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


METHODS
Splenocytes of Swiss albino mice were irradiated with various doses (0-7 Gy) of gamma-rays (60Co) 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 4Gy. 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.
Agrawal
A
A
Free Radical Biology laboratory, School of Iife Sciences, Jawaharlal Nehru University, New Delhi-110067, India.
Shukla
S
S
Kale
R K
RK
D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis / Q000502:physiology / Q000528:radiation effects; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004307:Dose-Response Relationship, Radiation; D004789:Enzyme Activation / Q000528:radiation effects; D005720:Gamma Rays; D015227:Lipid Peroxidation / Q000528:radiation effects*; D008297:Male; D008560:Membrane Fluidity / Q000528:radiation effects*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D011829:Radiation Dosage; D013154:Spleen / Q000166:cytology / Q000378:metabolism* / Q000528:radiation effects*
D000975:Antioxidants; D016328:NF-kappa B; D009569:Nitric Oxide; D002118:Calcium