No abstract

Blood-brain barrier (BBB) breakdown after stroke is linked to the up-regulation of metalloproteinases (MMPs) and inflammation. This study examines the effects of progesterone (PROG) and its neuroactive metabolite allopregnanolone (ALLO) on BBB integrity following permanent middle cerebral artery occlusion (pMCAO). Rats underwent pMCAO by electro-coagulation and received intraperitoneal injections of PROG (8 mg/kg), ALLO (8 mg/kg) or vehicle at 1 h post-occlusion and then subcutaneous injections (8 mg/kg) at 6, 24, and 48 h. MMP activation and expression were analyzed by Western blot, immunohistochemistry and gelatin zymography 72 h post-pMCAO. Occludin1, claudin5, tumor necrosis factor-alpha (TNF-α) and Interleukin-6 (IL-6) were analyzed at 72 h post-pMCAO with Western blots. BBB permeability was measured by Evans blue extravasation and infarct size was evaluated by cresyl violet at 72 h after pMCAO. Ischemic injury significantly (p<0.05) increased the expression of MMP-9, MMP-2, TNF-α and IL-6, and reduced the level of occludin1 and claudin5. These changes were followed by increased infarct size (% contralateral hemisphere) and Evans blue extravasation into the brain indicating compromise of the BBB. PROG and ALLO attenuated BBB disruption and infarct size following pMCAO by reducing MMPs and the inflammatory response and by preventing the degradation of occludin1 and claudin5. We conclude that PROG and ALLO can help to protect BBB disruption following pMCAO.

Recent experimental evidence indicates that progesterone (PROG) protects against various models of brain injury, including ischemic stroke. Most human studies of pharmacologic treatments for acute cerebral stroke have failed despite initial success in animal models. To simulate better the typical human stroke without reperfusion, the present study was conducted to examine the efficacy of PROG on infarct volume and functional outcome in a permanent model of stroke, using direct cauterization of the middle cerebral artery (MCA). Twenty-four male adult Sprague-Dawley rats underwent pMCAO by electro-coagulation and sham operation. After induction of permanent MCA occlusion (pMCAO), the rats received an initial intraperitoneal injection of PROG (8 mg/kg) or vehicle at 1h post-occlusion followed by subcutaneous injections at 6, 24 and 48 h. Functional deficits were tested on the rotarod and grip strength meter at 24, 48 and 72 h after pMCAO. The rats were killed 72 h after surgery and isolated brain was sectioned into coronal slices and stained with 2, 3, 5-triphenyltetrazolium chloride (TTC). PROG-treated rats showed a substantial reduction (54.05%) in the volume of the infarct (% contralateral hemisphere) compared to vehicle controls. In addition there was a significant improvement in ability to remain on an accelerating rotarod and increased grip strength observed in the pMCAO rats treated with PROG compared to vehicle. Taken together, these data indicate that PROG is beneficial in one of the best-characterized models of stroke, and may warrant further testing in future clinical trials for human stroke.

No abstract

We tested the hypothesis that the phosphatidylinositol-3 kinase (PI3K/Akt) pathway mediates some of the neuroprotective effects of progesterone (PROG) after ischemic stroke. We examined whether PROG acting through the PI3K/Akt pathway could affect the expression of vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF). Rats underwent permanent focal cerebral ischemia (pMCAO) by electro-coagulation and received intraperitoneal injections of PROG (8mg/kg) or vehicle at 1h post-occlusion and subcutaneous injections at 6, 24, and 48h. PAkt/Akt levels, apoptosis and apoptosis-related proteins (pBAD, BAD, caspase-3, and cleaved caspase-3) were analyzed by TUNEL assays, Western blotting and immunohistochemistry at 24h post-pMCAO. VEGF and BDNF were analyzed at 24, 72h and 14 days post-pMCAO with Western blots. Following pMCAO, PROG treatment significantly (p<0.05) reduced ischemic lesion size and edema. Treatment with PROG significantly (p<0.05) decreased VEGF at 24 and 72h but increased VEGF expression 14d after injury. The treatment also increased BDNF, and attenuated apoptosis by increasing Akt phosphorylation compared to vehicle-alone. The selective PI3K inhibitor Wortmannin compromised PROG-induced neuroprotective effects and reduced the elevation of pAkt levels in the ischemic penumbra. Our findings lead us to suggest that the PI3K/Akt pathway can play a role in mediating the neuroprotective effects of PROG after stroke by altering the expression of trophic factors in the brain.