Conclusions
Our results suggest that INF and MTX are useful antagonists in apoptosis and mitochondrial oxidative stress in the neutrophils of RA patients. INF and MTX decreased the Ca2+ concentration through inhibition of the TRPM2 channel in the neutrophils of RA patients. It may be a new pathway in the mechanisms of anti-rheumatic drugs.
Material and methods
Neutrophils were isolated from 10 patients with newly diagnosed RA and 10 healthy controls. They were divided into four groups (control, RA, RA + MTX, RA + INF) and incubated with MTX and INF. In the cell viability (MTT) test, the ideal non-toxic dose and incubation time of MTX were found to be 0.1 mM and 1 h, respectively. The neutrophils were also incubated with the TRPM2 channel blocker N-(p-amylcinnamoyl) anthranilic acid (ACA).
Methods
Neutrophils were isolated from 10 patients with newly diagnosed RA and 10 healthy controls. They were divided into four groups (control, RA, RA + MTX, RA + INF) and incubated with MTX and INF. In the cell viability (MTT) test, the ideal non-toxic dose and incubation time of MTX were found to be 0.1 mM and 1 h, respectively. The neutrophils were also incubated with the TRPM2 channel blocker N-(p-amylcinnamoyl) anthranilic acid (ACA).
Results
Intracellular free Ca2+ concentration, intracellular reactive oxygen species (ROS) production, mitochondrial depolarization, lipid peroxidation, apoptosis, and caspase 3 and caspase 9 activities were found to be significantly higher in the neutrophils of RA patients compared to controls. MTT, reduced glutathione (GSH) level, and glutathione peroxidase (GSHPx) activity were significantly lower in the neutrophils of RA patients. However, MTT, GSH and GSHPx values were detected to be significantly increased with INF and MTX therapies. The Ca2+ concentrations were further decreased by the ACA therapy. Conclusions: Our results suggest that INF and MTX are useful antagonists in apoptosis and mitochondrial oxidative stress in the neutrophils of RA patients. INF and MTX decreased the Ca2+ concentration through inhibition of the TRPM2 channel in the neutrophils of RA patients. It may be a new pathway in the mechanisms of anti-rheumatic drugs.