Systemic Administration of Thrombin Peptide TP508 Enhances VEGF-Stimulated Angiogenesis and Attenuates Effects of Chronic Hypoxia

Abstract

Revascularization of chronic wounds and ischemic tissue is attenuated by endothelial dysfunction and the inability of angiogenic factors to stimulate angiogenesis. We recently showed that TP508, a nonproteolytic thrombin peptide, increases perfusion and NO-dependent vasodilation in hearts with chronic ischemia and stimulates NO production by endothelial cells. In this study, we investigated systemic in vivo effects of TP508 on VEGF-stimulated angiogenesis in vitro using aortic explants in normoxic and hypoxic conditions. Mice were injected with saline or TP508 and 24 h later aortas were removed and cultured to quantify endothelial sprouting. TP508 injection increased endothelial sprouting and potentiated the in vitro response to VEGF. Exposure of control explants to hypoxia inhibited basal and VEGF-stimulated endothelial cell sprouting. This effect of hypoxia was significantly prevented by TP508 injection. Thus, TP508 systemic administration increases responsiveness of aortic endothelial cells to VEGF and diminishes the effect of chronic hypoxia on endothelial cell sprouting. Studies using human endothelial cells in culture suggest that protective effects of TP508 during hypoxia may involve stimulation of endothelial cell NO production. These data suggest potential clinical benefit of using a combination of systemic TP508 and local VEGF as a therapy for revascularization of ischemic tissue.

Nuclear Countermeasure Activity of TP508 Linked to Restoration of Endothelial Function and Acceleration of DNA Repair

“…Our previous studies showed that TP508 stimulates eNOS to produce NO (28) and reverses effects of hypoxia-induced ED to restore myocardial function (30) and prevent the loss of endothelial cell responses to VEGF (29). Because exposure to radiation has been shown to induce ED (6–8, 12, 13), we conducted studies to determine whether irradiation of HCAEC would affect NO production by endothelial cells in response to VEGF or TP508, which stimulate NO production by two different mechanisms (28).…”

“…injection of TP508 stimulated aortic endothelial cell sprouting and prevented hypoxia-induced loss of endothelial function when aortic explants were removed from nonirradiated mice 24 h after injection and cultured on Matrigel ex vivo (29). Thus, we chose to use this modified aortic sprouting assay to determine whether systemic injection of TP508 could protect endothelial function in vivo after whole-body gamma irradiation.…”

“…Mice were injected through the tail vein 1 h postirradiation with 100 μl of saline alone or 100 μl of saline containing 500, 100 or 20 μg of TP508. Thoracic aortas were isolated 24 h later and aortic explants were cultured ex vivo on Matrigel-coated 24-well plates and cultured in EGM media containing endothelial cell growth factors as previously described (29). …”

“…This peptide drug reversed hypoxia-induced ED to restore VEGF-stimulated NO signaling and VEGF-stimulated angiogenic responses (29). Moreover, TP508 is also effective in reversing ED-related effects of chronic myocardial ischemia (30) and acute myocardial infarct (31, 32).…”

“…Moreover, this effect of TP508 reverses hypoxia-induced ED to restore VEGF responsiveness, NO signaling and aortic endothelial cell sprouting (29). TP508 has also been shown to restore vascular function and prevent ischemic damage in porcine models of chronic myocardial ischemia (30) and acute myocardial infarct (31, 32).…”