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The role of tumor necrosis factor alpha on insulin resistance and vascular dysfunction

Helena Dominguez


This PhD thesis recapitulates a series of physiologic studies performed in humans published in papers 1 and 2 respectively.
Background. Persons with type 2 diabetes develop diffuse atherosclerosis and consequent organ damage. In patients with atherosclerotic disease as well as in patients with type 2 diabetes there is an increased production and increased blood levels of the cytokine TNF-α. In the surroundings of atherosclerotic plaques there is accumulation of TNF-α, especially around culprit lesions. Atherosclerotic plaques are attached to the endothelium, which is the innermost layer of the vessels. Among other functions the endothelium helps to maintain a relaxed vascular tone by producing nitric oxide (NO). It has been shown that TNF-α impairs the intracellular signal that is initiated when insulin binds to its receptors, what results in insulin resistance. Since one of the endpoints of the insulin signal in endothelial cells is the activation of eNOS, that catalyzes NO production, it is reasonable to believe that TNF-α has a substantial role in the development of atherosclerosis in insulin-resistant states. The purpose of this PhD is to contribute to the clarification of this role.
Methods. Phase I: The effect of TNF-α on insulin-stimulated endothelial function and glucose uptake was studied in young, healthy, lean men. Forearm blood flow was measured by venous occlusion plethysmography during intra-arterial infusion of increasing doses of the NO-agonist acetylcholine to assess the endothelium capacity to produce NOmediated vasodilatation. The capacity of insulin to enhance this vasodilatation was examined co-infusing insulin and acetylcholine. Forearm glucose uptake was measured before and at the end of the insulin infusion, and repeated after TNF-α was added to these infusions. The effect of TNF-α independent of insulin was assessed in control studies where insulin was substituted by its vehicle. The effect of TNF-α on non-endothelium mediated vasodilatation was assessed in control studies where acetylcholine was substituted by infusion of the NO-donor sodium nitroprusside. Phase II: 20 adipose patients with type 2 diabetes were randomized to treatment with etanercept, a TNF-α inhibitor, in an open-label parallel study. A 25 mg subcutaneous injection of etanercept was administered twice weekly for 4 weeks. Vascular function and forearm glucose uptake were assessed at baseline and at the end of the etanercept treatment or an equivalent Page 9 of 40 control period. Furthermore, the patients randomized to etanercept treatment were examined with: intravenous glucose tolerance tests (to assess β−cell function) and hyperinsulinemic euglycemic clamps (to assess whole body insulin sensitivity). Oral glucose tolerance tests and measurement of systemic inflammatory markers were performed in both groups.
Results. In phase I a short TNF-α infusion that increased 100-fold TNF-α levels impaired simultaneously insulin effects on endothelium-mediated vasodilatation and on forearm glucose uptake. In phase II etanercept provoked a marked and significant decrease on the systemic inflammatory markers C-reactive protein and interleukin-6. In spite of this, there was no improvement on insulin effect or on other measurements of vascular function. Insulin effect on forearm glucose uptake and insulin sensitivity during the clamps did not improve either. Conclusion. An acute and marked rise of TNF-α impairs simultaneously vascular and metabolic insulin sensitivity. Nevertheless, in the low-grade chronic inflammation that occurs in obesity and type 2 diabetes, a decrease of systemic inflammatory markers achieved by TNF-α blockade with etanercept does not reverse insulin resistance.