目的 基于神经特异性跨膜蛋白240（TMEM240）/Rho GDP解离抑制因子β（ARHGDIB）通路探讨穗花杉双黄酮改善脑缺血再灌注损伤小鼠神经炎症的作用机制。方法 采用线栓法建立脑缺血再灌注损伤模型，将造模成功小鼠随机分为模型组、穗花杉双黄酮组、穗花杉双黄酮＋sh-NC组、穗花杉双黄酮＋sh-TMEM240组，另设假手术组，每组24只。检测Zea-longa评分、平衡木测试评分、脑梗死体积比、脑组织含水量变化；HE染色检测缺血侧脑组织病理；免疫荧光染色检测缺血侧脑组织中离子钙接头蛋白1（Iba-1）⁺诱导型一氧化氮合酶（iNOS）⁺、Iba-1⁺ 精氨酸酶1（Arg-1）⁺阳性细胞数；ELISA检测缺血侧脑组织中肿瘤坏死因子-α（TNF-α）、白细胞介素（IL）-1β、IL-6、IL-10、转化生长因子-β1（TGF-β1）含量；Western blotting检测缺血侧脑组织中TMEM240、ARHGDIB、p-NF-κB p65/NF-κB p65蛋白表达。结果 与模型组比较，穗花杉双黄酮组缺血侧脑组织病理损伤明显改善，表现为神经元形态较完整、水肿减轻及炎性浸润减少，Zea-longa评分、脑梗死体积比、脑组织含水量、缺血侧脑组织Iba-1⁺ iNOS⁺阳性细胞数，TNF-α、IL-1β、IL-6水平，p-NF-κB p65/NF-κB p65蛋白表达显著降低，平衡木测试评分、缺血侧脑组织Iba-1⁺ Arg-1⁺阳性细胞数、IL-10、TGF-β1水平、TMEM240、ARHGDIB蛋白表达显著升高（P＜0.05）。sh-TMEM240逆转了穗花杉双黄酮对模型小鼠神经炎症的改善作用。结论 穗花杉双黄酮通过上调TMEM240/ARHGDIB信号轴，抑制NF-κB通路的过度活化进而改善脑缺血再灌注损伤小鼠的神经炎症。

Objective To investigate the mechanism of amentoflavone improves neuroinflammation in mice with cerebral ischemia-reperfusion injury based on TMEM240/ARHGDIB pathway. Methods A cerebral ischemia-reperfusion injury model was established using the suture occlusion method. Successfully modeled mice were randomly divided into model group, amentoflavone group, amentoflavone + sh-NC group, amentoflavone + sh-TMEM240 group, and sham-operated group with 24 mice in each group. Assessments included the Zea-longa score, balance beam test score, cerebral infarction volume ratio, and brain water content. Histopathological changes in the ischemic brain tissue were evaluated by HE staining. Immunofluorescence staining was used to detect the numbers of Iba-1⁺iNOS⁺ and Iba-1⁺Arg-1⁺ positive cells. ELISA measured the levels of TNF-α, IL-1β, IL-6, IL-10, and TGF-β1 in the ischemic brain tissue. Western blotting analyzed the protein expression of TMEM240, ARHGDIB, and p-NF-κB p65/NF-κB p65. Results Compared with the model group, the pathological damage of the ischemic side brain tissue in the Zea-longa flavone group was significantly improved, manifested as more intact neuronal morphology, reduced edema, and decreased inflammatory infiltration. The scores of Zea-longa, the ratio of cerebral infarction volume, the water content of brain tissue, the number of Iba-1⁺ iNOS⁺ positive cells in the ischemic side brain tissue, the levels of TNF-α, IL-1β, and IL-6, and the protein expression of p-NF-κB p65/NF-κB p65 were significantly decreased. The balance beam test score, the number of Iba-1⁺ Arg-1⁺ positive cells in the ischemic side brain tissue, the levels of IL-10 and TGF-β1, the protein expression of TMEM240 and ARHGDIB were significantly increased (P < 0.05). sh-TMEM240 reversed the improvement effect of amentoflavone on the neuroinflammation in the amentoflavone group. Conclusion Amentoflavone improves neuroinflammation in cerebral ischemia-reperfusion injury mice by upregulating the TMEM240/ARHGDIB signaling axis and subsequently inhibiting the overactivation of the NF-κB pathway.

R286.1

河南省青年科学基金项目（242300421515）；河南省高等学校重点科研项目计划（24A320029）