Adenovirus-mediated hypoxia-inducible factor-1 alpha gene transfer induces angiogenesis and neurogenesis following cerebral ischemia in rats

BACKGROUND: Hypoxia-inducible factor-1 (HIF-1) accumulates under conditions of hypoxia. HIF-1α target genes have pleiotropic effects on neurogenesis, neuroprotection and angiogenesis in the brain. OBJECTIVE: To investigate whether a recombinant adenovirus carrying HIF-1α can increase the expres- sion of HIF-1α in vivo and thus promote angiogenesis and neurogenesis in a rat model of focal cerebral ischemia. DESIGN, TIME AND SETTING: The randomized, controlled experiment was performed at the Department of Neurobiology, Third Military Medical University of Chinese PLA from September 2006 to October 2007. MATERIALS: 68 healthy adult male Sprague-Dawley (SD) rats, weighing 230-250 g, were used. HIF-1α antibody was purchased from Wuhan Boster Company. Vascular endothelial growth factor (VEGF) antibody was purchased from Santa Cruz Biotech Company. METHODS: All 68 rats were induced with a transient middle cerebral artery occlusion (MCAO), according to the method of intra-luminal vascular occlusion. 54 rats, in which MCAO was successfully induced, were randomly divided into adenovirus (Ad) group and recombinant adenovirus with HIF-1αgene (Ad-HIF-1α) group (27 rats for each group). Rats were injected with 10 μL Ad (Ad group) or Ad-HIF-1α (Ad-HIF-1α group) into the lateral ventricle, 1 day after MCAO induction. MAIN OUTCOME MEASURES: Reverse transcription polymerase chain reaction was used to measure the expression of HIF-1α and of VEGF. Immunohistochemistry was used to detect the localization of HIF-1α, VEGF and factor Ⅷ in ischemic penumbra. Rat newborn nerve cells were labeled with 5-bromodeoxyuridine (BrdU) after ischemia. BrdU/neurofilament 200 (NF200) and BrdU/glial fibrillary acidic protein (GFAP) double labeled immunofluorescent histochemistry was used to identify the differentiation of newborn cells. Neurological function was evaluated using the modified neurological severity score (NSS). RESULTS: Compared with Ad, Ad-HIF-1αenhanced the expression of HIF-1αand VEGF (P < 0.01). The numbers of factor VIII, BrdU, BrdU/NF200 and BrdU/GFAP positive cells were increased significantly (P < 0.01) in the Ad-HIF-1α group compared to the Ad group. Levels of HIF-1α and VEGF mRNA in the Ad-HIF-1α group were enhanced compared with those in the Ad group. NSS scores of the Ad-HIF-1α group were superior to those of the Ad group at days 7, 14, 21, and 28 after MCAO (P < 0.05). CONCLUSION: HIF-1α gene therapy can increase angiogenesis and neurogenesis, and thus improve neurological function following cerebral ischemia in rats.