JLX001 ameliorates cerebral ischemia injury by modulating microglial polarization and compromising NLRP3 inflammasome activation via the NF-κB signaling pathway.

Ischemic stroke is a devastating disease with high morbidity and mortality rates, and the proinflammatory microglia-mediated inflammatory response directly affects stroke outcome. Previous studies have reported that JLX001, a novel compound with a structure similar to that of cyclovirobuxine D (CVB-D), exerts antiapoptotic, anti-inflammatory and antioxidative effects on ischemia-induced brain injury. However, the role of JLX001 in microglial polarization and nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome regulation after ischemic stroke has not been fully investigated. In this study, we used the middle cerebral artery occlusion (MCAO) method to establish a focal cerebral ischemia model and found that JLX001 attenuated the brain infarct size and improved cerebral damage. Moreover, the expression levels of proinflammatory cytokines (interleukin [IL]-1β and tumor necrosis factor [TNF]-α) were significantly reduced while those of the anti-inflammatory cytokine IL-10 were increased in the JLX001-treated group. Immunofluorescence staining and flow cytometry revealed an increased number of anti-inflammatory phenotypic microglia and a reduced number of proinflammatory phenotypic microglia in JLX001-treated MCAO mice. Western blotting analysis showed that JLX001 inhibited the expression of NLRP3 and proteins related to the NLRP3 inflammasome axis in vivo. Furthermore, JLX001 reduced the number of NLRP3/Iba1 cells in ischemic penumbra tissues. Finally, mechanistic analysis revealed that JLX001 significantly inhibited the expression of proteins related to the NF-κB signaling pathway. Additionally, pyrrolidine dithiocarbamate (PDTC), an NF-κB inhibitor, ameliorated cerebral ischemia-reperfusion injury by suppressing microglial polarization towards the proinflammatory phenotype and NLRP3 activation in vivo, further suggesting that these protective effects of JLX001 were mediated by inhibition of the NF-κB signaling pathway. These results suggest that JLX001 is a promising therapeutic approach for ischemic stroke.

Bian HJ ，Xu SY ，Li HQ ，Jia JQ ，Ye L ，Shu S ，Xia SN ，Gu Y ，Zhu X ，Xu Y ，Cao X ... -  《-》

Curcumin Ameliorates White Matter Injury after Ischemic Stroke by Inhibiting Microglia/Macrophage Pyroptosis through NF-κB Suppression and NLRP3 Inflammasome Inhibition.

NLRP3 inflammasome-mediated pyroptosis is a proinflammatory programmed cell death pathway, which plays a vital role in functional outcomes after stroke. We previously described the beneficial effects of curcumin against stroke-induced neuronal damage through modulating microglial polarization. However, the impact of curcumin on microglial pyroptosis remains unknown. Here, stroke was modeled in mice by middle cerebral artery occlusion (MCAO) for 60 minutes and treated with curcumin (150 mg/kg) intraperitoneally immediately after reperfusion, followed by daily administrations for 7 days. Curcumin ameliorated white matter (WM) lesions and brain tissue loss 21 days poststroke and improved sensorimotor function 3, 10, and 21 days after stroke. Furthermore, curcumin significantly reduced the number of gasdermin D+ (GSDMD+) Iba1+ and caspase-1+Iba1+ microglia/macrophage 21 days after stroke. In vitro, lipopolysaccharide (LPS) with ATP treatment was used to induce pyroptosis in primary microglia. Western blot revealed a decrease in pyroptosis-related proteins, e.g., GSDMD-N, cleaved caspase-1, NLRP3, IL-1β, and IL-18, following in vitro or in vivo curcumin treatment. Mechanistically, both in vivo and in vitro studies confirmed that curcumin inhibited the activation of the NF-κB pathway. NLRP3 knocked down by siRNA transfection markedly increased the inhibitory effects of curcumin on microglial pyroptosis and proinflammatory responses, both in vitro and in vivo. Furthermore, stereotaxic microinjection of AAV-based NLRP3 shRNA significantly improved sensorimotor function and reduced WM lesion following curcumin treatment in MCAO mice. Our study suggested that curcumin reduced stroke-induced WM damage, improved functional outcomes, and attenuated microglial pyroptosis, at least partially, through suppression of the NF-κB/NLRP3 signaling pathway, further supporting curcumin as a potential therapeutic drug for stroke.

Ran Y ，Su W ，Gao F ，Ding Z ，Yang S ，Ye L ，Chen X ，Tian G ，Xi J ，Liu Z ... -  《-》

Aloe-emodin alleviates cerebral ischemia-reperfusion injury by regulating microglial polarization and pyroptosis through inhibition of NLRP3 inflammasome activation.

Li X ，Yao M ，Li L ，Ma H ，Sun Y ，Lu X ，Jing W ，Nie S ... -  《-》

Baicalein ameliorates ischemic brain damage through suppressing proinflammatory microglia polarization via inhibiting the TLR4/NF-κB and STAT1 pathway.

Microglial polarization mediated neuroinflammation plays an important role in the pathological process of stroke. The aim of this study is to determine whether baicalein indirectly ameliorates neuronal injury through modulating microglial polarization after stroke and if so, then by what mechanism. The effects of baicalein on microglial polarization were revealed through the middle cerebral artery occlusion mouse model (MCAO, n = 6), the lipopolysaccharide (LPS) + interferon-γ (IFN-γ) and oxygen-glucose deprivation (OGD) induced neuroinflammatory microglia model (BV2, n = 3), respectively. Mice were treated with baicalein (100 mg/kg, i.g.) after reperfusion, and followed by daily administrations for 3 days. Results showed that the infarct volumes at 3 d in vehicle and baicalein-treated MCAO mice were 91.18 ± 4.02% and 55.36 ± 4.10%. Baicalein improved sensorimotor functions (p < 0.01) after MCAO. Real-time PCR revealed that baicalein decreased proinflammatory markers expression (p < 0.05), while elevated the anti-inflammatory markers (p < 0.05) in vivo and in vitro. Both western blot and immunofluorescent staining further confirmed that baicalein reduced proinflammatory marker CD16 levels (p < 0.01) and enhanced anti-inflammatory marker CD206 or Arg-1 levels (p < 0.05). Notably, baicalein suppressed the release of proinflammatory cytokines (p < 0.05) and nitric oxide (NO, p < 0.001). Mechanistically, baicalein prevented increases in TLR4 protein levels (p < 0.001), the phosphorylation of IKBα and p65 (p < 0.01), and the nuclear translocation of NF-κB p65 (p < 0.05). The NF-κB inhibitor, BAY 11-7085, enhanced the inhibitory effect of baicalein on the proinflammatory microglial polarization. Baicalein also inhibited the phosphorylation of signal transducer and activator of transcription 1 (STAT1, p < 0.001). A microglia-neuron co-culture system revealed that baicalein driven neuroprotection against OGD induced neuronal damage through modulating microglial polarization (p < 0.05). Baicalein indirectly ameliorates neuronal injury after stroke by polarizing microglia toward the anti-inflammatory phenotype via inhibition of the TLR4/NF-κB pathway and down-regulation of phosphorylated STAT1, suggesting that baicalein might serve a potential therapy for stroke.

Ran Y ，Qie S ，Gao F ，Ding Z ，Yang S ，Tian G ，Liu Z ，Xi J ... -  《-》

Phthalide derivative CD21 ameliorates ischemic brain injury in a mouse model of global cerebral ischemia: involvement of inhibition of NLRP3.

The activation of NLRP3 inflammasome is closely related to ischemic brain injury and inhibition of NLRP3 inflammasome activation may be a new therapeutic strategy for ischemic stroke. Our previous studies showed that ligustilide (LIG) had a dose-dependent neuroprotective effect on various models of cerebral ischemia and dementia in vivo and in vitro. CD21, a kind of phthalide derivative, was modified from LIG. In this study, we established a global cerebral ischemia-reperfusion model in mice by bilateral common carotid artery ligation (2VO), and explored the neuroprotective effect of CD21 and its anti-inflammatory mechanism on cerebral ischemia mice. CD21 significantly improved weight loss, neurobehavioral deficits and neurons loss in hippocampal CA1 and caudate putamen (CPu) subregions, which were induced by 2VO in mice. CD21 significantly inhibited the overactivation of astrocyte and microglia, and decreased the mRNA level of IL-6, TNF-α and IL-1β. Moreover, CD21 significantly inhibited the activation of TLR4/NF-κB signaling pathway mediated by HMGB1 and NLRP3/ASC/Caspase-1 signaling pathway mediated by Cathepsin B, thus inhibiting the activation of NLRP3 inflammasome. Our results demonstrated that CD21 may exert a neuroprotection by inhibiting NLRP3 inflammasome activation after cerebral ischemia. These findings provide a new strategy for the treatment of ischemic stroke.

Li X ，Shi MQ ，Chen C ，Du JR ... -  《-》