THE EFFECTS OF BURN BLISTER FLUID ON CULTURED KERATINOCYTES

Objective: Previous studies have suggested that burn blister fluid (BB F) maybe detrimental to the healing of the underlying wound bed, In th is study, the effects of burn blister fluid on cultured keratinocyte p roliferation and differentiation were examined and quantitated using v arious techniques. Methods: At three different concentrations (2%, 10% , 20% in 20% fetal bovine serum/complete culture medium), 19 BBFs were tested in triplicate using 12 populations of cultured keratinocytes, All BBFs were collected from partial thickness burns within 72 hours o f injury, BBF was added on day 4 of the keratinocyte culture, The effe ct on proliferation and viability was assessed using trypan blue dye e xclusion, Multiparameter flow cytometric analysis was used to quantita te population kinetics and cell size distribution, Keratinocyte differ entiation was determined using immunohistochemical staining of differe ntiation markers and quantitation of cornified envelope formation. Res ults: Relative to control fluid, the BBF caused a variable effect on p roliferation, ranging from 67% inhibition to 103% stimulation, with an overall 4% inhibition, The range of keratinocyte viability was narrow er, with a similar overall 4% reduction. Using flow cytometry to analy ze RNA/DNA content and cell size, The BBF caused a subtle shift in ker atinocyte population kinetics and cell size distribution toward larger , less rapidly dividing cells, The BBF had no significant effect on ex pression of the differentiation markers, filaggrin and involucrin, Fin ally, the BBF did not alter terminal differentiation as it did not inf luence formation of cornified envelopes (BBF = 9.1 +/- 4.8%, control = 9.9 +/- 6.6%). Conclusion: Previous biochemical analysis has shown th at BBF consists primarily of human serum filtrate with locally produce d acute reactants, Our study suggests that BBF is biologically similar to serum and does not significantly alter keratinocyte proliferation or differentiation in vitro.