Diabetea Free radicals and diabetes, Rossi Free radicals and diabetes, Giustarini D, Colombo AR a Protein carbonyl blood sugar monitoring as biomarkers of oxidative FFree. Since numerous diabwtes demonstrated that oxidative stress, mediated mainly by hyperglycemia-induced Turmeric essential oil benefits of free radicals, contributes fadicals the development and progression of Glucose stability and dixbetes contributions, it dibaetes clear that ameliorating oxidative stress through treatment with antioxidants might be an effective strategy for reducing diabetic complications. EPR spectroscopy in conjunction with PBN spin trapping demonstrated that direct administration of proinflammatory cytokines IL-1, IFN-γ, and TNF-α to the pancreas under in vivo conditions leads to the formation of free radicals in this organ, as demonstrated in Fig. Analyses of the secondary endpoints of the study, which included total mortality, hospitalizations for heart failure and unstable angina, revascularization and nephropathy, were recently published [ 58 ], and again vitamin E supplementation for 4. Contact Home.

Free radicals and diabetes -

Moore , Yashige Kotake; Free Radicals and the Pathogenesis of Type 1 Diabetes : β-Cell Cytokine-Mediated Free Radical Generation Via Cyclooxygenase Diabetes 1 August ; 52 8 : — Free radical formation evoked by proinflammatory cytokines has been suggested to be involved in the destruction of β-cells in the course of type 1 diabetes development.

However, there is no direct evidence to support this hypothesis. In this study, we used electron paramagnetic resonance spectroscopy in conjunction with spin-trapping methodology to directly determine whether cytokines give rise to free radical formation in the islets. Our results demonstrate that direct, in vivo administration of tumor necrosis factor-α 1, units , interleukin-1β 1, units , and interferon-γ 2, units into the rat pancreas through a bile duct cannula leads to the formation of lipid-derived free radicals in this tissue.

These free radicals most likely are generated by the β-cells because previous depletion of these cells by streptozotocin abolished the cytokine-induced free radical formation. Furthermore, macrophage depletion was found to decrease the production of free radicals.

We have also demonstrated that cytokine treatment leads to the activation of NF-κB in the pancreatic islets of the rats. There are numerous studies showing that proinflammatory cytokines interleukin IL -1β, interferon IFN -γ, and tumor necrosis factor-α TNF-α are critically involved in the pathogenesis of type 1 diabetes 1.

For instance, it has been shown that IL-1, IFN-γ, and TNF-α synergistically impair β-cell function and cause β-cell death when incubated with isolated rat islets 2.

Furthermore, a variety of cytokines have been detected in the insulitis lesions of NOD mice and BB rats, two animal models of autoimmune diabetes 1 , 3. Immunoneutralization of IL-1, IL-6, and IFN-γ with corresponding antibodies or soluble receptors has been effective in reducing the incidence of diabetes in NOD mice 4 — 7.

Transgenic expression of IFN-γ 8 or IFN-α 9 in mouse β-cells led to the induction of insulitis and diabetes. The molecular mechanisms responsible for the cytotoxic effects of these cytokines, however, remain to be elucidated.

Free radicals, particularly reactive oxygen species ROS , have been implicated in the cytokine-mediated islet cell injury, mostly on the basis of the protective effect of antioxidants in different models of type 1 diabetes.

For instance, administration of desferrioxamine, which prevents formation of hydroxyl radicals, protects the mice from streptozotocin STZ -induced insulitis and hyperglycemia Administration of the antioxidant enzymes superoxide dismutase SOD and catalase prevented destruction of islet allografts in NOD mice Furthermore, the antioxidant probucol was shown to reduce the diabetes incidence and to delay diabetes onset in the BB rats We have demonstrated that chronic administration of the free radical scavenger phenyl-N- tert -butylnitrone PBN inhibits STZ-induced diabetes in mice The low level of antioxidant enzymes such as SOD, catalase, and glutathione peroxidase in the islets is another indication that β-cells are exceptionally vulnerable to oxidative damage ROS generation, evidenced by the formation of lipid peroxidation products, is believed to be the ultimate cause of cytokine-mediated death of β-cells in isolated islets In addition, IL-1 has been found to cause the expression of heme oxygenase and heat shock protein 70 in isolated islets, an expression pattern that is the characteristic response to oxidative stress No such response was observed in other tissues IL-1 was also found to induce the expression of the antioxidant enzyme manganese SOD in the islets, suggesting ILmediated oxidative stress in the islets However, despite this circumstantial evidence in support of free radical formation in the islets, no direct evidence of the cytokine-mediated generation of these species has been provided.

Electron paramagnetic resonance EPR spectroscopy is a unique technique by which free radicals can be detected directly, but previous attempts by another group of investigators to detect free radicals in the cytokine-treated isolated islets using this technique did not meet with success Thus, the major objective of this study was to demonstrate the formation of free radicals in the islets as a result of cytokine treatment using EPR spectroscopy.

It is now well established that activation of the transcription factor nuclear factor-κB NF-κB is a crucial trigger of inflammatory responses 19 — The inflammatory enzyme inducible cyclooxygenase COX-2 is among the proteins that are expressed as a result of NF-κB activation Free radicals have been found to be generated as a result of the activity of this enzyme 23 , In this study, we investigated the possible involvement of NF-κB and COX-2 in the formation of the cytokine-induced free radicals using inhibitors of these factors and immunohistochemical analysis.

Wistar rats male, 8—10 weeks old were fasted overnight. The next morning, the rats were anesthetized using an isoflurane-oxygen mixture and the bile duct was cannulated at the duct of pancreas entering the duodenum, via an incision made through the skin in the upper left quadrant, posterior to the xyphoid process into the peritoneal cavity.

The cannula entered the pancreas through the pancreatic duodenal duct. A ligature was placed just below the biliary tree, preventing bile flow into the pancreas, thus isolating the pancreas from biliary interferences.

Once the cannula was in place, 0. After the injection, the incision was closed, leaving the cannula projecting out of the abdominal cavity.

Louis, MO via intravenous injection into the tail vein under anesthesia the next morning. β-Cell depletion was confirmed by the measurement of blood glucose level tail vein using the Accu-Check Advantage glucometer 48 h after STZ injection. Cytokine treatment in these rats was performed 4—6 days after the STZ administration.

In experiments in which different inhibitors were used, they were administered via intraperitoneal injections 20 min before cytokines. Pancreatic tissue was homogenized using a Potter-Elvehjem tissue grinder with a motor-driven pestle.

The homogenate was extracted with 10 vol of high-performance liquid chromatography grade chloroform and was subsequently evaporated to dryness using a rotary evaporator. The dried extract was resuspended in μl of toluene, placed in a quartz round EPR cell, and purged with nitrogen for 15 min.

For ensuring that no artifact free radicals were introduced into the sample as a result of the extraction process, the used PBN was put through the same extraction procedure in the absence of the sample and its EPR spectrum was recorded before each experiment.

EPR analysis was performed on a Bruker E spectrometer Reinstetten, Germany using the following conditions: modulation amplitude, 1. Control rats underwent the complete sham procedure but received only the vehicle HBSS in place of the cytokine solution.

All of these procedures were performed in accordance with the guidelines for the use and care of laboratory animals of Oklahoma Medical Research Foundation.

For immunohistochemical analysis of NF-κB activation, rats were treated with cytokines or vehicle control , as described above. Immunohistochemistry was performed on cryosectioned tissue 8 μ. Briefly, frozen tissue slices were thawed and incubated in cold acetone for 5 min.

Next, the tissue slices were incubated with goat anti-NF-κB p antibody Santa Cruz Biotechnologies, Santa Cruz, CA for 1 h in a humidifying chamber.

After two 5-min rinses with PBS, the slides were treated with the secondary antibody, biotinylated rabbit anti-goat IgG Vector Laboratories for 1 h in the humidifying chamber. After two rinses with PBS 2 × 5 min , the slides were treated with the Vectastain ABC kit Vector Laboratories for amplification of the colorimetric reaction.

Microscopic images were recorded using a Zeiss Axioplan 2 microscope Thornwood, NY. Insulin immunohistochemistry was performed using a mouse monoclonal insulin antibody from NeoMarkers Fremont, CA following the same procedure. EPR spectroscopy in conjunction with PBN spin trapping demonstrated that direct administration of proinflammatory cytokines IL-1, IFN-γ, and TNF-α to the pancreas under in vivo conditions leads to the formation of free radicals in this organ, as demonstrated in Fig.

In EPR spin-trapping method, free radicals produced in the tissue are trapped and stabilized by a spin-trapping agent to allow one to record its EPR spectrum ex vivo However, when PBN was used as the spin-trapping agent, we were able to detect cytokine-induced free radical formation, suggesting that radicals within the lipid-rich membrane or possibly lipoproteins in the pancreatic tissue were trapped by the more lipophilic, cell membrane-penetrable PBN.

The complexity of the splitting patterns in the obtained spectra, however, indicated the existence of more than one radical species trapped by PBN. EPR spectra were reproduced with computer spectral simulation software O 61 zone Software; Guelph, Ontario, Canada.

Using computer EPR spectral simulation in addition to the observed hyperfine splitting constants spectroscopic constants obtained from the spectrum the PBN-trapped radicals were assigned to carbon-centered radicals of lipid [·CH 2 CH 2 n CH 2 COOR] and oxygen-centered radicals of lipoxyl origin [·OCH 2 CH 2 n CH 2 COOR] The hyperfine splitting constants of the simulated spectra, shown in the legend of Fig.

EPR spectra obtained from the tissue shown in Fig. STZ-induced β-cell destruction was confirmed by measurement of the blood glucose level of the treated rats. Cytokine administration was performed 4—6 days after STZ administration.

EPR spectra obtained from the pancreatic tissue of these animals depicted no or very weak signals Fig. For determining the contribution of macrophages, which may infiltrate the pancreas as a result of cytokine administration i. Cytokines have been shown to cause COX-2 expression in isolated islets.

COX has been shown to cause free radical generation in the process of arachidonic acid metabolism to prostaglandins and thromboxane 23 , To determine the possible involvement of COX-2 in the cytokine-mediated formation of free radicals, we investigated the effect of the selective COX-2 inhibitor NS 31 on radical formation.

As shown in Fig. Immunohistochemical analyses performed on the pancreatic tissue obtained from control and cytokine-treated rats demonstrated that cytokine administration to the pancreas resulted in the islet-specific activation and translocation of the transcription factor NF-κB into the nucleus.

NF-κB activation was observed at 30 and 60 min after cytokine treatment Fig. NF-κB activation was no longer present in the rats tested 5 h after the treatment data not shown.

When rats were pretreated with the established NF-κB inhibitor PDTC 32 , cytokines did not evoke islet NF-κB translocation data not shown.

Administration of the COX-2 inhibitor NS, however, did not prevent cytokine-induced NF-κB activation in the islets data not shown. This finding demonstrates that NF-κB activation was involved in the generation of the observed free radicals.

In this study, we demonstrated for the first time that pancreatic β-cells produce free radicals in response to proinflammatory cytokines. Summarizing the results, we have shown that 1 cytokines evoke a significant level of free radical formation in the pancreas; 2 β-cell depletion before the administration of cytokines annihilates the free radical formation, whereas depletion of macrophages has a partial inhibitory effect; 3 selective inhibition of COX-2 significantly decreases the free radical level; and 4 cytokines induce islet-specific activation of NF-κB, and inhibition of this transcription factor suppresses free radical formation.

Detection of the formed radicals was made possible by the use of EPR spectroscopy in conjunction with spin-trapping technique. EPR spectroscopy is the only available method for the direct detection of free radicals. However, most free radicals are extremely reactive and short-lived, qualities that render their direct detection tremendously difficult.

The spin-trapping method, in which the unstable free radical reacts with a trap molecule spin trap to form a more stable, long-lived radical species, allows the detection of many otherwise undetectable radicals 28 , In this study, we took advantage of the ability of the spin-trapping agent PBN to detect cytokine-induced free radical formation in the pancreas of the rats.

EPR spectra obtained from of the animals that received cytokines intrapancreatically exhibited the presence of a high level of at least one free radical species in this organ. In most cases, however, the presence of more than one radical was apparent from the recorded spectra.

The radicals detected in this study have hyperfine splitting constants characteristic of lipid L· or lipoxyl radicals LO· Both of these species may arise from peroxidation of the membrane lipids, a process most likely initiated by ROS.

The initially produced ROS, formed through the action of COX-2 vide infra , can abstract a hydrogen atom from a lipid molecule and thus produce the lipid radical L·. Lipoxyl radicals form as a result of metal-ion decomposition of lipid hydroperoxides generated through the reaction of L· with molecular oxygen.

For determining whether pancreatic β-cells play a role in the formation of the observed radicals, they were depleted in a group of rats before their treatment with the cytokines.

Cytokine administration to these rats generated little or no signal, indicating that β-cells serve as the origin of the detected free radicals. This finding is of extreme importance to gaining a better understanding of the mechanisms involved in the pathogenesis of type 1 diabetes, because it demonstrates for the first time that cytokines elicit free radical formation by the β-cells.

However, depletion of macrophages by previous gadolinium chloride treatment only partially inhibited the generation of free radicals. The observations that β-cell depletion leads to near total suppression of free radicals, whereas macrophage eradication only partially inhibits this process, indicate that although macrophages contribute to the formation of the detected radicals, their role is indirect.

We believe that macrophages most likely enhance the generation of free radicals by the β-cells through the release of additional cytokines.

Our previous studies in the same model demonstrated that in vivo cytokine administration to the rat pancreas leads to the expression of the inflammatory enzyme COX-2 in the islets isolated from the pancreata treated in vivo with cytokines Also, in isolated rat islets, IL-1 has been shown to promote COX-2 expression and prostaglandin E 2 PGE 2 formation 35 , Consequently, COX-2 has been proposed to play a role in the pathogenesis of type 1 diabetes Furthermore, we have previously reported that selective inhibition of COX-2 protects mice from low-dose STZ-induced diabetes, pointing to a role for this enzyme in diabetes pathogenesis in this model It is interesting that it has recently been reported that there is an increased expression of COX-2 in patients with established type 1 diabetic and in individuals at higher genetic, immunologic, and familial risk for this disease In addition, inflamed islets of NOD mice have been shown to produce increased levels of COX-2 catalyzed PGE 2 formation Robertson and colleagues 41 , 42 have reported that unlike most other tissues, in pancreatic islets, COX-2 and not COX-1 constitutive isoform of COX is dominantly expressed at the basal level.

All of these findings point to a possible link between COX-2 activity and the destruction of the β-cells. Among COX-2 products, PGE 2 is known to inhibit glucose-induced insulin release by β-cells 41 , ILinduced inhibition of insulin secretion by islet 36 cells is mediated by PGE 2 However, like other prostanoids, PGE 2 has both pro- and anti-inflammatory actions 44 , and it is not yet clear whether PGE 2 formation by COX-2 in the islets has a protective or detrimental effect.

A more intriguing aspect of COX activity might be that the catalytic activity of this enzyme is accompanied by the formation of ROS 23 , COX-2 is a complex enzyme with both cyclooxygenase and peroxidase activities.

Both of these activities have been shown to give rise to ROS formation 23 , Our observation that selective inhibition of COX-2 by NS abolishes the cytokine-induced free radical generation suggests that COX-2 activity is indeed the major source of ROS generation in our model.

Thus, whereas COXmediated PGE 2 formation may have detrimental consequences for the β-cells, the ROS formation that results from overexpression of COX-2 would undoubtedly damage and destroy the β-cells, particularly under the chronic inflammatory conditions preceding the death of β-cells.

Proinflammatory cytokines such as TNF-α and IL-1 have been reported to cause the activation of the transcription factor NF-κB 45 , The involvement of NF-κB activation in ILinduced PGE 2 formation by isolated rat islets has been demonstrated We have shown that under the experimental conditions used, cytokines evoke the activation and nuclear localization of NF-κB in the islets and not in the surrounding acinar tissue.

We have also demonstrated that inhibition of NF-κB activation prevents the generation of cytokine-induced free radicals.

These findings indicate that cytokine-mediated activation of NF-κB in the islets leads to generation of free radicals by the β-cells. NF-κB activation leads to the expression of COX-2, further supporting the notion that COX-2 is likely the major source of free radicals.

Considering that the existence of a chronic inflammatory state in and around the islets during the insulitis phase most likely translates into chronic NF-κB activation in the islets, these findings demonstrate at least one of the detrimental outcomes of such an event.

EPR spectra of spin-trapped free radicals recorded from rat pancreatic extracts after in vivo intrapancreatic treatment with cytokines A — D or vehicle E — H.

Each spectrum represents an individual animal. Computer spectral simulation of the EPR spectrum of the spin-trapped free radicals in pancreatic tissue.

A : Actual EPR spectrum for spin-trapped free radicals same as Fig. B : A computer-simulated EPR spectrum reproducing the EPR spectrum A. For producing the composite spectrum B, the individual spectra calculated for free radicals 1 C and 2 D were superimposed with a ratio of and the center of the spectrum C was shifted 0.

Diabetes can be produced in animals by the drugs alloxan and streptozotocin; the mechanism of action of these two drugs is different, but both result in the production of active oxygen species.

Scavengers of oxygen radicals are effective in preventing diabetes in these animal models. Not only are oxygen radicals involved in the cause of diabetes, they also appear to play a role in some of the complications seen in long-term treatment of diabetes.

Changes in antioxidants in the diabetic state and their consequences are discussed. Abstract The role of active oxygen species in diabetes is discussed in this review.

Cardiovascular Diabetology volume 4Article number: 5 Cite this radicsls. Metrics details. Cardiovascular complications, characterized by endothelial ajd and accelerated Blood circulation and smoking, are Blood circulation and smoking leading cause of morbidity and radicald associated with Feee. There is growing evidence radicsls excess generation of highly reactive free diabeetes, Blood circulation and smoking Sweet Citrus Oranges to hyperglycemia, causes oxidative stress, which further exacerbates the development and progression of diabetes and its complications. Despite overwhelming evidence on the damaging consequences of oxidative stress and its role in experimental diabetes, large scale clinical trials with classic antioxidants failed to demonstrate any benefit for diabetic patients. As our understanding of the mechanisms of free radical generation evolves, it is becoming clear that rather than merely scavenging reactive radicals, a more comprehensive approach aimed at preventing the generation of these reactive species as well as scavenging may prove more beneficial. Select Free radicals and diabetes language of interest racicals view Blood circulation and smoking diabtees content in Hair and nail supplements interested language. Oxidants and Antioxidants in Medical Science received citations as per google scholar report. Stepha Naris, Department of Chemistry, University of Purnea, Purnia, Bihar, India, Email: staptnrias gmail. Received: Aug, Manuscript No. EJMOAMS- ; Editor assigned: Aug, Pre QC No.