Association of POC-based Measurement of Skin AGEs with Serum AGEs as well as AGE Biomarkers in Individuals with and without Glucose Intolerance.

Accumulation of advanced glycation end products (AGEs) occurs with aging and in various disease states. There are no reliable screening techniques to measure AGEs in clinical settings. In this study, a point-of-care (POC) device was used to validate skin AGE measurements with serum AGE levels and to assess its usefulness to identify individuals with abnormal glucose tolerance (AGT). MATERIALS AND METHODS: The study group comprised individuals with normal glucose tolerance (NGT: n = 47) and with AGT, that is, either diabetes or prediabetes (n = 68). Intrinsic AGE fluorescence was measured spectrofluorimetrically using multimode plate reader in the serum by exciting the samples at 370 nm and emission readouts at 440 nm. Skin AGEs were acquired using a CE-marked Scout DS commercial device. Serum levels of biomarkers carboxymethyl lysine (CML), carboxyethyl lysine (CEL), and pentosidine were analyzed by enzyme-linked immunosorbent assay (ELISA). RESULTS: In subjects with AGT, the skin AGEs [61.3 vs 53.7 arbitrary units (AU), p<0.0001] and serum AGEs (3.5 vs 2.8 AU, p<0.0001) were significantly higher than in individuals with NGT. The levels of CML, CEL, and pentosidine were also significantly higher in the subjects with AGT when compared with NGT (138 vs 89 pg/mL; 2.4 vs 1.4 nmol/mL, and 64 vs 48 pmol/mL, p<0.0001), respectively. Pearson correlation analysis showed a significant positive association of skin AGEs with serum AGEs (r = 0.344) (p<0.001), CML (r = 0.323) (p<0.001), CEL (r = 0.308) (p<0.001), and pentosidine (r = 0.251) (p<0.001). In addition, it also showed a positive correlation with fasting plasma glucose (FPG) (p<0.001), 2-hour post-glucose (p<0.001), glycated hemoglobin (HbA1c) (p<0.001), and body mass index (BMI) (p<0.05). Multiple logistic regression analysis using AGT as a dependent variable showed that skin AGE scores were significantly (p<0.001) associated with AGT (odds ratio: 1.133, confidence intervals: 1.067-1.203). CONCLUSION: This study shows that the measurement of skin AGEs using a POC device may be suitable for mass screening of AGT even in low-resource settings.

Convergence of innate immunity and insulin resistance as evidenced by increased nucleotide oligomerization domain (NOD) expression and signaling in monocytes from patients with type 2 diabetes.

Despite the well known role of nucleotide oligomerization domain (NOD) receptor proteins in innate immunity, their association with diabetes is less explored. Here we report the transcriptional level of NODs and their downstream molecular signatures in CD14(+) monocytes from subjects with different grades of glucose tolerance. NOD1 and NOD2 mRNA expression were significantly up-regulated in monocytes from patients with type 2 diabetes (T2DM) and positively correlated with HOMA-IR and poor glycemic control. Patients with T2DM also exhibited increased monocyte activation markers (CD11b and CD36) and proinflammatory signals downstream of NOD (RIPK2 and NFκB) along with the increased circulatory levels of TNF-α and IL-6. In vitro stimulation of monocytes with NOD specific ligands-i-EDAP and MDP significantly up regulated the mRNA expression of NOD1 and NOD2 respectively in T2DM. Our study exposes up regulation of NODs in monocytes as an important component of inflammation and insulin resistance in patients with T2DM.

Coordinated augmentation of NFAT and NOD signaling mediates proliferative VSMC phenotype switch under hyperinsulinemia.

AIM: Although hyperglycemia has been demonstrated to play a significant role in the vascular disease associated with type 2 diabetes, the mechanisms underlying hyperinsulinemia mediated vascular dysfunction are not well understood. We have analyzed whether hyperinsulinemia could activate NFAT (Nuclear factor of activated T cells) signaling and thereby influence vascular smooth muscle cell (VSMC) migration and proliferation, a major event in the progression of atherosclerosis. METHODS AND RESULTS: Human aortic VSMCs upon chronic insulin treatment exhibited increased expression of NFATc1 both at the mRNA and protein levels. The mechanistic role of NFAT in VSMC migration and proliferation was examined using 11R-VIVIT, a cell permeable NFAT specific inhibitor, where it reduced the insulin effect on VSMC, which was further substantiated by over expression or silencing of NFATc1gene (p < 0.05). This study also report for the first time the role of NFAT in NOD (Nucleotide oligomerization domain) mediated innate immune signaling and its significance in insulin effect on VSMCs. mRNA expression of NOD was up regulated when cells were treated with insulin or ligands whereas pretreatment with 11R-VIVIT reversed this effect (p < 0.05). Our study uphold the clinical significance as we observed an increased mRNA expression of NFATc1 in monocytes isolated from patients with type 2 diabetes which correlated positively with insulin resistance and glycemic load (p < 0.05). DISCUSSION: This study suggests that targeted NFAT inhibition can be an effective strategy to coordinately quench insulin induced proliferative and inflammatory responses along with innate immunity alterations in vascular smooth muscle cells, which underlie atherosclerosis.

Association of neutrophil-lymphocyte ratio with glucose intolerance: an indicator of systemic inflammation in patients with type 2 diabetes.

BACKGROUND: The neutrophil-lymphocyte ratio (NLR) has been demonstrated to be a better risk factor than total white blood cell count in the prediction of adverse outcomes in various medical conditions. This study analyzed the association of NLR with different grades of glucose tolerance and insulin resistance in Asian Indians. SUBJECTS AND METHODS: Study subjects were recruited from Phase 3 of the Chennai Urban Rural Epidemiology Study (CURES). For this cross-sectional analysis, subjects with normal glucose tolerance (NGT) (n=237), impaired glucose tolerance (IGT) (n=63), and type 2 diabetes mellitus (DM) (n=286) were selected. The hemogram was done in all subjects using a five-part hematology analyzer (model SF-3000; Sysmex, Kobe, Japan). The NLR was calculated as the ratio between counts for neutrophils and total lymphocytes. Fasting insulin was measured by enzyme-linked immunosorbent assay, and insulin resistance was calculated using the homeostasis model assessment (HOMA-IR). RESULTS: Subjects with DM showed a significantly higher NLR (2.2 ± 1.12) compared with IGT subjects (1.82 ± 0.63), who in turn had a higher ratio than NGT subjects (1.5 ± 0.41) (P<0.01). Pearson correlation analysis showed a significant positive correlation of NLR with glycated hemoglobin (r=0.411), fasting plasma glucose (r=0.378), and HOMA-IR (r=0.233) (P<0.001). Regression analysis showed a linear increase in NLR with increasing severity of glucose intolerance even after adjusting for age, waist circumference, blood pressure, triglycerides, and smoking. CONCLUSIONS: This is the first report on the correlation of NLR with different grades of glucose intolerance and insulin resistance. NLR can be used as an adjuvant prognostic marker for macro- and microvascular complications in patients with glucose intolerance.