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JOURNAL ARTICLE
RESEARCH SUPPORT, NON-U.S. GOV'T
Effect of small interference RNA targeting HIF-1alpha mediated by rAAV combined L: -ascorbate on pancreatic tumors in athymic mice.
Pathology Oncology Research : POR 2009 March
To study the effect of recombinant adeno-associated virus (rAAV) vector bearing small inference RNA (siRNA) targeting hypoxia inducible factor 1alpha (HIF-1alpha) combined L: -ascorbate on pancreatic tumors in athymic mice primarily. A cassette encoding siRNA targeting HIF-1alpha mediated by rAAV was constructed, giving rAAV-siHIF. In vitro, rAAV-hrGFP, rAAV-siHIF and L: -ascorbate which were used alone or in combination were delivered to exponentially growing MiaPaCa2 cells. Then, we examined the expression of HIF-1alpha mRNA and protein, the secretion of VEGF in MiaPaCa2 cells under hypoxic condition with Real-time PCR, Western Blot, ELISA, respectively. In vivo, MiaPaCa2 cells were inoculated subcutaneously on the back of nude mice. Nude mice with xenograft tumor were randomly divided into equal groups and were injected with rAAV-hrGFP or rAAV-siHIF or were fed with L: -ascorbate. Then, we measured the size of tumor every 3 days and drew a tumor growth curve. After 30 days, all mice were sacrificed and the tumors were dissected. At last, we examined the expression of HIF-1alpha, VEGF and CD34 by immunohistochemistry and counted micro-vessel density (MVD). In vitro, we found that rAAV-siHIF could inhibit the expression of HIF-1alpha mRNA and protein in MiaPaCa2 human pancreatic cancer cells but L: -ascorbate could only restrain the expression of HIF-1alpha protein. Moreover, rAAV-siHIF and L: -ascorbate could all inhibit the secretion of vascular VEGF. In vivo, we found that rAAV-siHIF could inhibit the growth of nude mice xenograft tumor and the expression of HIF-1alpha and VEGF and MVD while L: -ascorbate can only inhibit the growth of xenograft tumor in the early and middle stage. These results suggest that rAAV-siHIF and L: -ascorbate can inhibit the growth of nude mice xenograft tumor and HIF-1alpha could be a target of pancreatic cancer genetic and pharmacological therapy.
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