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Adeno-associated viral vector-mediated hypoxia-regulated VEGF gene transfer promotes angiogenesis following focal cerebral ischemia in mice

Gene Therapy volume 15, pages 30–39 (2008)Cite this article

Abstract

Uncontrolled expression of vascular endothelial growth factor (VEGF) in vivo may cause unexpected side effects, such as brain hemangioma or tumor growth. Because hypoxia-inducible factor-1 (HIF-1) is upregulated during cerebral ischemia and regulates gene expression by binding to a cis-acting hypoxia-responsive element (HRE), we therefore used a novel HRE, originating in the 3′-end of the erythropoietin (Epo) gene, to control gene expression in the ischemic brain. A concatemer of nine copies (H9) of the consensus sequence of HRE was used to mediate hypoxia induction. Three groups of adult CD-1 mice received AAVH9-VEGF, AAVH9-lacZ or saline injection, and then underwent 45 min of transient middle cerebral artery occlusion (tMCAO). Results show that HIF-1 was persistently expressed in the ischemic brain. VEGF was overexpressed in the ischemic perifocal region in AAVH9-VEGF-transduced mice. Double-labeled immunostaining showed that VEGF expressed in neurons and astrocytes but not endothelial cells, suggesting that adeno-associated virus (AAV) vectors transduced neurons and astrocytes predominantly. The total number of microvessels/enlarged microvessels was greatly increased in the AAVH9-VEGF-transduced mice (180±29/27±4) compared to the AAVH9-lacZ (118±19/14±3) or saline-treated (119±20/14±2) mice after tMCAO (P<0.05). Cell proliferation examination demonstrated that these microvessels were newly formed. Regional cerebral blood flow recovery in the AAVH9-VEGF-transduced mice was also better than in AAVH9-lacZ or saline-treated mice (P<0.05). Our data indicated that HRE is a novel trigger for the control of VEGF expression in the ischemic brain. VEGF overexpression through AAVH9-VEGF gene transfer showed stable focal angiogenic effects in post-ischemic repair process, providing an opportunity to rebuild injured brain tissue.

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Abbreviations

AAV:

adeno-associated viral vector

AAV-VEGF:

AAV encoding VEGF165 gene

AAV-lacZ:

AAV encoding β-galactosidase gene

PCNA:

proliferating cell nuclear antigen

VEGF:

vascular endothelial growth factor

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Acknowledgements

These studies were supported by NIH Grants: R01 NS027713 (WLY), R21 NS050668 (GYY) and P01 NS044145 (WLY, GYY). We thank Voltaire Gungab for editorial assistance, and the staff of the Center for Cerebrovascular Research (http://avm.ucsf.edu/) for their collaborative support.

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Authors and Affiliations

  1. Department of Anesthesia and Perioperative Care, Center for Cerebrovascular Research, University of California, San Francisco, CA, USA

    F Shen, Y Fan, Y Zhu, Y Chen, W Liu, W L Young & G-Y Yang

  2. Department of Neurology, Rui-Jin Hospital, Jiaotong University, Shanghai, China

    F Shen & G-Y Yang

  3. Department of Internal Medicine, University of California, San Francisco, CA, USA

    H Su

  4. Department of Neurological Surgery, University of California, San Francisco, CA, USA

    W L Young & G-Y Yang

  5. Department of Neurology, University of California, San Francisco, CA, USA

    W L Young

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Correspondence to G-Y Yang.

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Shen, F., Fan, Y., Su, H. et al. Adeno-associated viral vector-mediated hypoxia-regulated VEGF gene transfer promotes angiogenesis following focal cerebral ischemia in mice. Gene Ther 15, 30–39 (2008). https://doi.org/10.1038/sj.gt.3303048

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