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.

Antegrade flow and peripheral resistance determine the level of endogenous arteriogenesis in patients with superficial femoral artery occlusion.

OBJECTIVES: Occlusion in a limb artery leads to impaired blood supply and ischaemia. Collateral artery growth (arteriogenesis) is one of the most effective natural response mechanisms to compensate this pathologic situation. However, it is unknown if clinically important features, like poor run-off, have an impact on compensatory vessel growth. METHODS: Study population of this retrospective study consisted of 70 patients who suffered from lower limb ischaemia and underwent bypass surgery because of an occlusion of the superficial femoral artery. Clinical data were collected and pre- and postoperative angiograms were reviewed. Number of collateral vessels bypassing the occluded segment was counted. Features of inflow and outflow vessels were recorded. RESULTS: The mean number of collaterals was 13 + or - 0.5 per patient. In univariate analysis, short daily walking distance, chronic critical leg ischaemia, low ankle brachial index, low number of patent calf arteries and stenosed inflow arteries predicted low number of collateral arteries. In the multivariate analysis, only the quality of inflow and the number of patent calf vessels demonstrated an independent association (P < 0.05) with the number of collaterals. CONCLUSIONS: Ankle-brachial index, grade of symptoms and daily walking capacity could be used to predict collateral density. Importantly, a good antegrade flow and peripheral runoff seem to have a significant effect on collateral density, implying an impact on the activation of arteriogenesis.

Overexpression of VEGF-A induces neovascularization and increased vascular leakage in rabbit eye after intravitreal adenoviral gene transfer.

AIM: The aim of this study was to determine dose-response effects of vascular endothelial growth factor A as delivered using an adenoviral vector on vascular growth and pathological changes in the rabbit eye. Moreover, we wanted to develop a large animal model for angioproliferative diseases in the eye. METHODS: Seventeen New Zealand White rabbits were injected with adenoviral vascular endothelial growth factor-A (AdVEGF-A) intravitreally with different doses (10(9)-10(11) vp). Controls were injected with an empty virus (AdCMV). Some animals had a combination of AdVEGF-A and AdsKDR (a soluble form of the VEGF receptor-2). Animals were killed 6 days after the gene transfer. On the basis of these results, 14 rabbits were injected intravitreally with AdVEGF-A or adenoviral LacZ (AdLacZ) with 10(10) vp in a volume of 0.1 mL. Animals were killed 3, 6, 14 and 28 days after the gene transfer, eyes were removed and analysed histologically. RESULTS: In enzyme-linked immunosorbent assay (ELISA) analysis, human VEGF-A was present in vitreous humour in all VEGF-A transduced eyes. The amount of VEGF-A showed a dose-dependent increase with the AdVEGF-A dose and was the highest 6 days after the gene transfer. Histologic analyses revealed an increased capillary area and density in the AdVEGF-A eyes when compared with the AdLacZ eyes (P < 0.05). In the AdVEGF-A/AdsKDR eyes the average capillary area was not increased compared with AdLacZ eyes. CONCLUSION: This model could be useful for large animal studies regarding the pathogenesis of neoangiogenesis and for the development of new therapeutic strategies for angioproliferative diseases of the eye. Our results establish the key role of VEGF-A in the induction of neovascularization and pathological changes in the rabbit eye.