2) Comparative toxicity of antimicrobial agents on transformed human keratinocytes

Topical antibiotics and antiseptics are widely used to prevent and treat bacterial infections in wounds and ulcers. Experimental data suggest that some antimicrobial agents may have an adverse effect on several aspects of the tissue repair process, including retardation of wound epithelialization. The aim of this study was to investigate the comparative cytotoxic effects of a range of antiseptics and antibiotics using human keratinocytes transformed by Simian virus 40 (SVK14 cells). SVK14 cells were grown to semi-confluence by adding 6 × 1014 cells per petri dish. After 48 h the cells were exposed to serial dilutions of the therapeutic concentrations (shown in parentheses) of each of the following agents; hydrogen peroxide (3%), cetrimide (1%), sodium hypochlorite (o·5%), povidone iodine (4%), neomycin (1%), bacitracin (50 units/ml), polymyxin B sulphate (10000 units/ml). The cells were exposed to the drug for 15 min and then washed and incubated in the culture medium (RPMI 1640 plus 10% foetal calf serum) for 24 h. Dead cells were then washed off and the viable adherent cells were trypsinized and counted in a Coulter counter. At therapeutic concentrations none of the antibiotics was found to be cytotoxic, whereas all of the antiseptics produced 100% killing of SVK14 cells. Dilutions of therapeutic concentrations of the antiseptics ranging between 1000- and 20000-fold (depending on the agent) were needed in order to achieve no-effect levels. Calculations based on 100% killing values for the antiseptics indicate that their order of toxicity from highest to lowest is: sodium hypochlorite, cetrimide, povidone iodine and hydrogen peroxide. Whilst it is appreciated that more work is needed in order to assess the exact relevance of these findings to clinical practice, it should be noted that SVK14 cells share many of the characteristics of normal keratinocytes and that these preliminary results are broadly in line with in vitro, in vivo and human volunteer findings made by other investigators studing antimicrobial toxicity profiles. We conclude that this cell line may be useful in studying the epithelial cytotoxicity of drugs in vitro and that care should be exercised in the selection of antimicrobial agents for use in wound management.