Effect of glycated insulin on the blood-brain barrier permeability: An in vitro study.

Altered blood-brain barrier (BBB) permeability may contribute to pathogenesis of diabetes-related central nervous system disorders. Considering the presence of glycated insulin in plasma of type 2 diabetic patients, we hypothesized that glycated insulin could induce changes in paracellular permeability in BBB. Therefore, the authors decided to study the effect of glycated insulin on paracellular permeability in a BBB model and the change induced in insulin conformation upon glycation. In this study, the structural modification was examined by fluorescence and circular dichroism spectroscopies and dynamic light scattering. Cell proliferation and production of ROS in astrocytes and HUVEC cells were analyzed by MTT and spectrofluorometric assays, respectively. Apoptosis induction was determined and confirmed by flow cytometry and western blot analyses, respectively. The permeability was measured Lucifer yellow and FITC-Dextran. According to our results, glycated insulin presented altered conformation and more exposed hydrophobic patches than insulin. Formation of oligomeric species and advanced glycated end products (AGEs) were determined. Lower cell viability, higher apoptosis, and more ROS were detected upon treatment of cells with glycated insulin. Finally, glycated insulin led to increased Lucifer yellow and FITC-dextran transportation across the BBB model which could result from ROS producing and apoptosis-inducing activities of AGE-insulin.

Glycation restrains open-closed conformation of Insulin.

In hyperglycemic conditions, the level of reactive dicarbonyl metabolites concentration is found to be high, which plays a significant role in protein glycation. Despite decades of research, the effect of methylglyoxal on the structure and function of insulin is still unknown. Through a shift in conformation at the B-chain C-terminal (BT-CT) hinge from an "open" to a "wide-open" conformation, insulin binds to the receptor and activates the signal cascade. Insulin resistance, which is the main sign of Type 2 Diabetes, can be caused by a lack of insulin signaling. Methylglyoxal site-specific glycation in residue R22 at B chain forms AGE product Methylglyoxal-hydroimidazolone (MGH1) in insulin. In this work, we present molecular dynamics study of this glycated insulin R22MGH1, which revealed new insights into the conformational and structural changes. We find the following key results: 1) B-chain in insulin undergoes a closed conformational change upon glycation. 2) Glycated insulin shows secondary structure alteration. 3) Glycated insulin retains its closed shape due to an unusually strong hydrophobic contact between B-chain residues. 4) Wide open native conformation of insulin allows the B chain helix to be surrounded by more water molecules compared to the closed conformation of glycated insulin. The closed conformation of glycated insulin impairs its binding to insulin receptor (IR).