STEIN, J.R.*; HOFER, T.; LEEUWENBURGH, C.; JULIAN, D.; University of Florida; University of Florida; University of Florida; University of Florida: Cellular oxidative stress in H2S-exposed erythrocytes from a marine polychaete

Marine invertebrates endemic to environments characterized by high hydrogen sulfide concentrations utilize sulfide oxidation to reduce internal sulfide. However, it has been proposed that sulfide oxidation in these animals may cause oxidative stress through the formation of oxygen- and sulfur-centered free radicals (Tapley et al. 1999). Free radicals can cause oxidative stress, which may include damage to cellular organelles and biomolecules such as DNA. We exposed erythrocytes from the sulfide-tolerant marine polychaete Glycera dibranchiata to hydrogen sulfide in vitro for 1h. We then assayed overall intracellular oxidative stress by measuring the intracellular conversion of the free radical indicator H2DCF and the production of mitochondrial superoxide using MitoSOX. Oxidative damage to RNA and DNA was measured by detecting the oxidation of guanosine to 8-oxoG and deoxyguanosine to 8-oxodG using HPLC-EC-UV. Erythrocytes exposed to 290 μM, 730 μM, and 1.9 mM sulfide for 1h showed a 60% increase in intracellular oxidative stress at 1.9 mM sulfide (P<0.0001, n=4) and a 300% increase in superoxide production (P=0.00019, n=4) with no significant increases at lower sulfide concentrations. Erythrocytes exposed to 290 μM and 730 ÁM sulfide for 1h showed a 600% increase in oxidative RNA damage (p=0.0012, n=4-5) and a 300% increase in oxidative DNA damage (p<0.0001, n=5-6) at 730 ÁM but no significant increase at 290 ÁM. These results indicate that sulfide exposure increases oxidative stress, mitochondrial injury (as indicated by increased superoxide production) and oxidative damage to RNA and DNA in a sulfide-adapted animal. It remains to be determined whether free radical damage also occurs during in vivo sulfide exposure.