Human gingival fibroblast cell response to gox/cat

Hydrogen peroxide kills bacteria; however, control of a specific dose limited control over cellular response and function. In this study, the GOX/CAT system was used to independently provide and control the amount of H2O2 and oxygen in cell culture. The GOX/CAT system is composed of glucose oxidase (GOX) and catalase (CAT). Lipopolysaccharide (LPS) is molecules on the outer membrane of gram-negative bacteria. LPS elicits an inflammatory stimulus which can activate macrophages to produce pro-inflammatory cytokines, TNF-a, and specific mediators of inflammation including NO and prostaglandins. Macrophages are known to provide a prominent defense against bacteria. The aim of this experiment was to determine if GOX/CAT (10µMH2O2) plus conditioned media from LPS challenged macrophage cells will alter fibroblast cell numbers, viability and morphology after 24, 48, 78 hours. Human gingival fibroblast (HGF) CRL-2014 were treated with conditioned media from RAW 264.7 cells with 4µg/mL LPS, GOX/CAT (10µMH2O2) and 4µg/mL LPS + GOX/CAT (10µMH2O2) at 24, 48 and 72 hours. Cell protein, MDA and GSH levels and cell morphology were determined after each incubation periods. The results showed no significant difference in protein levels and MDA levels throughout the experiment. Reduced glutathione concentration in treated cells was statistically lower than control suggesting the possibility of intracellular damage. Overall, cell response to H2O2 did not alter cell proliferation and viability of LPS conditioned media treated fibroblast. Morphological results indicated a trend in possible adaptive response to LPS after 48 and 72 hours. Additional experiments are needed to determine chronic exposure on HGF.