Assessment of differential cytokine effects on angiogenesis using an in vivo model of cutaneous wound repair.

Angiogenesis, or new blood vessel formation, has been a subject of intense investigation in recent years. A major obstacle in this research has been the selection of an appropriate in vivo model with which comparisons to humans can be made as well as a reliable quantitative method. Using the porcine excisional wound healing model, we report a new and simple technique for obtaining objective assessments of the microvascular compartment. Factor VIII immunostaining of histological specimens was utilized for specific identification of endothelium devoid of background interference. This technique was coupled with morphometric analysis to quantitate the differential effects of tumor necrosis factor alpha (TNF alpha), transforming growth factor beta (TGF beta), basic fibroblast growth factor (bFGF), insulin-like growth factor-1 (IGF-1), and epidermal growth factor (EGF) within healing porcine wounds. All cytokines stimulated angiogenesis, with low dose TNF alpha and bFGF treatments exhibiting the most profound effects at 7 days postwounding. With increasing levels of TNF alpha (1 ng, 10 ng, 100 ng, and 2.5 micrograms), a step-wise decrease in microvascular area was noted. Although no significant dose responsive differences in angiogenesis were noted following bFGF treatments, a profound increase in capillary area was shown. Significant yet less dramatic increases were noted in capillary area following treatment with EGF or IGF-1. Comparison of the angiogenic effects of TGF beta at 7 and 10 days postwounding showed a significant decrease in the microvasculature as wounds matured. Our data are consistent with previous qualitative in vitro and in vivo reports, thereby confirming the validity of this new model. The data furthermore provide the first quantitative evidence of differential angiogenic responses to cytokines within a clinically relevant model of cutaneous wound repair.