**2.1. Increased polyol pathway flux**

**2. Mechanisms by which diabetes increases cardiovascular disease**

responsible for hyperglycemic damage [5] (**Figure 1**).

34 Diabetes Food Plan

All forms of diabetes are characterized by chronic hyperglycemia and the development of diabetes-specific macrovascular disease affecting the coronary arteries. Large prospective clinical studies show a strong correlation between hyperglycemia, insulin resistance and diabetic macrovascular complications in both type 1 and type 2 diabetes mellitus [5]. Five major molecular mechanisms have been implicated in hyperglycemia-induced tissue damage [6]: (1) increased polyol pathway flux, (2) increased advanced glycation end products (AGEs), (3) activation of protein kinase C (PKC), (4) increased hexosamine pathway flux, and (5) activation of the 12/15-lipoxygenase (12/15-LO) pathway [5]. Hyperglycemia-induced overproduction of superoxide is the causal link between high glucose concentration and the pathways

**Figure 1.** Pro-atherogenic mechanisms of diabetes associated with hyperglycemia. Four hyperglycemia-related mechanisms may promote diabetic atherosclerosis: (1) the polyol pathway, (2) formation of advanced glycation end products (AGEs), (3) activation of protein kinase C (PKC) isoforms, (4) the 12/15-lipoxyenase pathway, and (5) the hexosamine pathway. All four mechanisms result in increased formation of reactive oxygen species (ROS) and promote diabetic atherosclerosis by various mechanisms as depicted in the figure. Boxes in arrows, cells and ECM indicate relevant pathway. 12/15-LO = 12-/15-lipoxygenase, AR = aldose reductase, EC = endothelial cell, ECM = extracellular matrix, Fruc = fructose, GFAT = glutamine-fructose-6-phopshate amidotransferase, Glc = glucose, Mo = monocyte, Mφ = macrophage, RAGE = receptor for advanced glycation end products, SDH = sorbitol dehydrogenase, VSMC = vascular smooth muscle cell, other abbreviations are explained in the text. Reprinted with permission from [5].

Aldose reductase (alditol:NADP+ 1-oxidoreductase) is a cytosolic NADPH-dependent oxidoreductase that catalyzes the reduction of glucose to sorbitol, which is further processed to fructose [7]. Aldose reductase (AR) has a low affinity (high *K*m) for glucose and, under euglycemic conditions, this pathway plays a minor role in glucose metabolism [6]. Excess glucose is also channeled into the accessory polyol pathway, where it is reduced to polyalcohol sorbitol by AR, an NADPH-dependent enzyme [8]. In the polyol pathway, sorbitol is oxidized to fructose by sorbitol dehydrogenase, with NAD+ reduced to NADH. Under hyperglycemia, this pathway can account for 25–30% of total glucose metabolism [9]. Overexpression of human AR in low-density lipoprotein (LDL) receptor (LDLR) deficient mice resulted in increased atherosclerotic lesion size if mice became diabetic by administration of streptozotocin (STZ) [5, 10]. Atherosclerotic lesions in normoglycemic LDLR−/− did not differ significantly between AR-overexpressing mice and mice with normal AR expression [11]. Long-term polyol pathway activation also increased intimal thickening in dog coronary arteries, an effect that could be blunted by AR inhibition [12]. Polyol pathway activation also triggered abnormalities in endothelium-dependent relaxation in aortas from STZ-diabetic rats and decreased nitric oxide (NO) release and functionality [13, 14].

#### **2.2. Increased intracellular formation of advanced glycation end products (AGEs)**

One of the important mechanisms responsible for accelerated atherosclerosis in diabetes is the Maillard reaction—a type of non-enzymic browning which involves the reaction of carbonyl compounds, especially reducing sugars, with compounds which possess a free amino group, such as amino acids, amines, and proteins [15]. This reaction is subdivided into three main stages. In an early stage, the protein glycation process starts with a nucleophilic addition between free ε-amino or NH<sup>2</sup> -terminal groups of proteins and the carbonyl group of reducing sugars (normally glucose or glyceraldehyde) to form a reversible Schiff base [16]. By structural irreversible rearrangements, more Amadori products—stable keto-amines—are formed (i.e., hemoglobin A1c (Hb A1c) [17]. In an intermediate stage, breakdown of Amadori products results in a variety of reactive dicarbonyl compounds such as glyoxal, methylglyoxal, and deoxyglucosones. In the late stage of glycation due to oxidation, dehydration, and cyclization reactions, irreversible compounds called AGEs are formed [18]. AGEs act either by modifying substrates, or by interacting with specific receptors [16]. AGEs-induced damage can occur to the vasculature, vascular cells, and cells implicated in vascular homeostasis via at least the following 4 mechanisms [19, 20]: (1) AGEs modify intracellular proteins, including those involved in the regulation of gene transcription; (2) precursors of AGEs leave the cells via diffusion and modify nearby extracellular matrix molecules, subsequently altering the signaling between matrix and cells and ultimately causing cellular dysfunction; (3) AGEs and their precursors modify circulating proteins in the bloodstream, thereby altering their function; (4) circulating proteins modified by AGEs bind to and activate AGE receptors, thereby altering the production of inflammatory cytokines and growth factors and causing tissue damage [19, 20].

The deleterious effects of AGEs on the vasculature can also be classified either as follow:

#### *2.2.1. Receptor-independent effects of AGEs*

Collagen in the blood vessel wall has a relatively long biological half-life, and with time undergoes significant non-enzymatic glycation, which may have a considerable bearing on atherosclerosis [21]. Soluble plasma proteins, such as low-density lipoprotein cholesterol (LDL-C) and immunoglobulin G (IgG), are also entrapped and covalently cross-linked by AGEs on collagen [20, 22]. Glycation of LDL-C decreases recognition of LDL-C particles by the LDL-receptor and enhances the uptake of LDL-C by a low-affinity high-capacity receptor pathway on macrophages. Decreased LDLR affinity of glycated LDL-C may result in increased oxidation of particles and may sufficiently alter their structure to render them immunogenic [23]. Glycated LDL-C is more susceptible to oxidative modification than non-glycated LDL-C. Being immunogenic, glycated LDL-C accumulates in plasma and may enhance cholesterol ester accumulation in macrophages and thus may increase the risk of atherogenic complications [23]. Glycation of apolipoprotein A1 (Apo-AI), the major protein of the protective HDL-C (high-density lipoprotein cholesterol) complex is increased in T2DM and has been shown to induce conformational changes and decreased stability of the lipid-protein interaction, as well as a reduction in the ability of the lipoprotein to self-associate [24, 25]. HDL-C glycated in vitro and Apo-AI isolated from diabetic subjects show decreased ability to activate lecithin-cholesterol acyltransferase, which drives reverse cholesterol transport by esterifying the cellular cholesterol removed by HDL-C [26, 27]. In human aortic endothelial cells, glycated and glycoxidized HDL-C induces H2 O2 formation, dampens the expression of endothelial nitric oxide synthases (eNOS) decreases NO production, promotes apoptosis associated with increased caspase 3 expression, attenuates caspase 3 inhibition, and increases release of cytochrome c into the cytosol [28, 29].

group is named after the protein kinase A, G, and C families that are closely related to the cAMPdependent protein kinase [36]. Twelve PKC isoforms have thus far been identified, which differ in terms of structure and substrate requirements [37]. Eight isoforms are activated by diacylglycerol (DAG) [6, 38]. Hyperglycemia can contribute to the direct and indirect production of ROS via the activation of the DAG-PKC pathway [6, 38]. Indirect PKC activation may be due to RAGE engagement or polyol pathway activation or activation of the12/15-lipoxygenase (12/15-LO) pathway [39]. Increased PKC levels associated with diabetes are found in several tissues including the aorta and the heart [40, 41]. Higher PKC activation triggers hyperglycemia-induced cardiometabolic perturbations such as changes in blood flow, basement membrane thickening, vascular permeability, angiogenesis, cell growth, and enzymatic activity alterations [42, 43]. PKC activation directly increases the permeability of albumin and other macromolecules through

the development of cardiac hypertrophy and fibrosis, which eventually leads to left ventricular dysfunction suggesting that PKCβ may play a central role in the development of diabetic cardio-

malities via inhibition of Akt (protein kinase B)-dependent endothelial nitric eNOS activity [48]. Restoration of Akt-eNOS-NO signaling has been shown to attenuate DCM and myocardial dysfunction [49]. Quantitative immunoblotting revealed a significant increase in membrane fraction expression of PKC-β1 and -β2 in failed human hearts [50]. Among the processes induced by hyperglycemia, activation of PKC may contribute to DCM by inhibiting the metabolic actions of insulin [51]. The PKC-β inhibitor ruboxistaurin (LY333531) is a class of bisindolylmaleimide

diastolic dysfunction in rats with STZ-induced diabetes. LY333531 suppresses the decreased expression of myocardial NO and phosphate endothelial eNOS [53]. Peroxisome proliferatoractivated receptors gamma (PPARs-γ), could directly affect vascular function because of their

The hexosamine biosynthesis pathway (HBP) is another side branch of glycolysis [56]. The reaction in which glucose 6-phosphate is changed to fructose 6-phosphate is catalyzed by glutamine fructose-6-phosphate amidotransferase (GFAT) [57]. The major product of HBP is UDP-N-acetylglucosamine (UDP-GlcNAc) [57]. UDP-GlcNAc regulates flux through HBP by regulating GFAT activity and is the obligatory substrate of O-GlcNAc transferase [57, 58]. Hyperglycemia stimulates the expression of PAI-1 in smooth vascular muscle cells and aortic endothelial cells. This effect is thought to be an important factor in the development of vascular disease in diabetes [59, 60]. Sp1 (a protein that in humans is encoded by the SP1 gene) was the first transcription factor identified as an O-GlcNAc modified protein [60]. It has multiple O-GlcNAc modification sites, and its phosphorylation on serine–threonine is inversely proportional to its O-GlcNAc modification [57, 61]. The glycosylated form of Sp1 seems to be more transcriptionally active than the deglycosylated form [62]. The major mechanism of glucose toxicity is the increased mitochondrial superoxide production; this event can account for the diverse manifestations in vascular cells, i.e., increased polyol pathway flux, increased AGE products, activation of PKC, and increased HBP [6, 63]. Inhibition of the rate-limiting

and PKCβ<sup>2</sup>

activation has been implicated in diabetes-associated abnor-

Dietary Recommendations for Patients with Cardiovascular Disease and Diabetes

are two of the classical isoforms

http://dx.doi.org/10.5772/intechopen.71391

37

activation and attenuates cardiac

overexpression and activation facilitates

barriers formed by endothelial cells [44]. PKCβ1

myopathy (DCM) [46, 47]. PKCβ<sup>2</sup>

(α, β, and γ) of PKC [45]. Of the two isoforms, PKCβ<sup>2</sup>

[52]. In vivo LY333531 treatment prevents excessive PKCβ<sup>2</sup>

**2.4. Increased glucose flux through the hexosamine pathway**

expression in endothelial cells and smooth vascular muscle cells [54, 55].
