Antiapoptotic Effect of Gene Therapy with Recombinant Adenovirus Vector Containing Hypoxia-inducible Factor-1α after Cerebral Ischemia and Reperfusion in Rats.

BACKGROUND: Mounting evidence has demonstrated that hypoxia-inducible factor-1α (HIF-1α) could attenuate brain injuries after cerebral ischemia and reperfusion (CIR). However, few reports have addressed the therapeutic efficacies of a recombinant adenovirus vector containing HIF-1α (AdHIF-1α) gene after ischemia and reperfusion. The aim of this study was to examine the antiapoptotic and neuroprotective effects of AdHIF-1α gene for cerebral injuries after ischemia and reperfusion in rats.

METHODS: From February to December 2016, male Sprague-Dawley rats were randomly divided into normal, sham, CIR, AdHIF-1α, and recombinant adenovirus (Ad) groups. Middle cerebral artery occlusion model was established by Longa's method and reperfusion resumed at 2 h postocclusion. AdHIF-1α solution, Ad solution, and phosphate-buffered saline were injected into the right lateral ventricle of rats in AdHIF-1α, Ad, and CIR groups. Brain tissue sections were observed under fluorescent microscope to confirm the definite expression of recombinant adenovirus in Ad and AdHIF-1α groups. The expressions of HIF-1α protein were analyzed by immunohistochemical staining at 6 h, 24 h, and 72 h postreperfusion. Brain water content and neurological deficit scores were evaluated at 6 h, 24 h, and 72 h postreperfusion. Pathological brain injuries were examined after hematoxylin and eosin stain and nerve cell apoptosis was measured after terminal-deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) stain at 72 h postreperfusion. Comparisons were conducted with one-way analysis of variance by post hoc Scheffe's test among different experimental groups.

RESULTS: Green fluorescent protein was successfully expressed in brain tissue of Ad and AdHIF-1α groups from 24 h to 21 days postinjection. As detected by immunohistochemical staining, the expressions of HIF-1α protein were obviously enhanced in AdHIF-1α group than those in CIR and Ad groups at 24 h and 72 h postreperfusion, respectively. There were significant reductions of brain water content (78.83% ± 0.34% vs. 83.21% ± 0.50% and 83.35% ± 0.32%; 84.13% ± 0.24% vs. 89.76% ± 0.34% and 89.70% ± 0.18%; respectively; all P < 0.05) and neurological deficit scores (2.90 ± 0.74 vs. 3.50 ± 0.52 and 3.60 ± 0.53 at 24 h; 2.40 ± 0.84 vs. 3.60 ± 0.52 and 3.50 ± 0.53 at 72 h; respectively; all P < 0.05) in AdHIF-1α group versus CIR and Ad groups at 24 h and 72 h postreperfusion, respectively. The pathologic changes of AdHIF-1α group were milder than those in CIR and Ad groups at 72 h postreperfusion. The percentage of TUNEL-positive cells in cerebral subcortex decreased significantly in AdHIF-1α group versus CIR and Ad groups at 72 h postreperfusion (P < 0.05).

CONCLUSION: AdHIF-1α has an obvious neuroprotective effect on ischemia and reperfusion in rat brains possibly through inhibiting the apoptosis of nerve cells.