No abstract

The label-free detection of oligonucleotides of 12-14 bases at a concentration of 10 −7 M (double helix) was achieved by using surface enhanced Raman scattering (SERS) and spermine as the aggregant of Ag sol. The wavenumbers of the produced SERS spectra of DNA are similar to those in the corresponding normal Raman spectra of free DNA, allowing the detailed assignment of the vibrational modes. The conformation of the adsorbed DNA, the adsorption geometry, and a molecular model of the interactions among DNA, spermine, and Ag nanoparticle are derived from the SERS spectra. The results show that the protonated amine groups of spermine interact with phosphodioxygens of DNA and N7s of dA and dG from the major groove. The protonated amines are also attracted to the negatively charged Ag surface and thus induce the adsorption of DNA on the metal surface. The DNA remains mostly in the B-conformation with a variation in the C5 -O dihedral angle. DNA lies flat with the bases perpendicular to the metal surface.

Diabetes is associated with hyperglycemia and increased thrombin production. However, it is unknown whether a combination of high glucose and thrombin can modulate the expression of NAPDH oxidase (Nox) subtypes in human aortic endothelial cells (HAECs). Moreover, we investigated the role of a diabetes-associated microRNA (miR-146a) in a diabetic atherothrombosis model. We showed that high glucose (HG) exerted a synergistic effect with thrombin to induce a 10.69-fold increase in Nox4 mRNA level in HAECs. Increased Nox4 mRNA expression was associated with increased Nox4 protein expression and ROS production. Inflammatory cytokine kit identified that the treatment increased IL-8 and IL-6 levels. Moreover, HG/thrombin treatment caused an 11.43-fold increase of THP-1 adhesion to HAECs. In silico analysis identified the homology between miR-146a and the 3′-untranslated region of the Nox4 mRNA, and a luciferase reporter assay confirmed that the miR-146a mimic bound to this Nox4 regulatory region. Additionally, miR-146a expression was decreased to 58% of that in the control, indicating impaired feedback restraint of HG/thrombin-induced endothelial inflammation. In contrast, miR-146a mimic transfection attenuated HG/thrombin-induced upregulation of Nox4 expression, ROS generation, and inflammatory phenotypes. In conclusion, miR-146a is involved in the regulation of endothelial inflammation via modulation of Nox4 expression in a diabetic atherothrombosis model.