Farnesoid X receptor knockout protects brain against ischemic injury through reducing neuronal apoptosis in mice
Farnesoid X receptor knockout protects brain against ischemic injury through reducing neuronal apoptosis in mice
复制标题
Farnesoid X受体敲除通过减少小鼠神经元凋亡保护大脑免受缺血性损伤
DOI：
10.1186/s12974-020-01838-w
复制
发表时间：
2020-05-25
影响因子：
9.3
通讯作者：
Wang, Yongting
中科院分区：
文献类型：
--
作者：
Shan, Hui-Min;Zang, Minhua;Wang, Yongting
背景法尼醇X受体（FXR）是一种核受体，在控制多种疾病中的细胞凋亡中发挥着关键作用。先前的研究表明，敲除FXR可以通过减少心肌缺血小鼠的心肌细胞凋亡来改善心脏功能。然而，FXR在脑缺血后的作用仍不清楚。本研究探讨FXR基因敲除(KO)对小鼠脑缺血再灌注后功能恢复的影响及机制。方法对成年雄性C57BL/6野生型和FXRKO小鼠进行90分钟短暂大脑中动脉闭塞(tMCAO)。将小鼠分为五组：假手术组、野生型tMCAO、FXRKO tMCAO、用钙激动剂Bayk8644处理的野生型tMCAO以及用Bayk8644处理的FXRKO tMCAO。使用免疫组织化学和蛋白质印迹检查 FXR 表达。分别于中风后第3天和第14天检查脑梗塞和脑萎缩体积。中风后 14 天进行神经行为测试。使用Western blot和RT-PCR检测凋亡因子（Bcl-2、Bax和Cleaved caspase-3）的蛋白水平和促炎因子（TNF-α、IL-6、IL-1β、IL-17和IL-18）的mRNA水平。使用共焦和双光子显微镜获得 TUNEL 染色和钙成像。结果 缺血性中风后 FXR 的表达上调，其位于神经元的细胞核中。发现与媒介物相比，FXRKO 可以减少 tMCAO 后的梗塞体积并促进神经行为恢复。与对照组相比，FXRKO 小鼠中凋亡因子和促炎因子的表达减少。与媒介物相比，FXRKO 小鼠梗塞周围区域的 NeuN+/TUNEL+ 细胞数量下降。我们进一步证明，抑制 FXR 可减少钙超载，并且添加离子霉素可以在体外逆转这种神经保护作用。更重要的是，体内结果表明，细胞内钙浓度的增加会加重缺血性损伤，并逆转FXRKO对小鼠的神经保护作用。结论FXRKO可以通过减少钙内流，促进神经行为恢复，减轻缺血性脑损伤、炎症释放和神经元凋亡，表明其作为中风治疗靶点的作用。
BackgroundFarnesoid X receptor (FXR) is a nuclear receptor that plays a critical role in controlling cell apoptosis in diverse diseases. Previous studies have shown that knocking outFXRimproved cardiac function by reducing cardiomyocyte apoptosis in myocardial ischemic mice. However, the role of FXR after cerebral ischemia remains unknown. In this study, we explored the effects and mechanisms ofFXRknockout (KO) on the functional recovery of mice post cerebral ischemia-reperfusion.MethodsAdult male C57BL/6 wild type andFXRKO mice were subjected to 90-min transient middle cerebral artery occlusion (tMCAO). The mice were divided into five groups: sham, wild-type tMCAO,FXRKO tMCAO, wild-type tMCAO treated with calcium agonist Bayk8644, andFXRKO tMCAO treated with Bayk8644. FXR expression was examined using immunohistochemistry and Western blot. Brain infarct and brain atrophy volume were examined at 3 and 14 days after stroke respectively. Neurobehavioral tests were conducted up to 14 days after stroke. The protein levels of apoptotic factors (Bcl-2, Bax, and Cleaved caspase-3) and mRNA levels of pro-inflammatory factors (TNF-α, IL-6, IL-1β, IL-17, and IL-18) were examined using Western blot and RT-PCR. TUNEL staining and calcium imaging were obtained using confocal and two-photon microscopy.ResultsThe expression of FXR was upregulated after ischemic stroke, which is located in the nucleus of the neurons.FXRKO was found to reduce infarct volume and promote neurobehavioral recovery following tMCAO compared to the vehicle. The expression of apoptotic and pro-inflammatory factors decreased inFXRKO mice compared to the control. The number of NeuN+/TUNEL+cells declined in the peri-infarct area ofFXRKO mice compared to the vehicle. We further demonstrated that inhibition of FXR reduced calcium overload and addition of ionomycin could reverse this neuroprotective effect in vitro. What is more, in vivo results showed that enhancement of intracellular calcium concentrations could aggravate ischemic injury and reverse the neuroprotective effect ofFXRKO in mice.ConclusionsFXRKO can promote neurobehavioral recovery and attenuate ischemic brain injury, inflammatory release, and neuronal apoptosis via reducing calcium influx, suggesting its role as a therapeutic target for stroke treatments.