Hydrated keratinocytes reduce collagen synthesis by fibroblasts via paracrine mechanisms.
Regulating collagen metabolism can control hypertrophic scars in cutaneous wounds. Hypertrophic scars can be reduced by occlusive dressings such as silicone sheeting; however, their mechanism is still unknown. We hypothesized that hydration of keratinocytes reduces the collagen secretion of fibroblasts by modifying the cytokine levels. Stratified human epidermal keratinocytes and confluent human dermal fibroblasts were co-cultured serum free for 72 hours. Keratinocytes were either kept at the air interface or hydrated. Messenger RNA (mRNA) levels of interleukin-1 (IL-1)alpha, IL-1beta, tumor necrosis factor alpha (TNF-alpha), keratinocyte growth factor (KGF), and procollagen-1 were analyzed by real-time reverse transcription-polymerase chain reaction. Secretion of cytokines into conditioned media was quantified by enzyme-linked immunosorbent assay and collagen content by Western blot. The content of collagen-I decreased by 44% in the presence of hydrated keratinocytes. Co-culture with air-treated keratinocytes decreased collagen-I only by 23%. Co-cultured hydrated keratinocytes had significantly higher TNF-alpha mRNA (172%) than hydrated keratinocytes. At the protein level, there was an overall trend toward increased TNF-alpha levels in hydrated cultures. IL-1beta secretion decreased significantly under hydration (42% monoculture, 58% co-culture). Co-culture stimulated a 240% increase of KGF mRNA in fibroblasts compared with monocultured fibroblasts. Fibroblasts secreted 4.5-fold more KGF in hydrated co-cultures and sixfold more KGF in air-treated co-cultures. Hydration of keratinocytes modifies important paracrine interactions between keratinocytes and fibroblasts and reduces collagen-1, which supports the hypothesis that hydration of the epidermis and restoration of water barrier function play an important role in scar formation.
Tandara
Andrea A
AA
Wound Healing Research Laboratory, Division of Plastic and Reconstructive Surgery, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.
Kloeters
Oliver
O
Mogford
Jon E
JE
Mustoe
Thomas A
TA
D000328:Adult; D002478:Cells, Cultured; D018920:Coculture Techniques; D003094:Collagen / Q000096:biosynthesis*; D016895:Culture Media, Serum-Free; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D051523:Fibroblast Growth Factor 7 / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007375:Interleukin-1 / Q000032:analysis; D015603:Keratinocytes / Q000502:physiology*; D009779:Occlusive Dressings; D019899:Paracrine Communication / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
D016895:Culture Media, Serum-Free; D007375:Interleukin-1; D014409:Tumor Necrosis Factor-alpha; D051523:Fibroblast Growth Factor 7; D003094:Collagen