**6. Vitamins**

High levels of serum AGE's have been documented in patients with type 2 diabetes mellitus and coronary heart disease [122]. Glycation increases susceptibility of low density lipoprotein (LDL) to oxidative modification which is considered a critical step in its atherogenicity [123]. Glycation end products can also enhance atherosclerosis by trapping LDL in the subendothe‐ lium and decrease the recognition of AGE-modified LDL by LDL receptor [124]. Modification of LDL and its increased localization in vessels increases foam cell production and accelerates atherosclerosis development [125]. Oxidative stress induces AGE's formation on collagen leading to cross-linking which is considered to play a role in diabetic cardiomyopathy [126]. The intermolecular collagen cross-linking caused by AGE increases vascular stiffness and interferes with arterial blood flow [127, 128] and this partly explains the diastolic dysfunction

**4.3. Protein kinase C (PKC) activation and ROS generation in diabetic complications**

their cell-surface receptors can result in enhanced activity of PKC isoforms [134, 135].

PKC likely regulates diabetic complications on multiple levels such as activation of eNOS, NAD(P)H oxidase, phospholipase A2 (PLA2), endothelin-1 (ET-1), Vascular endothelial growth factor (VEGF), Transforming growth factor-β (TGF-β), and by activating NF-KB. Diacylgly‐ cerol activated PKC alters the gene expression of key proteins leading to decrease blood flow, capillary occlusion, inflammation, free radicals generation and damage to cellular macromo‐

High glucose levels can stimulate ROS production via a PKC-dependent activation of NAD(P)H oxidase in cultured aortic endothelial cells, smooth muscle cells, and renal mesan‐ gial cells [84]. Nicotinamide adenine dinucleotide phosphate oxidase, which is primarily found in phagocytic cells, is the main source of ROS in non-phagocytic cells such as mesangial cells, endothelial cells [138], fibroblasts [139], podocytes [140] and smooth muscle cells [141]. The expression of NAD(P)H oxidase components is up-regulated in vascular tissues from animal models of diabetes and in patients with diabetes and coronary artery disease [142-144]. Experimental evidence indicates that NAD(P)H oxidase-dependent production of ROS may cause DNA damage in diabetic renal tissues leading to the development of nephropathy [145]. Increased activity of the NAD(P)H oxidase has also been reported in the retina of diabetic rats

suggesting its involvement in the development of diabetic retinopathy [146].

PKC activation is related to vasoconstriction, proliferation and overgrowth of smooth muscle cells as well as accelerated synthesis of extracellular matrix proteins, and thus plays significant roles in the onset and progression of vascular cell dysfunction in diabetes mellitus [129-131]. Two major pathways have been implicated in the activation of PKC in hyperglycemia. Persistent and excessive activation of several PKC isoforms result primarily from enhanced *de novo* synthesis of diacylglycerol (DAG) from glucose via increase in triose phosphate availability [90, 105, 132, 133]. There is also evidence that the interaction between AGE's and

and systolic hypertension seen in diabetic subjects.

32 Antioxidant-Antidiabetic Agents and Human Health

lecule [130-132, 136, 137].

Vitamin E is a fat-soluble vitamin. It has been shown that plasma α-tocopherol concentrations are lower in diabetics compared to controls [58] and appear to be even lower in diabetics with complications such as microangiopathy than in diabetics without complications [81]. Admin‐ istration of Vitamin E has proven to be beneficial in preventing cellular damage by inhibition of lipid peroxidation, protein oxidation, protein glycations and platelet aggregation [149-151] Vitamin E supplementation for two weeks (600 mg/day) lowered urinary F2-isoprostanes (a marker of lipid oxidation) in type 2 diabetics [152]. It was shown in a study that a decrease in plasma F2-isoprostanes was seen in type 2 diabetic patients after six weeks supplementation with Vitamin E [153].

Oxidative stress in the kidney of diabetics is usually associated with tissue damage that interferes with proper organ function, causing an increase in urinary protein excretion and blood urea nitrogen (BUN) [154]. Vitamin E supplementation (1000 IU/kg diet) to diabetic rats for 4 weeks significantly reduced urinary protein excretion and BUN suggesting a beneficial effect on kidney function [154]. Inhibitory effect of Vitamin E on glycation of hemoglobin in type I and type 2 diabetic rats has been documented [151, 155]. The ability of vitamin E to inhibit AGE's might be due to its antioxidant effect on the autoxidative pathways of AGE formation [156]. Vitamin E administration has also reduced oxidation of low density lipopro‐ tein (LDL) and development of atherosclerosis [157].

Numerous studies have shown that vitamin E normalized parameters of oxidative stress and inhibited vascular abnormalities caused by hyperglycemia-induced production of DAG and PKC activation in the retina, glomerulus and macrophages [158-160]. Supplementation with vitamin E reduced basement membrane thickening in diabetic rat retina and reduced vascular endothelial growth factor (VEGF) and aldose reductase activity, the abnormalities associated with diabetic retinopathy [161]. Dietary supplementation of vitamin E (2000 IU/kg) to diabetic rats for 8 weeks had cardioprotective effects which was simultaneously associated with an ability of vitamin E to blunt diabetes-induced amplification of myocardial 8-*iso* PGF2 and oxidized GSSG formation [162]. Clinical trials with vitamin E provided evidence that vitamin E may improve cardiovascular function [163, 164]. However, most large studies with vitamin E have not yielded positive benefits for decreasing the development or progression of diabetic microvascular and cardiovascular pathologies or mortality [165, 166].

flavonoids exhibits anti-inflammatory [181], anticarcinogenic [182], antiviral [183] and antiallergic properties. These effects are generally associated with free radical scavenging activity of flavonoids. The antioxidant effects of flavonoids are enhanced by the number and position of hydroxyl groups in the molecule. The catechol structure, presence of unsaturation and 4-oxo function in the C-ring also contributes to their radical scavenging activity [184, 185]. Flavonoids may be capable of binding the transition metal ions, which play a role in glycoxidation, thus preventing metal-catalysed formation of hydroxyl radicals or related

Oxidative Stress and Diabetic Complications: The Role of Antioxidant Vitamins and Flavonoids

http://dx.doi.org/10.5772/57282

35

Flavone Flavonol Flavanone

Isoflavone Flavan-3-ols

The potential beneficial effects of flavonoids in the prevention of diabetes mellitus and its associated complications have been investigated both *in vitro* and *in vivo* studies (Table 3). The inhibitory effect of flavonoids on glycation has been demonstrated and it is suggested that this effect is partly due to their antioxidant properties [188]. Epigallocatechin (EGC) has a beneficial effect in a rat model of diabetic nephropathy via suppressing hyperglycemia,

The potential beneficial effects of flavonoids in the prevention of diabetes mellitus and its associated complications have been investigated both *in vitro* and *in vivo* studies (Table 3). The inhibitory effect of flavonoids on glycation has been demonstrated and it is suggested that this effect is partly due to their antioxidant properties [188]. Epigallocatechin (EGC) has a beneficial effect in a rat model of diabetic nephropathy via suppressing hyperglycemia, proteinuria and lipid peroxidation. EGC also reduced renal accumulation of AGE's and their related oxidative stress [189]. Another study demonstrated the *in vitro* inhibitory effect of different flavonoids on pentosidine formation in collagen in the presence of glucose (250 mmol/L). The decreasing inhibitiory activity was observed from myricetin, quercetin, rutin, catechin and kaempferol in a structure and concentration dependent manner [190]. Kim and colleagues [191] also inves‐ tigated the effect of quercetin, isoquercitrin, hyperin and cacticin on formation of AGE's *in vitro*. At a concentration of 50 mΜ, the percentages of inhibition were 1.0, 89.6, 92.0 and 40.5, respectively. The inhibitory effect of hyperin on AGE formation was 6.5 times higher than that

flavonoids exhibits anti-inflammatory [181], anticarcinogenic [182], antiviral [183] and antiallergic properties. These effects are generally associated with free radical scavenging activity of flavonoids. The antioxidant effects of flavonoids are enhanced by the number and position of hydroxyl groups in the molecule. The catechol structure, presence of unsaturation and 4-oxo function in the C-ring also contributes to their radical scavenging activity [184, 185]. Flavonoids may be capable of binding the transition metal ions, which play a role in glycoxidation, thus preventing metal-catalysed formation of hydroxyl radicals or related

species from H2O2 [186].

**Figure 1**: Classes of flavonoids [187]

**Figure 1.** Classes of flavonoids [187]

species from H2O2 [186].

Vitamin C is an antioxidant vitamin which plays an important role in protecting free radicalinduced damage and a decrease in basal vitamin C levels has been documented in type 2 DM. Treatment of diabetic rats with vitamin C significantly decreased renal malondialdehyde, albuminuria, proteinuria, glomerular and tubulointerstitial sclerosis, suggesting the role of vitamin C in suppressing the progression of renal injury in diabetic rats [167]. Vitamin C also improved diabetes-induced endothelial dysfunction in a rat model by enhancing NO bioa‐ vailability [168].

The beneficial effects of vitamin C supplementation in humans are controversial. A study reported that vitamin C may improve glycemic control, lowering both fasting blood glucose and glycated haemoglobin (HbA1c) [169]. Chronic oral administration of vitamin C to patients with type 2 diabetes causes a decline in plasma free radicals that is associated with improved whole body glucose disposal [170,171] and improved endothelial function [172]. Recently, another study reported a reduction in the malondialdehyde (MDA) level, a major product of oxidative damage in both fasting and postprandial states of type 2 diabetic patients after vitamin C (1000 mg day-1) supplementation for 6 weeks although no effect was observed on lipid profiles [173]. Some studies have indicated that the intra-arterial infusion of vitamin c restores endothelium-dependent vasodilation in patients with type 1 or type 2 diabetes [174, 175] suggesting that hyperglycemia-induced oxidative stress mediates endothelial dysfunc‐ tion in diabetic patients.

However, in contrast to these promising results, other studies showed no beneficial effect with vitamin C treatment. Chen and colleagues [176] concluded that a high oral dose of vitamin C therapy was ineffective at improving endothelial dysfunction and insulin resistance in type 2 DM. It is important to note that complete replenishment of vitamin C levels was not achieved in the subjects. This is crucial since high concentrations of vitamin C (>80 μM) has been documented as a requirement for the preservation of NO-dependent endothelial function as vitamin C only competes with NO for superoxide anion at these high concentrations [177-178]. Also, in another study, no beneficial effects of oral vitamin C supplementation (1.5 g daily for 3 weeks) was observed on blood pressure, oxidative stress, and endothelial function in type 2 diabetes [179].
