concanavalin-a and Diabetes-Mellitus--Type-2

Among multiple causes, diabetic nephropathy (DN) is the major underlying renal disease that leads to end-stage renal disease (ESRD), and early diagnosis can effectively prevent or delay the progression to ESRD. Therefore, the current study aimed to develop noninvasive, accurate detection markers.. For this study, 62 diabetes mellitus (DM) patients, 59 DN patients and 21 healthy controls (HCs) were recruited. All participants' serum samples were subjected to concavanalin (Con) A affinity chromatography, which utilizes glycoproteins to discover potential markers.. From nano LC-MS and Western blot analysis, apolipoprotein A-IV (ApoA4) was selected which featured a gradual, almost twofold increase in the order of HC, DM and DN. In the Con A-based ELISA, the DM group was 1.91-fold higher than the HC group, while the DN group was 2.56-fold higher than the HCs and 1.33-fold higher than the DM group. In addition, significant positive correlations were observed between ApoA4 and blood urea nitrogen levels and between ApoA4 and creatine levels, while significant negative correlations were seen between serum protein levels and between serum albumin levels in comparisons of DM and DN samples.. Serum Con A-bound ApoA4 levels were higher in the DM group than in HCs, and further increased in the DN group. Levels of ApoA4 were positively correlated with blood urea nitrogen and creatine, but negatively correlated with serum protein and albumin. This evidence supports serum Con A-bound ApoA4 as a circulating marker for predicting the progression of renal impairment in DM patients.

Topics: Aged; Apolipoproteins A; Blood Proteins; Blood Urea Nitrogen; Blotting, Western; Case-Control Studies; Chromatography, Liquid; Concanavalin A; Creatinine; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Disease Progression; Enzyme-Linked Immunosorbent Assay; Female; Humans; Male; Mass Spectrometry; Middle Aged; Prognosis; Serum Albumin

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

Plasma cell differentiation antigen 1 (PC-1) is an inhibitor of insulinreceptor tyrosine-kinase. PC-1 content is elevated in muscle and adipose tissue from insulin-resistant subjects and its elevation correlates with in vivo insulin resistance. It is known that insulin resistance in uraemia may be improved with erythropoietin (EPO) and vitamin D therapy. Therefore, in this study the effects of human recombinant EPO and 1-alpha-D3 treatments on lymphocyte PC-1 expression in patients with end-stage renal failure on haemodialysis (HD) were investigated.. Lymphocyte basal, concanavalin A (Con A), and phorbol-12-myristate13-acetate (PMA)-stimulated PC-1 activity were investigated in HD patients before and after a two-month treatment with subcutaneous EPO (15 patients, 2000-3000 U thrice weekly) or oral 1-alpha-D3 (14 patients, 2 mug thrice weekly). Twenty-nine patients (16 men and 13 women), aged 22-68 years (49+/-7 years), on HD from 13 to 112 months, and 30 healthy controls participated in the study. None was obese and all had normal fasting plasma glucose.. A two-month EPO treatment produced a 41% haematocrit increase, with a rise in haemoglobin from 6.51+/-0.18 g/dL to 9.69+/-0.14 g/dL. Basal lymphocyte PC-1 activity in HD patients was found to be significantly increased (P<0.005) over the level in healthy controls. Treatment of patients with EPO decreased unstimulated lymphocyte PC-1 activity to values significantly lower than before the treatment (P<0.001). Lymphocyte Con A and PMA-stimulated PC-1 activity in patients on HD was found to be slightly increased over the level in healthy controls, but significantly reduced (P<0.005 and 0.05, respectively) after the EPO treatment. A two-month pulse oral 1-alpha-D3 treatment increased haematocrit by 21% and raised haemoglobin from 7.11+/-0.32 g/dL to 8.80+/-0.39 g/dL. This treatment normalized serum alkaline phosphatase activity and slightly reduced serum parathyroid hormone concentration. PC-1 in unstimulated and PMA-stimulated lymphocytes was unchanged, but significantly decreased (P<0.05) in Con A-stimulated lymphocytes after 1-alpha-D3 treatment. Fasting plasma glucose was not changed by the treatment.. An increased lymphocyte PC-1 activity over control was found in HD patients. A two-month EPO therapy significantly decreased PC-1 activity to the control values, suggesting that an effect on PC-1 expression could be implicated in the amelioration of insulin resistance in uraemic patients treated with EPO. Treatment with pulse oral 1-alpha-D3 had an effect only on PC-1 of Con A-transformed lymphocytes of haemodialysed patients and requires further investigation.

Topics: Adjuvants, Immunologic; Administration, Oral; Adult; Aged; Blood Glucose; Carcinogens; Concanavalin A; Diabetes Complications; Diabetes Mellitus, Type 2; Erythropoietin; Female; Humans; Hydroxycholecalciferols; Insulin Resistance; Kidney Failure, Chronic; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Phosphoric Diester Hydrolases; Pyrophosphatases; Recombinant Proteins; Renal Dialysis; Tetradecanoylphorbol Acetate

Superantigens have been implicated in the pathogenesis of type I diabetes and other immune-mediated diseases. We therefore tested the hypothesis of an abnormal reactivity of the immune system toward bacterial superantigens during the prediabetic phase. For this purpose, splenocytes from NOD (H-2g7) mice were exposed to two well-characterized superantigens: Staphylococcal aureus enterotoxin-B (SEB) and toxic shock syndrome toxin-1 (TSST-1). Cells from BALB/c (H-2d) and C57BL/6 (H-2b) mice as well as those from NON (H-2non) and NOR (H-2g7) mice were used as controls. After 72 h of co-culture with the superantigens or the mitogen concanavalin A (Con A), proliferative response and mitochondrial activity were determined. In the culture supernatants, the cytokines gamma-interferon (IFN-gamma) and interleukin 10 (IL-10) were measured. Striking similarities between NOD cells and major histocompatiblity complex (MHC)-identical NOR cells could be observed with regard to a low proliferative and mitochondrial response to SEB, accompanied by a normal response to TSST-1 and Con A, respectively. In addition, only NOD and NOR spleen cells were low producers of the T-helper 1 (Th1) cytokine IFN-gamma in response to SEB. Conversely, abnormally high IFN-gamma levels were induced by TSST-1 in NOD and NOR spleen cells. The cytokine response to Con A was also biased toward IFN-gamma in both NOD and NOR. Since IFN-gamma and IL-10 are crucial disease-promoting or -protecting mediators in prediabetic NOD mice, superantigens may affect pathogenesis by acting on the Th1/Th2 cytokine balance. The low responder status toward SEB in NOD spleen cells may be of pathogenetic relevance in view of recent findings that the insulin B-chain also interacts with the SEB binding site on MHC class II molecules. In conclusion, we show here that immune cells from mice with a diabetes-associated MHC type respond differently to common environmental superantigens than do immune cells from control strains.

Topics: Animals; Bacterial Toxins; Concanavalin A; Diabetes Mellitus, Type 2; Enterotoxins; Female; Histocompatibility Antigens Class II; Interferon-gamma; Interleukin-10; Lymphocyte Activation; Lymphocytes; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred NOD; Mitochondria; Spleen; Staphylococcus aureus; Superantigens

Peripheral blood mononuclear cells from patients with non-insulin-dependent diabetes mellitus (NIDDM) show reduced proliferative response to phytohemagglutinin (PHA) and other mitogens. This study was undertaken to determine whether this reduced lymphocyte proliferation is mediated by a decreased production of cytokine or decreased expression of interleukin-2 receptor (IL-2R). Mononuclear cells from NIDDM patients (n = 34) and healthy controls (n = 22) were cultured in RPMI-1640 media containing PHA, concanavalin-A and phorbol myristate acetate. NIDDM patients showed reduced [3H]thymidine uptake (57% of controls, P < 0.01), reduced percentage of IL-2R-positive cells (61% of controls, P < 0.02) and increased level of tumor necrosis factor (TNF)-alpha (200% of controls, P < 0.05). The percentage of complement receptor (CR) 3-positive monocytes from NIDDM patients was also decreased (72% of controls, P < 0.05). However, the production of IL-1 beta, IL-2 and interferon-gamma, the percentages of pan T cells (CD3), T helper cells (CD4), T suppressor cells (CD8), the ratio of CD4/CD8 and the expression of CR1 and Fc receptors for immunoglobulin G (Fc gamma RII and Fc gamma RIII) were not significantly different between NIDDM patients and healthy subjects. Human recombinant IL-2 was unable to restore the [3H]thymidine uptake by PHA-stimulated mononuclear cells from NIDDM patients. Elevation of glucose concentration up to 27.8 mmol/l in the culture medium did not suppress the [3H]thymidine uptake and IL-2R expression by activated lymphocytes from healthy subjects. The decreased expression of IL-2R on activated lymphocytes might be responsible for the insufficient lymphocyte proliferation in NIDDM patients. These findings suggest that decreased expression of CR3 on monocytes, decreased lymphocyte proliferation and decreased IL-2R expression despite a higher production of TNF-alpha may explain the impaired cell-mediated immunity seen in NIDDM patients.

Topics: CD4-CD8 Ratio; Cells, Cultured; Concanavalin A; Cytokines; Diabetes Mellitus, Type 2; Female; Humans; Immunity, Cellular; Lymphocyte Activation; Male; Middle Aged; Phytohemagglutinins; Receptors, Interleukin-2; Reference Values; T-Lymphocyte Subsets; T-Lymphocytes; Tetradecanoylphorbol Acetate; Thymidine