Long-term VEGF-A expression promotes aberrant angiogenesis and fibrosis in skeletal muscle

Vascular endothelial growth factor A (VEGF-A) induces strong angiogenesis and it has been widely used in proangiogenic gene therapy studies. However, little is known about long-term effects of VEGF-A expression in skeletal muscle. Here the long- term effects of adeno-associated virus (AAV) encoding human VEGF-A(165) (AAV-VEGF-A) gene transfer in normal and ischemic rabbit hindlimb skeletal muscles were studied. AAV-LacZ was used as a control. In one-year follow-up, a remarkable increase in skeletal muscle perfusion compared with AAV-LacZ was observed measured with Doppler and contrast pulse sequence ultrasound. Angiogenesis was also seen in histology as enlarged and sprouting capillaries. In addition to favorable angiogenic effects, aberrant vascular structures with CD31 positive cell layers were seen inside muscle fibers after AAV-VEGF-A gene transfer. Importantly, we found increased amounts of extracellular matrix with a high number of macrophages and fibrosis in AAV-VEGF-A transduced muscles. No changes in skeletal muscle morphology were detected in AAV-LacZ transduced muscles. Our results indicate that local AAV-VEGF-A gene transfer efficiently promotes long-term angiogenesis in large animal model. However, non-regulated expression of VEGF-A causes unfavorable changes in muscle morphology, which suggests the need for regulation of the transgene expression in long-term AAV-mediated VEGF-A gene transfer applications.