enzyme activities were prevented by cell treatment with ascorbic acid or N-acetylcysteine. These results indicate that antioxidant enzymes can also be affected by the same ROS they produce or neutralize if the time of exposure is long enough.     

Arterial hypertension exacerbates oxidative stress in early diabetic retinopathy

The present study was undertaken to investigate the redox status in the retina of an experimental model that combines hypertension and diabetes. Spontaneously hypertensive rats (SHR) and their control Wystar Kyoto (WKY) rats were rendered diabetic and, after 20 days, the rats were sacrificed and the retinas collected. The superoxide production was higher in diabetic than in control WKY (p<0.03) and SHR rats showed elevated superoxide production compared with WKY groups (p<0.009). The glutathione antioxidant system was diminished only in diabetic SHR (p<0.04). Tirosyne nitration was higher in diabetic WKY and control SHR compared with control WKY (p<0.03), and further increment was observed in diabetic SHR (p<0.02). The DNA damage estimated by immunohystochemistry for 8-OHdG was higher in control SHR than in WKY, mainly in diabetic SHR (p<0.0001). Hypertension aggravates oxidative-induced cytotoxicity in diabetic retina due to increasing of superoxide production and impairment of antioxidative system.