TMPDP, a tetramethylpyrazine derivative, protects vascular endothelial cells from oxidation damage by hydrogen peroxide.

A novel ligustrazine derivative, tetramethylpyrazine diphenylmethyl piperazidine (TMPDP), prepared by hybridization and bioisosteric replacement of the molecular structure of TMP, was studied for its protective effects on oxidative damage of human umbilical vein endothelial cells (HUVECs) in response to hydrogen peroxide (H2O2). The antioxidative effect of TMPDP was assessed by the 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) test. Cell viability was measured using a 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The activity of lactate dehydrogenase (LDH), superoxide dismutase (SOD) and glutathione peroxidase (GSH) and the content of malondialdehyde (MDA) in cells were determined by commercial kits. The intracellular formation of reactive oxygen species (ROS) and the concentration of free intracellular calcium ([Ca2+]i) were determined using DCFH-DA assay and with fura-2/AM fluorimetry, respectively. Results showed that TMPDP had a moderate antioxidative effect against DPPH. Cell viability was decreased markedly by exposure to H2O2. Introduction of TMPDP, however, significantly increased cell viability, markedly reduced LDH release from cells and decreased lipid peroxidation in response to H2O2 treatment. These effects of TMPDP were accompanied by increased activity of the endogenous antioxidant enzymes, SOD and GSH, reduced production of ROS and reduced intracellular concentration of Ca2+. These results suggest that TMPDP protects HUVECs against oxidative damage by scavenging ROS and regulates intracellular calcium concentration. This might have important implications for the development of new agents for the effective treatment of vascular disease.