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Molecular Imaging of RNA Interference Therapy Targeting PHD2 for Treatment of Myocardial Ischemia

  • Mei Huang
  • Joseph C. Wu
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 709)

Abstract

Coronary artery disease is the number one cause of morbidity and mortality in the Western world. It typically occurs when heart muscle receives inadequate blood supply due to rupture of atherosclerotic plaques. During ischemia, up-regulation of hypoxia inducible factor-1 alpha (HIF-1α) transcriptional factor can activate several downstream angiogenic genes. However, HIF-1α is naturally degraded by prolyl hydroxylase-2 (PHD2) protein. Recently, we cloned the mouse PHD2 gene by comparing the homolog gene in human and rat. The best candidate shRNA sequence for inhibiting PHD2 was inserted behind H1 promoter, followed by a separate hypoxia response element (HRE)-incorporated promoter driving a firefly luciferase (Fluc) reporter gene. This construct allowed us to monitor gene expression noninvasively and was used to test the hypothesis that inhibition of PHD2 by short hairpin RNA interference (shRNA) can lead to significant improvement in angiogenesis and contractility as revealed by in vitro and in vivo experiments.

Key words

RNA interference Molecular imaging Hypoxia inducible factor (HIF) Prolyl hydroxylases (PHD) Ischemic heart disease 

Notes

Acknowledgments

This work was supported in part by grants from the NIH HL095571 (JCW), NIH HL093172 (JCW), and AHA Western Postdoctoral Fellowship (MH).

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Mei Huang
    • 1
    • 2
  • Joseph C. Wu
    • 1
    • 2
  1. 1.Department of MedicineStanford University School of MedicineStanfordUSA
  2. 2.Department of RadiologyStanford University School of MedicineStanfordUSA

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