Dual roles of tumor necrosis factor-α receptor-1 in a mouse model of hindlimb ischemia

Signals in the tumor necrosis factor alpha (TNF-alpha) pathway are upregulated after ischemia, yet its role in peripheral ischemia remains unclear. We investigated the effect of TNF-alpha receptor 1 (TNFR-1) in acute limb ischemia of TNFR-1 knockout (TNFR-1(-/-)) and wild type (WT, TNFR-1(+/+)) mice. Laser Doppler scanning showed that although pre-ischemia blood flow levels were similar in these mice, the limb reperfusion after ischemia was significantly higher in TNFR-1(-/-) mice 1-7 days after injury. Consistently, fewer TUNEL-positive cells, less DNA fragmentation, and a lower ischemic score were detected in the TNFR-1(-/-) group when compared to WT controls. Western blot analysis revealed less expression of pro-apoptotic markers Bax and cleaved caspase-3 in TNFR-1(-/-) mice 1 day after ischemia, supporting the hypothesis that the absence of TNFR-1 results in a reduction of apoptosis. The rate of post-ischemia amputation was 50% in WT mice versus 0% in TNFR-1(-/-) mice. However, immunohistochemical co-staining of microvessel marker CD31 and cellular proliferation marker BrdU 21 days after ischemia showed an impaired angiogenic activity in the TNFR-1(-/-) mice. These data were supported by Western blot analysis, which indicated a decreased expression of angiopoietin-1 (Ang-1) and its receptor Tie-2 in TNFR-1(-/-) mice. Our results suggest that a deficiency in TNFR-1 prevents the activation of death-related proteins downstream to TNF-alpha and attenuates apoptosis in acute limb ischemia, but the lack of TNFR-1 signaling hinders the belated angiogenesis mediated by the Ang-1/Tie-2 pathway.