Molecular Link between Glo-1 Expression and Markers of Hyperglycemia and Oxidative Stress in Vascular Complications of Type 2 Diabetes Mellitus.

Chronic hyperglycemia and oxidative stress in Type 2 Diabetes Mellitus trigger cellular dysfunction via the formation of Advanced Glycation End Products (AGEs), resulting in dicarbonyl stress. Glyoxalase-1 (Glo-1) is the main defense against dicarbonyl stress. The aim of this study was to explore any cross-talk between Glo-1 and markers of hyperglycemia and oxidative stress. The siRNA-mediated downregulation of Glo-1 was performed in human microvascular endothelial cell line (HMEC-1). A Glo-1 transgenic rat model was developed. Glo-1 activity, as determined spectrophotometrically, and methylglyoxal were quantified using UPLC-MS/MS and the expression of representative markers of hyperglycemia and oxidative stress was performed using quantitative real-time PCR. A significant increase in the expression of Vascular Cell Adhesion Molecule-1 (VCAM-1) was observed in the case of the siRNA-mediated downregulation of Glo-1 in the microvasculature model under hyperglycemic conditions (p-value < 0.001), as well the as overexpression of Glo-1 in the macrovasculature (p-value = 0.0125). The expression of thioredoxin interacting protein (TXNIP) was found to be significantly upregulated in wildtype diabetic conditions vs. Glo-1 transgenic control conditions (p-value = 0.008), whereas the downregulation of Glo-1 had no impact on TXNIP expression. These findings substantiate the role of VCAM as an important marker of dicarbonyl stress (represented by Glo-1 downregulation), as well as of hyperglycemia, in diabetic vascular complications. Our findings also suggest a potential feedback loop that may exist between Glo-1 and TXNIP, as the highest expression of TXNIP is observed in cases of wildtype diabetic conditions, and the lowest expression of TXNIP is observed when Glo-1 transgene is being expressed in absence of dicarbonyl stress.

Molecular dynamics simulations and bioinformatics' analysis of deleterious missense single nucleotide polymorphisms in Glyoxalase-1 gene.

Glyoxalase-1 (Glo-1) is a key member of the Glyoxalase system, the primary line of defense against dicarbonyl stress which, in tandem, with reduced levels of expression or activity of Glyoxalase-1 enzyme, has been implicated in various human diseases like type 2 diabetes mellitus (T2DM) and its vascular complications. The association of Glo-1 single nucleotide polymorphisms with genetic susceptibility to T2DM and its vascular complications is yet to be explored. Therefore, in this study, we have employed a computational approach to identify the most damaging missense or nonsynonymous SNPs (nsSNPs) in Glo-1 gene. Initially, we characterized missense SNPs that are damaging to the structural and functional integrity of Glo-1 using various bioinformatic tools. These tools included SIFT, PolyPhen-2, SNAP, PANTHER, PROVEAN, PhD-SNP, SNPs&GO, I-Mutant, MUpro and MutPred2. One of these missense SNPs (rs1038747749; corresponding to amino acid change Arginine to Glutamine at position 38) was found to be highly conserved in evolution and is an important part of the enzyme's active site, glutathione binding site, as well as the dimeric interface based on the results obtained from ConSurf and NCBI Conserved Domain Search tools. Project HOPE reported that this mutation replaces a positively charged polar amino acid (Arginine) with a small, neutrally charged amino acid (Glutamine). Comparative modelling of wildtype and mutant (R38Q) Glo-1 proteins was performed in the run up to molecular dynamics simulation analysis which showed that rs1038747749 adversely impacts Glo-1 protein's stability, rigidity, compactness, hydrogen bonds/interactions as demonstrated by the results of various parameters computed during the analysis.Communicated by Ramaswamy H. Sarma.

The microRNA regulatory network: a far-reaching approach to the regulate the Wnt signaling pathway in number of diseases.

Wnt signaling pathway plays an important role in cell renewal, tumorigenesis, organogenesis, bone formation and bone resorption. Wnt signaling pathway is divided into two outlets: Wnt-ß-catenin pathway (canonical pathway) and Wnt-calcium pathway (non-canonical pathway). miRNAs play a key role in the regulation of Wnt signaling pathway. In this review, we highlight the basic indulgent of miRNAs-mediated regulation of Wnt signaling pathway. We focus on the role of miRNAs at different levels of Wnt signaling: signaling molecules, their associated signaling proteins, regulatory proteins, transcription factors and related cytokines. Finally, we concluded that these multiple levels of targeting may have diagnostic potential as well as therapeutic prospective in future treatment.