Real-time monitoring of keratin 5 expression during burn re-epithelialization.
BACKGROUND
Keratin is a major protein produced during epithelialization following burn injury and is a useful marker for assessing wound healing. Transgenic mice expressing enhanced green fluorescent protein (EGFP) driven by the keratin 5 (K5) promoter (K5GFP mice) were used to monitor keratin expression, and thus, re-epithelialization of burn wounds.


METHODS
K5GFP transgenic mice were created using conventional techniques, with PCR and Southern blot confirmation of transgene incorporation, followed by selection of the line with the most intense and consistent basal epithelial EGFP expression. Epi-fluorescent microscopy of 24 K5GFP mouse flanks and 10 negative littermate controls was used to characterize EGFP intensity, before wounding and serially for 30 days after administration of a standardized burn wound and excision. Biopsy sections of K5GFP and negative control mice were stained with K5 antibody and imaged with confocal microscopy to characterize the distribution of EGFP and K5 at baseline and after injury and to examine the correlation between K5 expression and EGFP expression during healing.


RESULTS
Green fluorescence intensity increased at the advancing wound margin of burned K5GFP mice, reaching a maximum between days 12 and 15 post-burn and then decreasing as healing completed. K5 and EGFP expression increased in parallel in burned K5GFP mice as demonstrated by confocal microscopy.


CONCLUSIONS
EGFP expression correlated with K5 expression during wound healing and therefore serves as a good marker of re-epithelialization. This transgenic model allows noninvasive, real-time assessment of in vivo K5 expression and will be useful in the study of wound healing.
Bruen
Kevin J
KJ
Department of Surgery, Chapel Hill, North Carolina 27599-7050, USA.
Campbell
Chris A
CA
Schooler
Wesley G
WG
deSerres
Suzan
S
Cairns
Bruce A
BA
Hultman
C Scott
CS
Meyer
Anthony A
AA
Randell
Scott H
SH
D000818:Animals; D002056:Burns / Q000378:metabolism* / Q000503:physiopathology; D005260:Female; D049452:Green Fluorescent Proteins; D053569:Keratin-15; D053555:Keratin-5; D007633:Keratins / Q000096:biosynthesis*; D008164:Luminescent Proteins; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D023421:Models, Animal; D008991:Monitoring, Physiologic / Q000379:methods; D012867:Skin / Q000378:metabolism* / Q000503:physiopathology; D014945:Wound Healing / Q000502:physiology*
D053569:Keratin-15; D053555:Keratin-5; C492053:Krt15 protein, mouse; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D007633:Keratins