Multiple effects of 2ME2 and D609 on the cortical expression of HIF‐1α and apoptotic genes in a middle cerebral artery occlusion‐induced focal ischemia rat model

Despite 2‐methoxyestradiol (2ME2) and tricyclodecan‐9‐yl‐xanthogenate (D609) having multiple effects on cancer cells, mechanistically, both of them down‐regulate hypoxia‐inducible factor‐1α (HIF‐1α) and vascular endothelial growth factor (VEGF). We hypothesize HIF‐1α plays an essential role in cerebral ischemia as a pro‐apoptosis regulator; 2ME2 and D609 decrease the levels of HIF‐1α and VEGF, that might contribute to protecting brain from ischemia injury. A total of 102 male Sprague–Dawley rats were split into five groups: sham, middle cerebral artery occlusion (MCAO), MCAO + dimethyl sulfoxide, MCAO + 2ME2, and MCAO + D609. 2ME2 and D609 were injected intraperitoneally 1 h after reperfusion. Rats were killed at 24 h and 7 days. At 24 h, 2ME2 and D609 reduce the levels of HIF‐1α and VEGF (enzyme‐linked immunosorbent assay), depress the expression of HIF‐1α, VEGF, BCL2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3) and cleaved caspase 3 (western blot and immunohistochemistry) in the brain infarct area. Double fluorescence labeling shows HIF‐1α positive immunoreactive materials are co‐localized with BNIP3 and terminal deoxynucleotidyl transferase biotin‐dUTP nick end labeling inside the nuclei of neurons. At 7 days, 2ME2 and D609 reduce the infarct volume (2,3,7‐triphenyltetrazolium chloride) and blood–brain barrier extravasation, decrease the mortality and improve the neurological deficits. In conclusion, 2ME2 and D609 are powerful agents to protect brain from cerebral ischemic injury by inhibiting HIF‐1α expression, attenuating the superfluous expression of VEGF to avoid blood–brain barrier disruption and suppressing neuronal apoptosis via BNIP3 pathway.

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