Angiostatin-like fragments are produced in a mouse model of chronic inflammation, suggesting that chronic inflammation and the regulation of angiogenesis may be linked (Falcone et al. 1998). Angiostatin is an anti-angiogenic molecule consisting of kringle domains 1-4 (K1-4) of plasminogen. Leg ulcers are associated with a chronic inflammatory state and we have previously shown that angiostatin-like fragments are generated by incubating leg ulcer exudate with plasminogen (Hoffman et al. 1998). In this study, we have investigated if anti-angiogenic fragments of plasminogen such as angiostatin are present in leg ulcer exudate.

Leg ulcer exudates were collected from patients following informed consent (ethics committee approval was obtained). Angiostatin-related compounds were resolved by western blotting using a polyclonal anti-plasminogen antibody and a monoclonal anti-angiostatin antibody. Wound exudate was fractionated on a lysine-Sepharose column and anti-angiogenic activity was assayed by monitoring tubule formation by human umbilical vein endothelial cells (HUVECs) plated on matrigel.

Angiostatin, as shown using both a polyclonal anti-plasminogen antibody and a monoclonal anti-angiostatin antibody, was detected in all four exudates examined, whereas the related plasminogen fragment K1-3 was present in two out of four exudates. Angiostatin and K1-3 were resolved by lysine-Sepharose chromatography and fractions were assayed for anti-angiogenic activity using the rate at which HUVEC tube formation occurred when plated onto Matrigel. Tube formation was assessed against a relative scale. Fractions containing angiostatin were anti-angiogenic, reducing the rates of tube formation up to fourfold over control, but fractions containing K1-3 had less anti-angiogenic activity, reducing the rate of tube formation less than twofold.

Using the example of leg ulcers, we have demonstrated that chronic inflammation in humans can be associated with the presence of the anti-angiogenic compound angiostatin.

Falcone, D.J., Khan, K.M., Layne, T. & Fernandes, L. (1998). J. Biol. Chem. 273 (47), 31480-31485.

Hoffman, R., Starkey, S. & Coad, J. (1998). J. Invest. Dermatol. 111 (6), 1140-1144.