Biological Evaluation of 2-Hydroxyethylmethacrylate (HEMA) Toxicity in Human Gingival Fibroblasts with Histochemical X-ray Microanalysis

Purpose: To analyze the cytotoxic effects of 2-hydroxyethylmethacrylate (HEMA) in human gingival fibroblasts using quantitative x-ray microanalysis (EXPMA) and two classical methods (DNA and LDH release in culture medium). Materials and Methods: Different concentrations of HEMA (5, 10, 20, 30, and 40 mM) in DMEM medium were used and the effects on human gingival fibroblasts after 6, 12, and 24 h were determined. As controls, fibroblasts cultured with DMEM culture medium (negative control) and fibroblast incubated in 1% triton X (positive control) were used. Results: The results showed that correlation between the concentrations of HEMA and the amount of LDH and DNA released to the medium were statistically significant for all times analyzed. LDH and DNA released from cells incubated in the lowest concentrations of HEMA (5 and 10 mM) were not significantly different to negative controls. In contrast, cells incubated in the highest HEMA concentrations (20, 30, 40 mM) showed a significant increase of both LDH and DNA released to the culture medium at 6, 12, and 24 h. On the other hand, the ionic concentration of the different elements analyzed in this work revealed that the contents of P, S, Cl, and K were significantly higher in the controls than in samples incubated for 6 h in 5 mM or 10 mM HEMA (p 0.01). K/ Na index (an excellent marker of cell viability) showed a significant decrease, and therefore, viability was significantly reduced. Conclusion: The results suggest that EXPMA is a sensitive method that is able to detect early cell damage even before the cell membrane is altered. Keywords: HEMA, cell death, apoptosis, necrosis, microanalysis, LDH, DNA