Mechanism for IL-1β-mediated neovascularization unmasked by IL-1β knock-out mice

We have reported that interleukin-1 beta (IL-1β) upregulates cardiac expression of vascular endothelial growth factor (VEGF) and VEGF receptor-2 (VEGFR-2), raising the possibility that IL-1β plays an important role in VEGF-mediated neovascularization. In this study, we examined the cellular mechanism for ischemia-induced neovascularization using IL-1β knock-out (–/–) mice. Recovery of blood perfusion in ischemic hindlimb in IL-1β–/– mice was markedly (43% decrease) impaired as compared with the wild-type mice. CD31+ vessel numbers and Ki-67+ neo-capillaries were significantly (P < 0.01) decreased 44% and 68%, respectively. IL-1β expression was localized in the capillary vessels in ischemic limb muscles. Ischemia-induced expressions of hypoxia-inducible factor 1 alpha (HIF-1α), VEGF, its receptor VEGFR-2 and vascular cell adhesion molecule-1 (VCAM-1) were markedly inhibited in the IL-1β–/– mice. Hindlimb ischemia-induced an increase (1.22% out of total nuclear cell) in CD34–/B220–/CD3–/Flk-1+ hematopoietic stem cell population in peripheral blood in the wild-type mice, whereas in the IL-1β–/– mice such increase was only 0.09%. Injection of IL-1β protein into the wild-type mice markedly increased the ratio of the CD34–/B220–/CD3–/Flk-1+ cell population (from 0.03% to 0.7%) in the peripheral blood associated with an increase in the number of endothelial cells. Such IL-1β-mediated increases in cell numbers were blocked by co-injection of anti-VEGF antibody. CD34–/B220–CD3–Flk-1+ cells trans-differentiated into eNOS- and CD31-expressing endothelial cells in vivo and in vitro. This study demonstrates that IL-1β plays a key role in ischemia-induced neovascularization by mobilizing CD34–/B220–CD3–Flk-1+ endothelial precursor cells in a VEGF-dependent manner as well as by upregulating expressions of VEGF, VEGFR-2 and adhesion molecules on endothelial cells.