Inhibition of proliferation of Pseudomonas aeruginosa by KGF in an experimental burn model using human cultured keratinocytes.
Experimental models showed the ability of Pseudomonas aeruginosa to interact with epidermal keratinocytes [Green H, Kehinde O, Thomas J. Growth of cultured human epidermal cells into multiple epithelia suitable for grafting. Proc Natl Acad Sci USA 1979;76(11):5665-8], stimulating these cells to produce specific peptides that start an immunological chain reaction in the epidermis [O'Connor NE, Mulliken JB, Banks-Schlegel S, Kehinde O, Green H. Grafting of burns with cultured epithelium prepared from autologous epidermal cells. Lancet 1981;(1):75-8]. The immune reaction causes the release of cytokines and growth factors. The objective of this study was to test whether the presence of keratinocyte growth factor (KGF) alters P. aeruginosa proliferation in an experimental burn model.


METHODS
Human keratinocytes derived from neonatal foreskins were isolated and cultured following standard methods [Gallico III, GG, O'Connor NE, Compton CC, Kehinde O, Green H. Permanent coverage of large burn wounds with autologous cultured human epithelium. N Engl J Med 1984;311(7):448-51]. Some of these cells were genetically modified to produce KGF, and the other cells were supplemented with different KGF concentrations in the culture media. Both groups of keratinocytes were seeded in collagen matrices and cultured to form stratified epithelia. A hot plate was used to produce burn defects. Each matrix was inoculated with luminescent P. aeruginosa strain. Experiments were made using keratinocytes without KGF, keratinocytes supplemented with different concentrations of KGF, and keratinocytes genetically modified to produce KGF. Statistical analyses were made using Wilcoxon paired test.


RESULTS
When KGF was added to P. aeruginosa in the presence of keratinocytes, bacterial growth was inhibited, and the same was observed when genetically modified keratinocytes were used.


CONCLUSIONS
Many studies have been done on KGF, where its known properties were defined as a mitogen for keratinocytes [Munster AM. Cultured skin for massive burns: a prospective, controlled trial. Ann Surg 1996;224(3):372-7]. This method allows for a qualitative and quantitative evaluation in real time of the bacterial growth in wound sites after bacterial inoculation. KGF was involved in the reduction of bacterial viability. However, as KGF alone did not produce any effect on P. aeruginosa, it seems to modulate the skin innate immune response.
Sobral
C S
CS
Federal University of São Paulo, Division of Plastic Surgery, São Paulo, SP, Brazil. chrisssobral@terra.com.br
Gragnani
A
A
Morgan
J
J
Ferreira
L M
LM
D000818:Animals; D002056:Burns / Q000382:microbiology / Q000473:pathology*; D049109:Cell Proliferation / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D051523:Fibroblast Growth Factor 7 / Q000494:pharmacology*; D006801:Humans; D015603:Keratinocytes / Q000166:cytology; D051379:Mice; D008954:Models, Biological; D041681:NIH 3T3 Cells; D011550:Pseudomonas aeruginosa / Q000254:growth & development*
D051523:Fibroblast Growth Factor 7