concanavalin-a and Hyperglycemia

Toll-like receptors (TLRs) play an important role in several inflammatory diseases such as diabetes. The present study was to determine whether hyperglycemia in diabetes interferes in reactive oxygen species (ROS) production in granulocytes stimulated with either TLR2/zymosan, TLR4/lipopolysaccharide (LPS) or TLR2,4,9/concanavalin A (ConA). NADPH (nicotinamide adenine dinucleotide phosphate) oxidase and mitogen-activated protein kinase (MAPK) signaling pathways associated with ROS generation in TLR-stimulated granulocytes were evaluated. Our results demonstrate that ROS generation in resting granulocytes derived from patients suffering from Type 2 diabetes mellitus (T2DM) is significantly higher than that observed in equivalent cells from healthy controls. However, ROS formed by TLR-stimulated granulocytes from T2DM patients and healthy subjects were comparable. ROS production by TLR4,9 depends on NADPH-oxidase and MAPK signaling pathways. In contrast, the activation of TLR2 leads to ROS production by a mechanism that is dependent on NADPH oxidase but independent of the MAPK. In conclusion our results suggest that hyperglycemia in diabetes may prime cells metabolically for ROS generation but does not exert any significant effect on TLR-stimulated ROS production and possibly does not aggravate the development of ROS-dependent diabetic complications.

Topics: Adult; Aged; Aged, 80 and over; Cells, Cultured; Concanavalin A; Diabetes Mellitus, Type 2; Female; Granulocytes; Humans; Hyperglycemia; Lipopolysaccharides; Male; Middle Aged; Mitogen-Activated Protein Kinases; NADPH Oxidases; Reactive Oxygen Species; Toll-Like Receptors; Zymosan

Diabetes mellitus (DM), one of the commonest metabolic disorders, can impair the function of cells involved in cellular and/or humoral immunity. This study sought to define potential effects upon cell-mediated immune cells due to an acute hyperglycemic state (in vitro) for comparison against those that might be attributable to a diabetic phenotype itself. Peripheral blood mononuclear cells (PBMC) were isolated from ten diabetic patients (5 with Type I disease and 5 with Type II) and 10 healthy controls. The cells were then challenged with 1 of 3 different mitogens (concanavalin A, phytohemagglutinin, pokeweed mitogen) in the presence of differing glucose concentrations (0, 100, 200, 400, or 800 mg/dl), and proliferative responses assessed. Neutrophils (PMNC) from the blood samples, exposed to the same experimental conditions, were analyzed for respiratory burst activity using nitroblue tetrazolium. The results indicated that there was significant inhibition of the proliferative responses to mitogens among the stimulated PBMC and in respiratory burst activity among the PMNC obtained from the diabetic patients. However, these effects were not affected by either the added presence of increasing amounts of exogenous glucose, the type of diabetes the patients had, the length of time the patient had had the disease, or whether or not the patients had been receiving insulin treatments. In contrast, the PBMC from healthy individuals appeared to display dose-trend decreases in responsiveness to mitogens; interestingly, similar effects on their PMNC were not evident. It was thus concluded that in situ ongoing repeated hyperglycemic states caused changes in cells of the immune system that could have been caused by repeated "continuous" exposures to excess sugar. Further studies are needed to more clearly identify hyperglycemia (sugar)-sensitive targets on/in these cells that could contribute to the appearance of the diabetic immunodeficiency in these types of patients.

Topics: Adult; Cell Proliferation; Concanavalin A; Diabetes Mellitus; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Female; Glucose; Humans; Hyperglycemia; Leukocytes, Mononuclear; Male; Middle Aged; Neutrophils; Nitroblue Tetrazolium; Phytohemagglutinins; Pokeweed Mitogens; Respiratory Burst

Severe diabetes with insulitis was produced in young diabetes-prone BB/W rats by passive transfer of concanavalin A-treated spleen cells from BB/W animals with acute diabetes. Spleen cells alone or in combination with lymph node cells were active in transferring disease.

Topics: Animals; Concanavalin A; Diabetes Mellitus; Hyperglycemia; Immunity, Cellular; Mice; Mice, Nude; Rats; Spleen; Transplantation, Heterologous; Transplantation, Homologous

Topics: Aging; Animals; Blood Glucose; Concanavalin A; Diabetes Mellitus, Experimental; Female; Glycosuria; Hyperglycemia; Islets of Langerhans; Islets of Langerhans Transplantation; Male; Pancreas; Rats; Tissue Preservation; Transplantation, Isogeneic

Supplementation of rat lymphocyte cultures with plasma from alloxan-diabetic rats produced a dose-dependent suppression of mitogen-induced blastogenic responses. Viability measurements indicated that this inhibition was not due to a direct cytotoxic effect of alloxan-diabetic plasma on rat mononuclear leukocytes in vitro. This inhibition was not explained by hyperglycemia alone and was observed when blastogenesis was induced in vitro by phytohemagglutinin (PHA), concanavalin A (con A), or allogeneic cells. Peripheral blood lymphocytes from alloxan-diabetic rats appeared to be more sensitive than normal cells to the inhibitory effect of diabetic plasma. Heating at 56 degrees for 60 minutes produced only a partial loss of the inhibition by diabetic plasma. Alloxan-diabetic rat plasma promoted an increased accumulation of adenosine 3',5'-monophosphate (cyclic AMP) in mononuclear leukocytes. Insulin (1 and 100 muU./ml.) in vitro enhanced PHA-induced blastogenesis but failed to reverse the inhibition caused by diabetic plasma. Ultrafiltration through a cellulose dialysis membrane with an exclusion size of 12,000 molecular weight did not remove the inhibitory factor(s). These results indicate that a depressed cellular immune response is produced during an insulin-deficient diabetic state. The suppressed in-vitro blastogenic response of lymphocytes from alloxan-diabetic rats appears to involve some circulating inhibitory factor(s) in diabetic plasma. This inhibition may be explained, in part, by the ability of diabetic plasma to elevate cyclic AMP in mononuclear leukocytes.

Topics: Animals; Antilymphocyte Serum; Blood; Cell Survival; Cells, Cultured; Concanavalin A; Cyclic AMP; Cytotoxicity Tests, Immunologic; Diabetes Mellitus, Experimental; Dose-Response Relationship, Drug; Hot Temperature; Hyperglycemia; Insulin; Lectins; Lymphocyte Activation; Male; Rats; Thymidine