Fibrocytes from burn patients regulate the activities of fibroblasts

Wound healing requires an elaborate interplay between numerous cell types that orchestrate a series of regulated and overlapping events. Fibrocytes are a unique leukocyte subpopulation implicated in this process. One role proposed for these cells in wound healing is to synthesize extracellular matrix. Interestingly, using mass spectrometry to quantify hydroxyproline, we discovered that the capacity of fibrocytes from normal subjects or from burn patients to produce collagen is much less than that of dermal fibroblasts. Therefore, we investigated whether fibrocytes could play an indirect, regulatory, role in the healing of burn wounds by affecting the functions of dermal fibroblasts. Dermal fibroblasts treated with medium conditioned by burn patient fibrocytes, but not by those derived from normal subjects, showed an increase in cell proliferation and migration. Using confocal microscopy, flow cytometry, and immunoblotting, we found the level of alpha-smooth muscle actin (alpha-SMA) expression to be increased in these treated dermal fibroblasts, which also showed an enhanced ability to contract collagen lattices. To determine whether these effects could be attributed to transforming growth factor beta (TGF-beta 1) or to connective tissue growth factor (CTGF), we measured total TGF-beta 1 levels in the conditioned medium by an enzyme-linked immunosorbtion assay and assessed levels of CTGF mRNA and protein in fibroblasts and fibrocytes by reverse transcription-polymerase chain reaction and Western blotting. The results showed significantly higher levels of TGF-beta 1 and CTGF produced by burn patient fibrocytes. In addition, the application of a TGF-beta 1 neutralizing antibody significantly reduced the effect of burn patient fibrocyte medium on dermal fibroblast proliferation, migration, and collagen lattice contraction. Our results suggest that in healing burn wounds, fibrocytes could regulate the activities of local fibroblasts.