Inhibition of keratinocyte growth in cell culture and whole skin culture by mast cell mediators

Mast cells are suggested to participate in regenerative processes, but thei r influence on epithelialization and wound healing has not been well studie d. Since mast cells can be found in contact with epidermis in chronic infla mmatory skin diseases and venous ulcers, the effect of mast cells on kerati nocyte growth was studied. Keratinocytes were cultured in serum-free condit ions with (complete medium) or without (basal medium) epidermal growth fact or (BPE) and bovine pituitary extract (BPE) to reach subconfluence in a 24- well plate, and the cells were treated with different mast cell mediators h istamine, heparin and tryptase, or lysate from HMC-1 cells, a human leukemi c mast cell line. Whole skin cultures were used as a model for in vitro wou nds to study the effect of mast cells on epithelial outgrowth from skin spe cimens. Histamine inhibited H-3-thymidine incorporation of keratinocytes do se-dependently by 29% at 1 mM, and 89% at 5 mM histamine. In whole skin cul ture, histamine inhibited epithelial outgrowth dose-dependently by 64% alre ady at 0.1 mM histamine and maximally (91%) at 1 mM histamine. Heparin inhi bited H-3-thymidine incorporation dose-dependently by up to 33% at 2 mug/ml in the absence, but not in the presence, of EGF/BPE. In contrast, in whole skin culture, heparin first inhibited the epithelial outgrowth by up to 27 % at 2 mug/ml, but then reversed the inhibition to 30% stimulation at 200 m ug/ml. Skin tryptase (0.0285 to 2.85 mug/ml) with or without heparin (0.5 t o 20 mug/ml) did not affect thymidine incorporation in keratinocytes. Lysat e from HMC-1 cells, but not that from control, neuroblastoma cells, inhibit ed 3H-thymidine incorporation in keratinocytes dose-dependently, and maxima l (47%) inhibition was reached with 16,700 lysed HMC-1 cells/ml. In whole s kin culture, HMC-1 lysate inhibited the epithelial outgrowth by up to 36% a t 67,000 lysed cells/ml. The results show that mast cells and their mediato rs are inhibitory to keratinocyte H-3-thymidine incorporation and epithelia l outgrowth in vitro, although, the inhibitory effect of histamine was seen at high concentrations suggesting a requirement for close morphologic vici nity of mast cells to keratinocytes. Thus, mast cells are assumed to contro l epidermal regeneration and to impair epithelialization of chronic ulcers.