121. VEGF-Induced Angiogenesis Does Not Have a Favorable Effect on Muscle Perfusion: A Relevant Role of Long-Term Angiopoietin-1 Expression in the Formation of Functional Blood Vessels
Authors: Serena Zacchigna, Ennio Tasciotti, Nikola Arsic, Oreste Sorace, Marini Cecilia, Silvia Pardini, Debora Petroni, Claudia Kusmic, Lucia Pattarini, Silvia Moimas, GianMario Sambuceti, Mauro Giacca
Journal: Molecular Therapy (2005)

Top of pageAbstract
While the ability of VEGF to drive a powerful angiogenic response is widely recognized, the central question as to whether the newly formed vessels support a physiological increase in tissue perfusion and meet the increased flow demand during exercise remains, as does the question whether other factors, such as the angiopoietins, might be required to promote further vessel maturation. To explore these issues, we exploited AAV vectors for the prolonged expression of VEGF165 and Ang1 in the skeletal muscle.

Eighty-four rats were injected in their right hindlimb with AAV-LacZ, AAV-VEGF, AAV-Ang1, or AAV-VEGF+AAV-Ang1 (n=21 for each group, 4|[times]|1011 particles for each vector). Muscular blood flow (MBF) and vascular permeability were assessed at 1, 2 and 4 months using PET and 13N-ammonia (3.7 Mbq iv), under both resting conditions and after 15 min of muscle activity, induced by electrical stimulation at 180 spikes/min.

Resting perfusion did not vary significantly in the animals injected with AAV-VEGF alone; surprisingly, in the same animals, perfusion after exercise markedly decreased over time (0.70|[plusmn]|0.13, 0.66|[plusmn]|0.02, 0.50|[plusmn]|0.02 ml/min/g at 1, 2 and 4 months respectively, vs. 0.89|[plusmn]|0.03 control; p<0.05 at every time point). In contrast, the simultaneous injection of VEGF165 and Ang1 determined a marked increase in resting perfusion at 1 month (0.61|[plusmn]|0.34 vs. 0.33|[plusmn]|0.05; p<0.05) and, most notably, a significant improvement after exercise (1.20|[plusmn]|0.24 vs. 0.82|[plusmn]|0.06; p<0.01) that persisted over time (1.17|[plusmn]|0.42 and 1.16|[plusmn]|0.09 at 2 and 4 months, respectively; p<0.01 vs. controls at both times). At all time points, Ang1 alone had no apparent effect on muscle perfusion.

The worsening of perfusion with VEGF alone contrasted with its strong angiogenic effect (an 8.2-fold increase in vessel number at histological examination; p<0.01) and the 6.2-fold apparent increase in the vascular volume as detected by the injection of fluorescent microspheres (p<0.01). We provided two possible explanations for this paradox, by showing that VEGF, besides its potent permeabilizing effect (a 6-fold increase in vascular permeability detected by 99Tc-DTPA injection followed by SPECT imaging and confirmed by the Miles test), promotes a consistent formation of artero-venous shunts, as detected by the injection of a bolus of 99Tc-microspheres (50-100 |[mu]|m diameter range) and subsequent SPECT imaging. Both the leakiness and the artero-venous shunting were significantly counteracted by the co-expression of Ang-1.

Taken together, these results contest the value of VEGF as the sole factor to induce angiogenesis and prompt the use of factor combinations to achieve competent blood vessel formation.