Activation of RAGE induces elevated O2− generation by mononuclear phagocytes in diabetes

Abstract

Oxidative stress has been found to play a role in the pathogenesis of diabetic complications. The aim of this study was to define the oxidative burst of diabetic monocytes to characterize the phenotype associated with poor diabetic control. Superoxide (O 2 -) is the first molecule generated during the respiratory burst of phagocytes by NADPH oxidase, and its generation by monocytes from 26 controls and 34 diabetic subjects was evaluated in this study. Under resting states or stimulation by PMA or opsonized zymosan, diabetic monocytes produce significantly more O 2 -than nondiabetic cells. The increased O 2 -generation was found to be correlated with glycemic control (glycated hemoglobin) of patients. To clarify the effects of hyperglycemia on O 2 -generation, normal human monocytes were treated with receptor for advanced glycation endproducts (RAGE) ligands (AGE protein and S100B) or high glucose media before stimulation. RAGE ligands and high glucose concentration increased O 2 -generation from human mononuclear phagocytes. RAGE ligands, specifically and potently, increased O 2 -generation from mononuclear phagocytes, and high-glucose effects were associated with correspondingly increased osmotic pressure. Differentiated THP-1 cells, from a human monocytic cell line, were used as a model of human monocytes to study the effects of S100B, the RAGE ligand. It was confirmed that RAGE is involved in the priming of O 2 -generation by S100B. This study demonstrates that RAGE ligands can contribute significantly to the hyperresponsive phenotype of diabetic monocytes, which might be reversible by blocking the RAGE or controlling the presence of RAGE ligands by controlling hyperglycemia.

Phosphorylation of Pleckstrin Increases Proinflammatory Cytokine Secretion by Mononuclear Phagocytes in Diabetes Mellitus

“…Because it is difficult to standardize the individual variations among human subjects with respect to monocyte/macrophage functions, we used THP-1 cells to study the mechanism of hyperglycemia-induced changes in PKC activation and involvement of pleckstrin in vitro. Preliminary experiments by our group and others confirm the expression of RAGE on THP-1 cells as well as functions such as oxidative burst and cytokine production (41,46). Furthermore, a similar phosphorylation pattern of PKC substrates responding to PMA stimulation between THP-1 cells and human monocytes (Fig.…”

“…AGE are nonenzymatically glycated molecules that are elevated in the serum of diabetics (46,52). Many cell types express RAGE (53) and it has be demonstrated that binding of AGE to innate immune cells leads to increased inflammation (54 -56).…”

“…We have recently reported that RAGE activation activates PKC (46). To study the role of pleckstrin in this process, we used a RAGE ligand, S100B, to treat monocytes/macrophages.…”

RAGE ligands induce apoptotic cell death of pancreatic β-cells via oxidative stress

“…Engagement of RAGE by its ligands generates ROS, in part via superoxide-producing enzyme NADPH oxidase and also, possibly, via mitochondrial pathways in both vascular and neuronal cells affected by diabetes (8)(9)(10). The source of the ROS in ß-cells also remains controversial with two proposed candidates, i.e., leakage from the mitochondrial electron transport chain and NADPH oxidase (30).…”

“…In pathological conditions, especially in diabetic vascular diseases, RAGE expression is often upregulated with increased expression of its ligands as well (6,7). Engagement of RAGE by its ligands was shown to generate reactive oxygen species (ROS) via receptor-dependent signaling and to activate inflammatory signaling cascades in vascular and inflammatory cells (8)(9)(10)(11).…”