Diabetic stress: new triumphs and challenges to maintain vascular longevity

Diabetes mellitus (DM) is present in more than 165 million individuals worldwide and has increasingly become a significant health concern, especially regarding vascular and cardiac disease [1]. By the year 2030, it is predicted that more than 360 million individuals will be afflicted with DM. At least 80% of all diabetics have Type 2 DM, which is increasing in incidence as a result of changes in human behavior that relate to diet and daily exercise. Although Type 1 insulin-dependent DM accounts for only 5–10% of all diabetics, it is increasing in adolescent minority groups. Of potentially greater concern is the incidence of undiagnosed diabetes, which consists of impaired glucose tolerance and fluctuations in serum glucose that can increase the risk for the development of DM. Both acute and long-term complications of Type 1 and Type 2 DM can result in complications of the vascular and cardiac systems. For example, DM can impair vascular integrity and alter cardiac output that eventually diminishes the capacity of sensitive cognitive regions of the brain, leading to functional impairment and dementia [2,3].

Diabetes & oxidative stress

Although a number of mechanisms can account for the degeneration of vascular cells in DM, the generation of cellular oxidative stress represents a significant component of the pathological complications that can occur with DM. Initial work in this field by early pioneers observed that increased metabolic rates could be detrimental to animals in an elevated oxygen environment. More current studies outline potential aging mechanisms and accumulated toxic effects for an organism that is tied to oxidative stress. The effects of oxidative stress are linked to the generation of oxygen free radical species in excessive or uncontrolled amounts during the reduction of oxygen. These oxygen free radicals are usually produced at low levels during normal physiological conditions and are scavenged by a number of endogenous antioxidant systems, such as superoxide dismutase, glutathione peroxidase, catalase and small molecule substances, such as vitamins C, E, D₃ and B₃[4].

However, free radical species that are not scavenged can ultimately lead to cellular injury and programmed cell death, also known as apoptosis. Interestingly, it has recently been shown that genes involved in the apoptotic process are replicated early during processes that involve cell replication and transcription, suggesting a much broader role for these genes than originally anticipated. Apoptosis-induced oxidative stress can contribute to a variety of disease states, such as diabetes, and lead to the impairment or death of vascular endothelial cells. Membrane phosphatidylserine (PS) externalization is an early event during cell apoptosis and involves the translocation of membrane PS residues from the inner cellular membrane to the outer surface, which is a necessary component under most conditions for the removal of apoptotic cells [5]. The loss of membrane phospholipid asymmetry has also been associated with clot formation in the vascular cell system. In contrast to the early externalization of membrane PS residues, the cleavage of genomic DNA into fragments is considered to be a later event during apoptotic injury. During oxidative stress, mitochondrial membrane transition pore permeability is also increased, a significant loss of mitochondrial NAD⁺ stores occurs and further generation of superoxide radicals leads to cell injury [6,7].

In disorders such as DM, elevated levels of ceruloplasmin have been suggested to represent increased reactive oxygen species and acute glucose fluctuations have been described as a potential source of oxidative stress [3]. Elevated serum glucose has also been shown to lead to increased production of reactive oxygen species in endothelial cells, but a prolonged duration of hyperglycemia will not necessarily lead to oxidative stress injury, since even short periods of hyperglycemia can generate reactive oxygen species in vascular cells. Recent clinical correlates support these experimental studies to demonstrate that acute glucose swings, in addition to chronic hyper-glycemia, can trigger oxidative stress mechanisms during Type 2 DM, illustrating the importance for therapeutic interventions during acute and sustained hyperglycemic episodes [8]. The maintenance of cellular energy reserves and mitochondrial integrity also becomes a significant factor in DM. During DM, fatty acid accumulation leads to both the generation of reactive oxygen species and mitochondrial DNA damage. Insulin resistance in the elderly has also been associated with an elevation in fat accumulation and a reduction in mitochondrial oxidative and phosphorylation activity [1,7].

Therapeutic strategies for diabetes & vascular disease

The potential pathways that can lead to diminished vascular longevity during DM are far reaching and involve multiple precipitating factors. Yet, oxidative stress-induced cellular signaling is believed to be one of the important factors responsible for cell injury that initially is set in motion following hyperglycemia. For example, studies shave shown that administration of insulin or insulin growth factor at concentrations that were insufficient to reverse hyperglycemia could nevertheless reduce oxidative stress injury to cells and maintain mitochondrial inner membrane potential [1,3]. As a result, cellular strategies that directly target the reduction of oxidative stress toxicity to vascular cells could offer viable therapeutic options to patients with DM, in addition to more conventional treatments to regulate serum glucose levels.

Erythropoietin

Treatment with agents that function as antioxidants during high glucose levels has been shown to inhibit free radical production, as well as prevent the production of advanced glycation end products known to produce reactive oxygen species and cellular injury during DM. Agents that can modulate oxidative stress and the injury of vascular cells offer attractive candidates for novel therapeutic strategies in DM. As a result, the 30.4-kDa glycoprotein erythropoietin (EPO) appears to fill such criteria. Although EPO is currently approved for the treatment of anemia, the role of this trophic factor and cytokine has become far more reaching and has dispensed with initial beliefs that EPO is required only for erythropoiesis [9]. In clinical studies with DM, plasma EPO is often low in diabetic patients whether or not anemia is present and is believed to have limited response to progressive anemia onset in diabetics [10]. Interestingly, EPO secretion is upregulated in diabetic pregnancies, which may suggest the body’s attempt to protect against the complications of DM. Treatment with EPO has been shown in diabetic patients with severe, resistant congestive heart failure to decrease fatigue, increase left ventricular ejection fraction and significantly decrease the number of hospitalization days [11]. In studies that examine the primary toxic effects of elevated glucose upon vascular cells, EPO prevents early apoptotic membrane PS exposure and late DNA degradation in vascular cells at concentrations that are clinically applicable to the cytoprotection associated with EPO during cardiac and renal disease. Vascular protection by EPO is closely tied to protein kinase B (Akt) and the maintenance of mitochondrial membrane potential to prevent cell injury and the subsequent induction of the apoptotic cascades [12–14].

Interestingly, EPO may require other substrates of the Akt pathway, such as the forkhead transcription factor FOXO3a, to prevent vascular cell compromise during DM. The mammalian forkhead transcription factor family functions as transcription factors involved in pathways responsible for cell metabolism, DM onset and diabetic complications. These family members contain over 100 forkhead genes and 19 human subgroups that are known to exist that extend from FOXA to FOXS [15]. In a prospective population-based study, haplotype analyses of FOXO1a revealed that carriers of haplotype 3 ‘TCA’ had higher hemoglobin A_1c levels to suggest evidence of at least an association of forkhead transcription factors with glucose intolerance [16]. Other work suggests that the forkhead proteins may confer resistance against oxidative stress for some cell populations, such as hema-topoietic stem cells [15]. In a similar manner for FOXO1, this transcription factor may be responsible for the prevention of β-cell injury during oxidative stress. Yet, the role of forkhead transcription factors can vary among different cells and tissues. For example, mice overexpressing FOXO1 in skeletal muscle suffer from reduced skeletal muscle mass and poor glycemic control [17] and prevention of FOXO3a activation by EPO during oxidative stress protects against vascular cell injury [13].

Nicotinamide

As the amide form of niacin or vitamin B₃, nicotinamide also plays a critical role in cellular metabolism and can offer significant vascular cell protection during a wide range of disorders that include DM. For example, nicotinamide can improve glucose utilization and prevent excessive lactate production in ischemic animal models [4]. With regards to vascular cell protection, nicotinamide can maintain endothelial cell membrane integrity during oxygen radical exposure and nicotinamide is believed to be responsible for the preservation of endocardial endothelial cell integrity during models of oxidative stress and to assist with left ventricular cardiac function [4]. Treatment with nicotinamide may be effective during DM, since nicotinamide can maintain normal fasting blood glucose in animals with streptozotocin-induced diabetes. Oral application of nicotinamide can also protect β-cell function and prevents clinical disease in islet-cell antibody-positive first-degree relatives of Type 1 DM [18]. In addition, treatment with nicotinamide in patients with recent-onset Type-1 DM combined with intensive insulin therapy for up to 2 years after diagnosis significantly reduced HbA_1c levels [19]. Potentially relevant to diabetic patients with renal failure, nicotinamide has also been shown to reduce intestinal absorption of phosphate and prevent the development of hyperphosphatemia and progressive renal dysfunction.

Wnt signaling pathways

Wnt proteins are secreted cysteine-rich glycosylated proteins that can be dependent upon Akt signaling and oversee embryonic cell proliferation, differentiation, survival and death [20]. Wnt signaling can prevent cell injury through a variety of mechanisms. Wnt prevents apoptosis through β-catenin/Tcf transcription-mediated pathways and can also protect cells during oxidative stress that may require the modulation of glycogen synthase kinase (GSK)-3β and β-catenin [5,21]. Abnormalities in the Wnt signaling pathways, such as with transcription factor 7-like 2 gene, may impart increased risk for Type 2 DM in some populations as well as having an increased association with obesity [1,3]. Yet, intact Wnt family members may offer glucose tolerance and increased insulin sensitivity as well as protect glomerular mesangial cells from elevated glucose-induced apoptosis [22]. New in vitro studies demonstrate that the Wnt1 protein is necessary and sufficient to impart cellular protection during elevated glucose exposure and is a vital component for vascular protection provided by EPO [23].

With regards to Wnt signaling and the pathways of GSK-3β, inactivation of GSK-3β by small molecule inhibitors or RNA interference prevents toxicity from high concentrations of glucose to suggest a possible targeting of GSK-3β during DM [24]. Clinical applications for GSK-3β are also attractive, especially in concert with EPO. For example, both the potential benefits of EPO to improve cardiovascular function in diabetic patients [11] and the positive effects of exercise to improve glycemic control during DM appear to rely upon the inhibition of GSK-3β activity. EPO blocks GSK-3β activity [25,26] and combined with exercise may offer synergistic benefits, since physical exercise also has been shown to phosphorylate and inhibit GSK-3β activity [1].

Future challenges for clinical care

Although the development of novel therapeutic strategies directed to improve vascular longevity in disorders such as DM may someday offer significant clinical success stories, treatments that center upon EPO, nicotinamide or Wnt signaling are far from perfect for some patients and ultimately lead to disease progression or other consequences. For example, adverse effects during treatment with EPO are well reported. These can include increased incidence of thrombotic vascular effects, progression of cardiac insufficiency, potential vascular stenosis, elevation in mean arterial pressure and increased metabolic rate and blood viscosity [9,27]. The progression of neoplastic disease has been another significant concern raised with EPO administration. Under some conditions, EPO expression has been suggested to block tumor cell apoptosis, enhance tumor progression, increase metastatic disease, and negate the effects of radiotherapy by assisting with tumor angiogenesis [28,29]. Similar to recent revelations with the links between EPO and cancer, Wnt signaling has been closely linked to tumorigenesis for a number of years [20,30].

If one looks further, nicotinamide also has been reported to have diverse biological roles that include cellular lifespan reduction. Prolonged exposure to nicotinamide in some studies can lead to impaired β-cell function and reduction in cell growth [1,4]. In addition, nicotinamide offers vascular protection in millimole concentrations against oxidative stress, but in relation to cell longevity, lower concentrations of nicotinamide can function as an inhibitor of sirtuins that have been associated with the promotion of increased lifespan yeast and metazoans [4,31]. Interestingly, it has been postulated that sirtuins may prevent nicotinamide from assisting with DNA repair by altering the accessibility of DNA damaged sites for repair enzymes. Given the intimate and inverse relationship between nicotinamide and sirtuins in controlling cell longevity and survival, alternative approaches for the treatment of vascular longevity may be required that may involve the prevention of intracellular nicotinamide accumulation.

For new therapeutic strategies to triumph against the complications of DM and address effective approaches to extend vascular cell longevity, further investigations must continue through basic as well as clinical research to overcome the present challenges of existing or developing therapies. Critical to this process is the targeted focus upon intricate cellular pathways governed by potential strategies, such as EPO, nicotinamide and Wnt signaling, to outline not only potential benefits, but also likely detriments of these therapies. In this way, new therapeutic strategies can hopefully bypass toxic complications, or at the very least avoid contraindications that can be predicted to complicate care, and offer effective, safe and robust clinical treatment for patients.

Financial & competing interests disclosure

This work was supported by the following grants: American Diabetes Association, American Heart Association (National), Bugher Foundation Award, Janssen Neuroscience Award, LEARN Foundation Award, MI Life Sciences Challenge Award, Nelson Foundation Award, NIH NIEHS (P30 ES06639), and NIH NINDS/NIA. The author has no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.