Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, 19140, USA.
J Neuroinflammation. 2024 Sep 28;21(1):244. doi: 10.1186/s12974-024-03230-4.
Alcohol consumption leads to neuroinflammation and blood‒brain barrier (BBB) damage, resulting in neurological impairment. We previously demonstrated that ethanol-induced disruption of barrier function in human brain endothelial cells was associated with mitochondrial injury, increased ATP and extracellular vesicle (EV) release, and purinergic receptor P2 × 7R activation. Therefore, we aimed to evaluate the effect of P2 × 7R blockade on peripheral and neuro-inflammation in ethanol-exposed mice. In a chronic intermittent ethanol (CIE)-exposed mouse model, P2 × 7R was inhibited by two different methods: Brilliant Blue G (BBG) or gene knockout. We assessed blood ethanol concentration (BEC), brain microvessel gene expression by using RT2 PCR array, plasma P2 × 7R and P-gp, serum ATP, EV-ATP, number of EVs, and EV mtDNA copy numbers. An RT2 PCR array of brain microvessels revealed significant upregulation of proinflammatory genes involved in apoptosis, vasodilation, and platelet activation in CIE-exposed wild-type animals, which were decreased 15-50-fold in BBG-treated-CIE-exposed animals. Plasma P-gp levels and serum P2 × 7R shedding were significantly increased in CIE-exposed animals. Pharmacological or genetic suppression of P2 × 7R decreased receptor shedding to levels equivalent to those in control group. The increase in EV number and EV-ATP content in the CIE-exposed mice was significantly reduced by P2 × 7R inhibition. CIE mice showed augmented EV-mtDNA copy numbers which were reduced in EVs after P2 × 7R inhibition or receptor knockout. These observations suggested that P2 × 7R signaling plays a critical role in ethanol-induced brain injury. Increased extracellular ATP, EV-ATP, EV numbers, and EV-mtDNA copy numbers highlight a new mechanism of brain injury during alcohol exposure via P2 × 7R and biomarkers of such damage. In this study, for the first time, we report the in vivo involvement of P2 × 7R signaling in CIE-induced brain injury.
酒精摄入可导致神经炎症和血脑屏障（BBB）损伤，从而导致神经功能障碍。我们之前的研究表明，乙醇诱导的人脑内皮细胞屏障功能障碍与线粒体损伤、ATP 和细胞外囊泡（EV）释放增加以及嘌呤能受体 P2×7R 激活有关。因此，我们旨在评估 P2×7R 阻断对乙醇暴露小鼠的外周和神经炎症的影响。在慢性间歇性乙醇（CIE）暴露小鼠模型中，通过两种不同的方法抑制 P2×7R：亮蓝 G（BBG）或基因敲除。我们通过 RT2 PCR 阵列评估了血液乙醇浓度（BEC）、脑微血管基因表达、血浆 P2×7R 和 P-糖蛋白、血清 ATP、EV-ATP、EV 数量和 EV mtDNA 拷贝数。大脑微血管 RT2 PCR 阵列显示，CIE 暴露的野生型动物中涉及凋亡、血管舒张和血小板激活的促炎基因显著上调，BBG 处理的 CIE 暴露动物中的这些基因上调倍数降低了 15-50 倍。CIE 暴露动物的血浆 P-糖蛋白水平和血清 P2×7R 脱落显著增加。P2×7R 的药理学或基因抑制将受体脱落降低至与对照组相当的水平。CIE 抑制后，EV 数量和 EV-ATP 含量的增加显著减少。CIE 小鼠的 EV-mtDNA 拷贝数增加，P2×7R 抑制或受体敲除后 EV 中的 EV-mtDNA 拷贝数减少。这些观察结果表明，P2×7R 信号在乙醇诱导的脑损伤中起关键作用。细胞外 ATP、EV-ATP、EV 数量和 EV-mtDNA 拷贝数的增加突出了通过 P2×7R 以及此类损伤的生物标志物在酒精暴露期间发生脑损伤的新机制。在这项研究中，我们首次报道了 P2×7R 信号在 CIE 诱导的脑损伤中的体内参与。
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