HUMAN RECOMBINANT TRANSFORMING GROWTH FACTOR-BETA-1 MODULATION OF BIOCHEMICAL AND CELLULAR EVENTS IN HEALING OF ULCER WOUNDS

The effects of recombinant human transforming growth factor beta-1 (rhTGF-beta-1) on wound healing were examined in a rabbit ear ulcer model in which rhTGF-beta-1 was applied to full-thickness biopsy ulcers on the ears. The influence of perichondrium on healing was studied by comparing ulcers with and without perichondrium on 1) formation of total healing wound area (HWA, the newly formed connective and granulation tissues within the ulcer) over time and 2) the amount of collagen synthesized by the wound tissue at day 5. The HWA of ulcers with intact perichondrium increased sharply with time and reached a plateau at day 7, whereas a slower healing occurred in the perichondrium-free model where maximal HWA appeared at day 14. Topical application of 100 ng of rhTGF-beta-1 per wound accelerated healing by increasing HWA in both models. The enhancement of healing by rhTGF-beta-1 was associated with increased collagen synthesis. The percent collagen synthesis in the rhTGF-beta-1 treated wound tissue (HWA plus the overlying epithelium) was doubled in the perichondrium-intact ulcers and increased 40% in the perichondrium-free ulcers. DNA synthesis in the perichondrium-intact ulcers was not altered by rhTGF-beta-1 when measured at day 5 by in vitro labeling with [H-3]thymidine ([H-3]TdR). Autoradiography indicated that the primary cells labeled in the wound tissue were epithelial cells and rhTGF-beta-1 enhanced the migration of these cells from the wound margin towards the center. To evaluate the effects of rhTGF-beta-1 on fibroblasts derived from the granulation tissue of the wound, cells were treated with increasing concentrations of rhTGF-beta-1 and DNA and collagen synthesis were determined. rhTGF-beta-1 elicited a biphasic change in percent collagen synthesis with a maximal increase of 50% at 20 pM followed by a decline. A twofold increase in [H-3]TdR incorporation that plateaued at 1 nM was also observed. Our results indicate that the cellular responses to rhTGF-beta-1 differ in vivo and in vitro. The perichondrium-intact ulcers contain more wound tissue and have larger responses to rhTGF-beta-1 stimulation, which allows better examination of biochemical and cellular events. The in vivo mechanisms are multi-factorial, which may involve cell migration and recruitment as results of numerous cell/cell and cell/matrix interactions.