Increased flux of free radicals in cells subjected to hyperthermia: detection by electron paramagnetic resonance spin trapping

Abstract

It has been hypothesized that hyperthermia promotes oxygen-centered free radical formation in cells; however, to date there is no direct evidence of this heat-induced increase in oxygen free radical flux. Using electron paramagnetic resonance (EPR) spin trapping, we sought direct evidence for free radical generation during hyperthermia in intact, functioning cells. Rat intestinal epithelial cell monolayers were exposed to 45°C for 20 min, after which the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was added. Compared to control cells at 37°C, heat-exposed cells had increased free radical EPR signals, consistent with the formation of DMPO/ ⋅OH (a N=a H=14.9 G). These findings indicate that heat increases the flux of cellular free radicals and support the hypothesis that increased generation of oxygen-centered free radicals and the resultant oxidative stress may mediate in part, heat-induced cellular damage.