Cytoprotective and proangiogenic activity of Ex-vivo netrin-1 transgene overexpression protects the heart against ischemia/reperfusion injury

Shazia Durrani, Khawaja Husnain Haider, Rafeeq P.H. Ahmed, Shujia Jiang, Muhammad Ashraf

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

In continuation of a previous work that transgene expression of sonic hedgehog promoted neo-vascularization via netrin-1 release, the current study was aimed at assessing the anti-apoptotic and pro-angiogenic role of netrin-1 transgene overexpression in the ischemic myocardium. pLP-Adeno-X ViralTrak vectors containing netrin-1 cDNA amplified from rat mesenchymal stem cells (Ad-netrin) or without a therapeutic gene (Ad-null) were constructed and transfected into HEK-293 cells to produce Ad-netrin and Ad-null vectors. Sca-1+-like cells were isolated and propagated in vitro and were successfully transduced with Ad-netrin transduced Sca-1+ cells ( NetSca-1+) and Ad-null transduced Sca-1+ cells (NullSca-1+). Overexpression of netrin-1 in NetSca-1+ was confirmed by reverse transcription- polymerase chain reaction and western blot. Neonatal cardiomyocytes and rat endothelial cells expressed netrin-1 specific receptor Uncoordinated-5b and the conditioned medium from NetSca-1+ cells was protective for both the cell types against oxidant stress. For in vivo studies, the rat model of myocardial ischemia/reperfusion injury was developed in female Wistar rats by left anterior descending coronary artery occlusion for 45 min followed by reperfusion. The animals were grouped to receive 70 μL of Dulbecco's modified Eagle's medium without cells (group-1), containing 2×106 NullSca-1+ cells (group-2) and NetSca-1 + cells (group-3). NetSca-1+ cells significantly reduced ischemia/reperfusion injury in the heart and preserved the global heart function in group-3 (P<0.05 vs. groups-1 and group-2). Ex-vivo netrin-1 overexpression in the heart increased NOS activity in the heart. Blood vessel density was significantly higher in group-3 (P<0.05 vs. controls). We concluded that netrin-1 decreased apoptosis in cardiomyocytes and endothelial cells via activation of Akt. Netrin-1 transgene expression was proangiogenic and effectively reduced ischemia/reperfusion injury to preserve global heart function.

Original languageEnglish (US)
Pages (from-to)1769-1778
Number of pages10
JournalStem Cells and Development
Volume21
Issue number10
DOIs
StatePublished - Jul 1 2012
Externally publishedYes

ASJC Scopus subject areas

  • Hematology
  • Developmental Biology
  • Cell Biology

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