Homocysteine induced SH-SY5Y apoptosis through activation of NADPH oxidase in U251MG cells

Epidemiological studies have indicated a correlation between homocysteinemia and dementia, including Alzheimer's disease. However, the mechanism by which homocysteine (Hcy) induces neuronal cell death remains unknown. We found that micromolar concentrations of Hcy induced neuroblastoma SH-SY5Y cell death only when co-cultured with glioblastoma U251MG cells. In this culture system, cysteine had no effect on SH-SY5Y cell death. There was an increase in TUNEL-positive cells and loss of mitochondrial membrane potential following treatment with 100 μM Hcy. Addition of conditioned medium prepared from U251MG cells in the presence of 100 μM Hcy also reduced SH-SY5Y cell viability, while this effect was prevented when using conditioned medium from U251MG cells exposed to 100 μM Hcy + apocynin, a specific NADPH oxidase inhibitor. Following exposure to 100 μM Hcy in U251MG cells, expression of Rac1, a compartment of NADPH oxidase, was translocated to the plasma membrane, and the active form of Rac1 was increased. There was no change in peroxide concentration in the medium of U251MG cells after addition of Hcy. Overall, these data suggest that Hcy stimulates Rac1 activation and NADPH oxidase, resulting in superoxide anion production that may induce SH-SY5Y cell apoptosis.