Role of Ca2+ in radiation-induced damage in murine splenocytes

Purpose: To address the links between calcium, peroxidation, cell damage and death and the response of the enzymes involved in free radical metabolism, in splenocytes of mice irradiated with gamma-rays. Materials and methods: Splenocytes of Swiss albino mice were irradiated with various doses ( 0 - 7 Gy) of gamma-rays (Co-60) at a dose-rate of 0.0575 Gy s(-1). Membrane peroxidation and fluidity were determined by the thiobarbituric acid-reactive substances (TBARS) method, and fluorescence polarization of 1,6-diphenyl-1,3,5- hexatriene (DPH), respectively. Apoptosis was analysed by nucleosomal ladder formation and activity of NF-kappaB by electrophoretic mobility shift assay ( EMSA). The specific activities of the antioxidant enzymes, lactate dehydrogenase (LDH), levels of nitric oxide (NO.) and glutathione were determined spectrophotometrically. Modulatory effects of Ca2+ were examined at 3 Gy using different concentrations ( 1, 3 and 5 mM) in the presence or absence of the ionophore A23187. Results: Irradiation of splenocytes resulted in enhanced peroxidative damage, membrane fluidity, apoptosis and DNA binding activity of NF-kappaB. The specific activities of LDH and antioxidant enzymes superoxide dismutase (SOD), DT-diaphorase (DTD), glutathione S-transferase (GST) and levels of glutathione (GSH) and NO. were increased with radiation dose up to 4 Gy. Ca2+ augmented the radiation-induced responses. The presence of ionophore A23187 potentiated the modulatory effects of Ca2+. Conclusions: These findings show that Ca2+ augments radiation damage and is more effective intracellularly. Ca2+, peroxidation, cellular damage and apoptosis are possibly interlinked through signals, as is evident from the increased activity of NF-kappaB and generation of NO.. The enhanced antioxidant status suggests an attempt made by the irradiated cells to maintain their normal functions.