Atorvastatin ameliorates early brain injury after subarachnoid hemorrhage via inhibition of AQP4 expression in rabbits.

The therapeutic effects of atorvastatin on early brain injury (EBI), cerebral edema and its association with aquaporin 4 (AQP4) were studied in rabbits after subarachnoid hemorrhage (SAH) using western blot analysis and the dry-wet method. Seventy-two healthy male New Zealand rabbits weighing between 2.5 and 3.2 kg were randomly divided into three groups: the SAH group (n=24), sham-operated group (n=24) and the SAH + atorvastatin group (n=24). A double SAH model was employed. The sham-operated group were injected with the same dose of saline solution, the SAH + atorvastatin group received atorvastatin 20 mg/kg/day after SAH. All rabbit brain samples were taken at 72 h after the SAH model was established successfully. Brain edema was detected using the dry-wet method after experimental SAH was induced; AQP4 and caspase-3 expression was measured by western blot analysis, and neuronal apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) staining at 72 h after SAH. The results indicated that brain edema and injury appeared soon after SAH, while brain edema and EBI were ameliorated and increased behavior scores were noted after prophylactic use of atorvastatin. Compared with the SAH group, the level of AQP4 and the cerebral content of water was significantly decreased (P<0.01) by atorvastatin, and TUNEL staining and studying the expression of caspase-3 showed that the apoptosis of neurons was reduced markedly both in the hippocampus and brain cortex by atorvastatin. The results suggest that atorvastatin ameliorated brain edema and EBI after SAH, which was related to its inhibition of AQP4 expression. Our findings provide evidence that atorvastatin is an effective and well-tolerated approach for treating SAH in various clinical settings.

Curcumin attenuates brain edema in mice with intracerebral hemorrhage through inhibition of AQP4 and AQP9 expression.

AIM: Aquaporins (AQPs) are the water-channels that play important roles in brain water homeostasis and in cerebral edema induced by brain injury. In this study we investigated the relationship between AQPs and a neuroprotective agent curcumin that was effective in the treatment of brain edema in mice with intracerebral hemorrhage (ICH). METHODS: ICH was induced in mice by autologous blood infusion. The mice immediately received curcumin (75, 150, 300 mg/kg, ip). The Rotarod test scores, brain water content and brain expression of AQPs were measured post ICH. Cultured primary mouse astrocytes were used for in vitro experiments. The expression of AQP1, AQP4 and AQP9 and NF-κB p65 were detected using Western blotting or immunochemistry staining. RESULTS: Curcumin administration dose-dependently reduced the cerebral edema at d 3 post ICH, and significantly attenuated the neurological deficits at d 5 post ICH. Furthermore, curcumin dose-dependently decreased the gene and protein expression of AQP4 and AQP9, but not AQP1 post ICH. Treatment of the cultured astrocytes with Fe(2+) (10-100 µmol/L) dose-dependently increased the expression and nuclear translocation of NF-κB p65 and the expression of AQP4 and AQP9, which were partly blocked by co-treatment with curcumin (20 µmol/L) or the NF-κB inhibitor PDTC (10 µmol/L). CONCLUSION: Curcumin effectively attenuates brain edema in mice with ICH through inhibition of the NF-κB pathway and subsequently the expression of AQP4 and AQP9. Curcumin may serve as a potential therapeutic agent for ICH.

Effect of Shenfu on inflammatory cytokine release and brain edema after prolonged cardiac arrest in the swine.

OBJECTIVE: Shenfu injection (SFI), a traditional Chinese formulation, has been confirmed to be protective against brain during ischemia and reperfusion injury. In this exploratory study, we investigated the action of SFI in regulating the inflammatory response and brain edema after cardiopulmonary resuscitation. METHODS: After 8 minutes of untreated ventricular fibrillation (VF), pigs in the cardiopulmonary resuscitation group (n = 24) received a central venous injection of either SFI (SFI group; 1.0 mL/kg), epinephrine (EP group; 0.02 mg/kg), or saline (SA group). Levels of porcine-specific tumor necrosis factor α and interleukin in sera were measured using enzyme-linked immunosorbent assay at 0.5, 1, 2, 4, 6, and 24 hours after return of spontaneous circulation (ROSC). Surviving pigs were killed 24 hours after ROSC, and the brains were removed for electron microscopy, Western blotting, and quantitative real-time polymerase chain reaction analysis. RESULTS: Compared with the EP and SA groups, SFI decreased the levels of tumor necrosis factor α and interleukin-6 in serum and the brain (P < .05) and decreased the expression of nuclear factor κB and aquaporin-4 messenger RNA in the brain (P < .05). Shenfu injection also inhibited the expression of nuclear factor κB, matrix metalloproteinase 9, and aquaporin-4 protein after ROSC (P < .05). Observation of brain tissue ultrastructure showed that injury was alleviated in the SFI group compared with the SA and EP groups. CONCLUSIONS: Our exploratory experiments demonstrated that SFI reduced cerebral damage in a porcine model of VF, which may be related to suppression of the inflammatory reaction and decreased brain edema after ROSC.

Bolus injection of hypertonic solutions for cerebral edema in rats: challenge of homeostasis of healthy brain.

Hypertonic solutions are mainstay of osmotherapy to cerebral edema. How hypertonic solutions affect healthy brain homeostasis, however, is not fully understood. Using rat model of cerebral edema induced by local cryoinjury, we found with immunohistochemistry that less microglial activation in healthy hemishere 24 h after hypertonic saline (HS, 3% NaCl) administration, compared to mannitol (20%, the same osmotic concentration of 3% NaCl) while dehydrating the brain tissue. To see whether blood-brain barrier (BBB) or aquaporin-4 (AQP4) contribute to this difference, HS or mannitol was intra-arterially injected to normal rats, and BBB opening, ultrastructure and AQP4 immunoreactivity were examined. Evans blue extravasation indicated that BBB was opened much lighter in HS group than mannitol group at the same time points. Electron microscopy also showed edema around the capillaries slightly lighter in HS than mannitol group 24 h after injection. Meanwhile, HS injection led to AQP4 down regulation in expression similarly as mannitol, compared with NS group. These data suggested that bolus injection of hypertonic agents may lead to microglia activation in healthy brain in different extent, due to BBB compromise, instead of water movement or AQP4 expression. Hence in clinical application, BBB of healthy brain should be considered in perspective to maintain the brain homeostasis.

[Regulative effect of electroacupuncture on aquaporin-4 in rats with focal cerebral ischemia/reperfusion].

OBJECTIVE: To study the effect of electroacupuncture (EA) on changes of aquaporin-4 (AQP4) expression in rats with cerebral ischemia/reperfusion(CI/R). METHODS: One hundred and seventy male Sprauge-Dawley rats were randomized into control (n = 50), CI/R model (n = 60) and EA (n = 60) groups, and the later 2 groups were further divided into 6 h, 12 h, 24 h, 48 h and 72 h subgroups, with 12 cases in each. CI/R model was established by occlusion of the middle cerebral artery (MCAO). EA (3.85/6.25 Hz, 0.8 - 1.0 mA) was applied to "Shuigou" (GV 26) and "Baihui" (GV 20) for 30 min. Transcardial perfusion was conducted with normal saline + paraformaldehyde in rats of different groups for taking brain tissue samples. The degree of brain edema was detected by cresyl violet stain method, permeability of the blood-brain barrier (BBB) reflected with IgG value in brain tissue was displayed by immunohistochemistry, and AQP4 protein and mRNA expression of corpora striata (CS) were detected by using immunohistochemistry (ABC) and RT-PCR techniques respectively. RESULTS: Twelve hours after CI/R, the hemisphere of brain began to get swelling significantly and the exosmose state of IgG became evidently; compared with the corresponding phases of model group, the swell rates in 24 h, 48 h and 72 h subgroups of EA group were significantly lower (P < 0.05, 0.01), and nearly no IgG immunoreaction positive products were found in different phases of EA group. The expression of AQP4 protein and mRNA increased significantly from 12 h and 24 h on separately in model group, while the expression in EA group was markedly weaker. The ratios of AQP4 mRNA and beta-actin mRNA of CS on the affected side of the brain from 24 h to 72 h of EA group were significantly lower than those of model group (P < 0.05, 0.01). CONCLUSION: EA can reduce the degree of brain edema induced by CI/R and improve the degree of BBB injury, which may be related to its effect in down-regulating AQP4 expression.