Abstract
Blockading P2X7 receptor(P2X7R) provides neuroprotection toward various neurological disorders, including stroke, traumatic brain injury, and subarachnoid hemorrhage. However, whether and how P2X7 receptor suppression protects blood-brain barrier(BBB) after intracerebral hemorrhage(ICH) remains unexplored. In present study, intrastriatal autologous-blood injection was used to mimic ICH in rats. Selective P2X7R inhibitor A438079, P2X7R agonist BzATP, and P2X7R siRNA were administrated to evaluate the effects of P2X7R suppression. Selective RhoA inhibitor C3 transferase was administered to clarify the involvement of RhoA. Post-assessments, including neurological deficits, Fluoro-Jade C staining, brain edema, Evans blue extravasation and fluorescence, western blot, RhoA activity assay and immunohistochemistry were performed. Then the key results were verified in collagenase induced ICH model. We found that endogenous P2X7R increased at 3 hrs after ICH with peak at 24 hrs, then returned to normal at 72 hrs after ICH. Enhanced immunoreactivity was observed on the neurovascular structure around hematoma at 24 hrs after ICH, along with perivascular astrocytes and endothelial cells. Both A438079 and P2X7R siRNA alleviated neurological deficits, brain edema, and BBB disruption after ICH, in association with RhoA activation and down-regulated endothelial junction proteins. However, BzATP abolished those effects. In addition, C3 transferase reduced brain injury and increased endothelial junction proteins’ expression after ICH. These data indicated P2X7R suppression could preserve BBB integrity after ICH through inhibiting RhoA activation.
Highlights
Spontaneous intracerebral hemorrhage (ICH) is a common fatal stroke subtype due to small vessel bleed within brain parenchyma and subsequent hematoma[1]
Double immunofluorescence staining revealed that the P2X7 receptor immunoreactivity was mainly found peri-hematoma astrocytes and endothelial cells in ipsilateral striatum, which were marked with glial firillary acidic protein (GFAP) and von Willebrand factor (vWF), respectively (Fig. 1C)
We found that endogenous P2X7 receptor increased at 3 hours after ICH with a peak at 24 hours, returned to normal level at 72 hours after ICH
Summary
Spontaneous intracerebral hemorrhage (ICH) is a common fatal stroke subtype due to small vessel bleed within brain parenchyma and subsequent hematoma[1]. Decreased expression and disarrangement of the endothelial junction proteins after ICH indicate blood brain barrier integrity disruption and increased paracellular permeability[6,7]. Our previous studies demonstrated that P2X7 receptor involved in the neuroinflammation and apoptosis after subarachnoid hemorrhage[14,15]. It is unclear whether endogenous P2X7 receptor participates in the pathophysiology process, especially in BBB disruption, of the early brain injury after ICH. Recent studies indicate that Ras homolog gene family member A (RhoA) might be the downstream signal of P2X7 receptor involved in blood brain barrier integrity modulation[16,17]. In the present study, we sought to investigate the potential role and mechanism of P2X7 receptor in the blood brain barrier integrity after ICH
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