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Angiogenesis in ischemic tissue is a complex and multi-gene event. In the study, we constructed hypoxic re-sponse elements (HRE) and the Tet-On advanced double-controlled systems and investigated their effects on the expression of hVEGF165 and angiopoietin-1 (Ang-1) genes in rat cardiomyocytes exposed to hypoxia and pharma-cologic induction. We infected neonatal rat cardiomyocytes with recombinant rAAV-rtTA-Rs-M2/rAAV-TRE-Tight-Ang-1 and rAAV-9HRE- hVEGF165. Our results indicated that the viral titer was 1×1012 vg /mL and the viral purity exceeded 98%. hVEGF165 expression was induced by hypoxia, but not by normoxia (P < 0.001). Ang-1 expression was evident under doxycycline induction, but undetectable without doxycycline induction (P < 0.001). Immunofluorescence staining showed that positively stained hVEGF165 and Ang-1 protein appeared only under both hypoxia and doxycycline induction. We demonstrate here that HRE and the recombinant Tet-On advanced double gene-controlled systems sensitively regulate the expression of hVEGF165 and Ang-1 genes in an altered oxygen environment and under pharmacological induction in vitro.
In the study, we constructed hypoxic re-sponse elements (HRE) and the Tet-On advanced double-controlled systems and investigated their effects on the expression of hVEGF165 and angiopoietin-1 We in neonatal rat cardiomyocytes with recombinant rAAV-rtTA-Rs-M2 / rAAV-TRE-Tight-Ang-1 and rAAV-9HRE-hVEGF165. Results indicated that the viral titer was 1 × 1012 vg / mL and the viral purity exceeded 98%. hVEGF165 expression was induced by hypoxia but not by normoxia (P <0.001). Ang-1 expression was evident under doxycycline induction, but undetectable Without doxycycline induction (P <0.001). Immunofluorescence staining showed that positively stained hVEGF165 and Ang-1 protein had only under both hypoxia and doxycycline induction. We demonstrate here that HRE and the recombinant Tet-On advanced double gene-controlled systems sen sitively regulate the expression of hVEGF165 and Ang-1 genes in an altered oxygen environment and under pharmacological induction in vitro.