Expression of matrix metalloproteinases, cytokines, and connexins in diabetic and nondiabetic human keratinocytes before and after transplantation into an ex vivo wound-healing model.
OBJECTIVE
Wound healing is known to require a well-organized balance of numerous factors, e.g., cytokines, matrix metalloproteinases (MMPs), and their inhibitors, as well as direct cell-cell communication (connexins). Disruption of this balance may lead to the formation of chronic wounds such as diabetic foot ulcers. The transplantation of autologous keratinocytes is a promising therapy for diabetic foot ulcers; however, little is known about their characteristics on a molecular level. Therefore, we intended to characterize transplanted keratinocytes from diabetic and nondiabetic origin before and after transplantation.


METHODS
We isolated human keratinocytes from diabetic and nondiabetic origins and transplanted them into an ex vivo wound healing model. To characterize the keratinocytes, we investigated mRNA expression of MMP-1, MMP-2, and MMP-9; tissue inhibitor of MMP (TIMP)-1 and TIMP-2; interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha; Cx26 (connexin 26) and Cx43; and, for connexins, immunolocalization.


RESULTS
We found no significantly increased expression of the molecules investigated in cultured keratinocytes from diabetic compared with nondiabetic origin, even though there were significant differences for MMP-2, IL-1beta, and TNF-alpha in skin biopsies. Expression of IL-1beta was significantly lower in keratinocytes from diabetic origin. In the course of wound healing, differences in the dynamics of expression of MMP-1, IL-1beta, and Cx43 were observed.


CONCLUSIONS
Our results suggest that keratinocytes from diabetic origin are as capable for transplantation into chronic wounds as keratinocytes from healthy origin at the starting point of therapy. However, differences in expression dynamics later on might reflect the systemic influence of diabetes resulting in a memory of the transplanted keratinocytes.
Brandner
Johanna M
JM
Department of Dermatology and Venerology, University Hospital Hamburg-Eppendorf, Hamburg, Germany.
Zacheja
Steffi
S
Houdek
Pia
P
Moll
Ingrid
I
Lobmann
Ralf
R
D000328:Adult; D000368:Aged; D001706:Biopsy; D018929:Cell Culture Techniques; D000072259:Connexin 26; D017630:Connexins / Q000235:genetics*; D016207:Cytokines / Q000235:genetics*; D017931:DNA Primers; D003920:Diabetes Mellitus / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D005260:Female; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000473:pathology / Q000502:physiology*; D008297:Male; D020782:Matrix Metalloproteinases / Q000235:genetics*; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012016:Reference Values
D017630:Connexins; D016207:Cytokines; D017931:DNA Primers; C000607042:GJB2 protein, human; D000072259:Connexin 26; D012313:RNA; D020782:Matrix Metalloproteinases