灯盏花乙素通过调节线粒体融合-分裂平衡抑制氧糖剥夺/复糖复氧诱导的N2a细胞焦亡

目的: 探讨灯盏花乙素对氧糖剥夺/复氧复糖(OGD/R)诱导的N2a细胞焦亡的影响及作用机制。方法: 建立N2a细胞OGD/R损伤模型,设立对照组、模型组、灯盏花乙素给药组和线粒体分裂抑制剂Mdivi-1组。通过CCK-8检测N2a细胞增殖能力,检测各组细胞培养基中LDH和IL-1β水平,通过JC-1探针检测线粒体膜电位,Western blot检测线粒体融合分裂关键蛋白(Drp-1、Mfn-1、Mfn-2、OPA-1)和细胞焦亡标志蛋白(Caspase-1、NLRP-3和GSDMD)表达。结果: 与对照组相比,OGD/R可显著降低N2a细胞活力,灯盏花乙素可明显提高N2a细胞活力。与模型组相比,灯盏花乙素可减少LDH的释放,降低Caspase-1和IL-1β水平,提高线粒体膜势能,降低Drp-1的表达水平,上调Mfn-1、Mfn-2和OPA-1的表达,并减少细胞焦亡关键蛋白NLRP-3和GSDMD的表达。进一步给予Drp-1抑制剂Mdivi-1,与模型组相比,Mdivi-1可明显提高线粒体膜势能,减少Caspase-1、NLRP-3和GSDMD的表达。结论: 灯盏花乙素可通过抑制线粒体过度分裂,改善线粒体融合-分裂失衡与功能异常,进而缓解OGD/R所致N2a细胞焦亡。

Inhibitory effect of scutellarin on oxygen glucose deprivation/reperfusion induced pyroptosis by adjusting mitochondrial fusion-division balance in N2a cells

OBJECTIVE To investigate the effect of scutellarin on oxygen glucose deprivation/reperfusion (OGD/R) induced N2a cell pyroptosis and the underlying mechanism.METHODS An OGD/R model was established on N2a cells, and N2a cells were divided into control group. model group, scutellarin group and mitochondrial division inhibitor Mdivi-1 group. CCK-8 was used to detect the proliferation of N2a cells, and the levels of LDH and IL-1β in the cell culture media were determined as well. The mitochondrial membrane potential (MMP) was detected by JC-1 probe. Western blotting was applied to detect the expression of the key proteins in mitochondrial fusion-division (Drp-1, Mfn-1, Mfn-2, OPA-1), and the cell pyroptosis marker proteins (Caspase-1, NLRP-3 and GSDMD) in N2a cells.RESULTS Compared with the control group, OGD/R declined the viability of N2a cells, while scutellarin enhanced the viability. Compared with the model group, scutellarin decreased the release of LDH, lowered the levels of Caspase-1 and IL-1β, increased the NMP, decreased Drp-1 expression level, up-regulated the expression of Mfn-1, Mfn-2 and OPA-1, and inhibited the expression of the key proteins in pyroptosis (NLRP-3 and GSDMD). Compared with the model group, Drp-1inhibitor Mdivi-1 could increase the NMP, and inhibit the expression of Caspase-1, NLRP-3 and GSDMD.CONCLUSION Scutellarin can alleviate OGD/R induced N2a cells pyroptosis by attenuating mitochondrial disturbance caused by excessive mitochondrial fission.