Dl-3-n-butylphthalide improves stroke outcomes after focal ischemic stroke in mouse model by inhibiting the pyroptosis-regulated cell death and ameliorating neuroinflammation.

Recent studies have highlighted the involvement of pyroptosis-mediated cell death and neuroinflammation in ischemic stroke (IS) pathogenesis. DL-3-n-butylphthalide (NBP), a synthesized compound based on an extract from seeds of Apium graveolens, possesses a broad range of biological effects. However, the efficacy and the underlying mechanisms of NBP in IS remain contentious. Herein, we investigated the therapeutic effects of NBP and elucidated its potential mechanisms in neuronal cell pyroptosis and microglia inflammatory responses. Adult male mice underwent permanent distal middle cerebral artery occlusion (dMCAO), followed by daily oral gavage of NBP (80 mg/kg) for 1, 7, or 21 consecutive days. Gene Expression Omnibus (GEO) dataset of IS patients peripheral blood RNA sequencing was analyzed to identify differentially expressed pyroptosis-related genes (PRGs) during the ischemic process. Our results suggested that NBP treatment effectively alleviated brain ischemic damage, resulting in decreased neurological deficit scores, reduced infarct volume, and improved neurological and behavioral functions. RNA sequence data from human unveiled upregulated PRGs in IS. Subsequently, we observed that NBP downregulated pyroptosis-associated markers at days 7 and 21 post-modeling, at both the protein and mRNA levels. Additionally, NBP suppressed the co-localization of pyroptosis markers with neuronal cells to variable degrees and simultaneously mitigated the accumulation of activated microglia. Overall, our data provide novel evidence that NBP treatment significantly attenuates ischemic brain damage and promotes recovery of neurological function in the early and recovery phases after IS, probably by negatively regulating the pyroptosis cell death of neuronal cells and inhibiting toxic neuroinflammation in the central nervous system.

Neuroprotective effects of cilostazol are mediated by multiple mechanisms in a mouse model of permanent focal ischemia.

The phosphodiesterase (PDE) 3 inhibitor cilostazol, used as an anti-platelet drug, reportedly can also ameliorate ischemic brain injury. Here, we investigated the effects of cilostazol in a permanent focal ischemia mice model. Male Balb/c mice were subjected to permanent middle cerebral artery occlusion. Mice were then treated with either cilostazol (10 or 20mg/kg) or vehicle administered at 30min and 24h post-ischemia, and infarct volumes were assessed at 48h post-ischemia. Mice treated with 20mg/kg of cilostazol or vehicle were sacrificed at 6h or 24h post-ischemia and immunohistochemistry was used for brain sections. Treatment with 20mg/kg of cilostazol significantly reduced infarct volumes to 70.1% of those with vehicle treatment. Immunohistochemistry results for 8-hydroxydeoxyguanosine (OHdG) expression showed that some neurons underwent oxidative stress around the ischemic boundary zone at 6h post-ischemia. Cilostazol treatment significantly reduced the percentage of 8-OHdG-positive neurons (65.8±33.5% with vehicle and 21.3±9.9% with cilostazol). Moreover, NADPH oxidase (NOX) 2-positive neurons were significantly reduced with cilostazol treatment. In contrast, immunohistochemistry results for phosphorylated cyclic-AMP response element binding protein (pCREB) showed that there were significantly more pCREB-positive neurons around the ischemic boundary zone of cilostazol-treated mice than in those of vehicle-treated mice at 24h post-ischemia. These results suggested that cilostazol might have multiple mechanisms of action to ameliorate ischemic tissue damage, by attenuating oxidative stress mediated by suppressing NOX2 expression by ischemic neurons and an anti-apoptotic effect mediated through the pCREB pathway.

Targeting CD36-mediated inflammation reduces acute brain injury in transient, but not permanent, ischemic stroke.

AIMS: The pathology of stroke consists of multiple pro-death processes, and CD36 has been suggested as a multimodal target to reduce oxidative stress and inflammation in ischemic stroke. Using CD36-deficient mice and SS-31, a cell permeable tetrapeptide known to down-regulate CD36 pathways, the current study investigated whether targeting CD36 is effective in transient and permanent ischemic stroke. METHODS: Wild-type or CD36-deficient mice were subjected to either 30-min transient or permanent focal ischemic stroke. In parallel, a cohort of mice subjected to either transient or permanent stroke received either vehicle or 5 mg/kg of SS-31. Monocyte chemoattractant protein-1 (MCP-1) and its receptor CCR2, mRNA levels, and infarct volume and percent hemispheric swelling were measured in the postischemic brain. RESULTS: CD36 deficiency or SS-31 treatment significantly attenuated MCP-1 or CCR2 mRNA up-regulation and injury size in the transient ischemic stroke. However, the approaches failed to show the protective effect in permanent ischemic stroke. CONCLUSION: The study revealed that targeting CD36 has a beneficial effect on transient but not permanent focal ischemic stroke. The study thus precludes a generalized strategy targeting CD36 in ischemic stroke and suggests careful consideration of types of stroke and associated pathology in developing stroke therapies.

Achyrocline satureioides (Lam.) DC. (marcela) reduces brain damage in permanent focal ischemia in rats / Achyrocline satureioides (Lam.) DC. (marcela) reduce el daño cerebral en la isquemia focal permanente en ratas

Introduction: Achyrocline satureioides is a plant which has been widely used in popular medicine and experimental studies confirm its antioxidant and anti-inflammatory effects, attributable to the presence of flavonoids, mainly quercetin. Objectives: to evaluate the neuroprotective effects of a chronic oral pre-administration to rats with an Achyrocline satureioides decoction (2 %). Methods: for decoction, dried flowers of Achyrocline satureioides were used. The consumption of food and AS decoction/water of the rats was evaluated daily and weight gain weekly; quercetin content in the decoction and in the plasma of the rats was evaluated by high performance liquid chromatography. The cerebral damage was assessed with a tetrazolium salt (TTC) and a behavioral test was performed previously. Nissl staining and Fluoro-Jade histochemistry were used. Results: the pre-treatment with Achyrocline satureioides in all groups reverted the functional deficit and, during 21 days, the infarction volume also decreased significantly. Nissl staining showed a higher percentage of preserved neuronal population and the Fluoro-Jade showed a decreased of the neurons in degeneration. Conclusions: the quercetin levels in the decoction and plasma of rats could explain the preventive benefits of Achyrocline satureioides due to the antioxidant and anti-inflammatory properties described for this flavonoid.

Introducción: Achyrocline satureioides es una planta que ha sido ampliamente utilizada en la medicina popular y los estudios experimentales confirman sus efectos antioxidantes y antiinflamatorios, atribuibles a la presencia de flavonoides, principalmente quercetina. Objetivos: evaluar los efectos neuroprotectores de la pre-administración oral crónica a ratas con una decocción de Achyrocline satureioides 2 %. Métodos: para la decocción se utilizaron flores secas de Achyrocline satureioides. Se cuantificaron, diariamente, el consumo de alimentos, la decocción y el agua; y cada semana, la ganancia de peso. El contenido de quercetina en la decocción y en el plasma de las ratas se evaluó utilizando la técnica de cromatografía líquida de alta resolución. El daño cerebral se cuantificó con una sal de tetrazolio y antes se realizó una prueba de comportamiento. Se utilizaron la tinción de Nissl y el fluoro Jade. Resultados: el pretratamiento con Achyrocline satureioides en todos los grupos revirtió el déficit funcional, y la decocción durante 21 días también decreció de modo significativo el volumen del infarto. La tinción de Nissl mostró alto porcentaje de población neuronal conservada y el fluoro Jade presentó un decrecimiento en las neuronas en degeneración. Conclusiones: los niveles de quercetina en la decocción y el plasma de las ratas podrían explicar los beneficios preventivos de Achyrocline satureioides, debido a las propiedades antioxidantes y antiinflamatorias descritas para este flavonoide.