Pathway of reactive oxygen species production in rabbit vascular smooth muscle cells

Objective To study the pathway of reactive oxygen species (ROS) production in rabbit vascular smooth muscle cells ( VSMCs). Methods Primary cell culture of VSMCs and MTT assay were performed to detect the cell viability after being treated with different concentrations of H2O2. 100 μmol/L H2O2 was used on VSMC as the final concentration, intracellular ·O2- production was measured by flow cytometry and mRNA expression of subunits of VSMC NADPH oxidase was detected by RT-PCR at different time points after treatment. Results VSMC viability was decreased gradually with the increase of the concentration of H2O2. When the concentration ≥30 μmol/L, the A value at each time point was decreased significantly (P 0.05). Under the condition of 100 μmol/L H2O2, intracellular· O2- production was increased and reached the peak value at 24th h after treatment (24.01% of DHE-positive cell rates) ; the expression of p22phox mRNA was increased gradually and reached the peak value 1 h after treatment (2 times of 0 h), while gp91phox and noxl mRNA expression decreased, especially the expression of noxl mRNA decreased down to the lowest 1 h after treament (as 15 % as 0 h). Conclusion Certain concentration of exogenous H2O2 can contribute to VSMC· O2- prduction, and NADPH oxidase may involves in these cellular events.