Neuroinflammation and oxidative stress plays an important role in the pathogenesis of early brain injury (EBI) after subarachnoid hemorrhage (SAH). This study is the first to show that activation of autophagy protein nuclear receptor binding factor 2 (NRBF2) could reduce endoplasmic reticulum stress (ERS)-associated inflammation and oxidative stress after SAH. Male C57BL/6J mice were subjected to endovascular perforation to establish a model of SAH. NRBF2 overexpression adeno-associated virus (AAV), NRBF2 small interfering RNAs (siRNA), lysosomal inhibitor-chloroquine (CQ), and late endosome GTPase Rab7 receptor antagonist-CID1067700 (CID) were used to investigate the role of NRBF2 in EBI after SAH. Neurological tests, brain water content, western blotting and immunofluorescence staining were evaluated. Our study found that the level of NRBF2 was increased after SAH and peaked at 24 h after SAH. In addition, we found that the overexpression of NRBF2 significantly improved neurobehavioral scores and reduced ERS, oxidative stress, and neuroinflammation in SAH, whereas the inhibition of NRBF2 exacerbated these phenotypes. In terms of mechanism, NRBF2 overexpression significantly promoted autophagosome maturation, with the downregulation of CHOP, Romo-1, TXNIP, NLRP3, TNF-α, and IL-1β expression through interaction with Rab7. The protective effect of NRBF2 on ERS-associated neuroinflammation and oxidative stress after SAH was eliminated by treatment with CQ. Meanwhile, it was also reversed by intraperitoneal injection of CID. Moreover, the MIT domain of NRBF2 was identified as a critical binding site that interacts with Rab7 and thereby promotes autophagosome maturation. Our data provide evidence that the autophagy protein NRBF2 has a protective effect on endoplasmic reticulum stress-associated neuroinflammation and oxidative stress by promoting autophagosome maturation through interactions with Rab7 after SAH.

中文翻译：

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SAH后自噬蛋白NRBF2通过与Rab7相互作用促进自噬体成熟，减轻内质网应激相关的神经炎症和氧化应激

神经炎症和氧化应激在蛛网膜下腔出血（SAH）后早期脑损伤（EBI）的发病机制中起重要作用。这项研究首次表明激活自噬蛋白核受体结合因子 2 (NRBF2) 可以减少 SAH 后内质网应激 (ERS) 相关的炎症和氧化应激。雄性C57BL/6J小鼠经血管腔内穿孔建立SAH模型。NRBF2 过表达腺相关病毒 (AAV)、NRBF2 小干扰 RNA (siRNA)、溶酶体抑制剂-氯喹 (CQ) 和晚期内体 GTPase Rab7 受体拮抗剂-CID1067700 (CID) 用于研究 NRBF2 在 SAH 后 EBI 中的作用. 评估了神经学测试、脑含水量、蛋白质印迹和免疫荧光染色。我们的研究发现，SAH 后 NRBF2 水平升高，并在 SAH 后 24 小时达到峰值。此外，我们发现 NRBF2 的过表达显着改善了 SAH 的神经行为评分并降低了 ERS、氧化应激和神经炎症，而 NRBF2 的抑制加剧了这些表型。就机制而言，NRBF2过表达显着促进自噬体成熟，通过与Rab7的相互作用下调CHOP、Romo-1、TXNIP、NLRP3、TNF-α和IL-1β的表达。NRBF2对SAH后ERS相关的神经炎症和氧化应激的保护作用被CQ治疗消除。同时，腹腔注射CID也可逆转。而且，NRBF2 的 MIT 结构域被确定为与 Rab7 相互作用的关键结合位点，从而促进自噬体成熟。我们的数据提供的证据表明，自噬蛋白 NRBF2 在 SAH 后通过与 Rab7 的相互作用促进自噬体成熟，从而对内质网应激相关的神经炎症和氧化应激具有保护作用。