Department of Critical Care Medicine, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, China.
Department of Rehabilitation Medicine, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, China.
Brain Res Bull. 2024 Oct 1;216:111055. doi: 10.1016/j.brainresbull.2024.111055. Epub 2024 Aug 20.
Cerebral ischemia is characterized by its rapid onset and high rates of recurrence, morbidity, and mortality, with blood-brain barrier (BBB) permeability playing a vital role in brain injury. Therefore, it is important to understand the molecular mechanism which regulates the BBB during cerebral ischemia.
An in vitro model of oxygen-glucose deprivation (OGD) and an in vivo model of cerebral ischemia/reperfusion (I/R) were constructed. PD-1 overexpression vectors and vectors containing si-RNA were transfected and injected into in vitro and in vivo models. Western blotting, real-time quantitative PCR (qPCR), immunofluorescence (IF) analysis, and immunohistochemical staining were employed to evaluate the expression levels of programmed cell death-1 (PD-1), microglia M1 and M2 biomarkers, and tight junction proteins. Flow cytometry and ELISA were used to measure the levels of pro-inflammatory cytokines. The BBB permeability of brain tissues was evaluated by Evans blue dye (EBD) extravasation and transendothelial electrical resistance (TEER). Brain water content was measured to assess the extent of inflammatory exudation. The infarct volume and neurological severity score (NSS) were used to assess the severity of brain injury. Brain cell apoptosis was assessed by the TUNEL assay and hematoxylin-eosin (H&E) staining.
PD-1 helped to convert the microglia M1 phenotype to the M2 phenotype and to reduce BBB permeability both in vitro and in vivo. Overexpression of PD-1 promoted a shift of the M1 phenotype to the M2 phenotype and reduced BBB permeability via the ERK and p38 MAPK signaling pathways. PD-1 reduced inflammatory exudation, BBB permeability, cell apoptosis, and brain injury in vivo.
Our present study verified that PD-1 exerts an anti-inflammatory effect by converting the microglia M1 phenotype to the M2 phenotype, reducing BBB permeability, and thereby relieves brain injury caused by cerebral ischemia. PD-1 is potential therapeutic target for brain injury caused by cerebral ischemia.
脑缺血具有起病迅速、复发率高、发病率和死亡率高的特点，血脑屏障（BBB）通透性在脑损伤中起着至关重要的作用。因此，了解脑缺血时调节 BBB 的分子机制非常重要。
构建了氧葡萄糖剥夺（OGD）体外模型和脑缺血再灌注（I/R）体内模型。转染 PD-1 过表达载体和 si-RNA 载体，并注入体外和体内模型。采用 Western blot、实时定量 PCR（qPCR）、免疫荧光（IF）分析和免疫组织化学染色评估程序性细胞死亡-1（PD-1）、小胶质细胞 M1 和 M2 生物标志物以及紧密连接蛋白的表达水平。流式细胞术和 ELISA 用于测量促炎细胞因子的水平。通过 Evans 蓝染料（EBD）渗出和跨内皮电阻（TEER）评估脑组织的 BBB 通透性。测量脑水含量以评估炎症渗出的程度。用脑梗死体积和神经功能缺损评分（NSS）评估脑损伤的严重程度。通过 TUNEL 检测和苏木精-伊红（H&E）染色评估脑细胞凋亡。
PD-1 有助于在体外和体内将小胶质细胞 M1 表型转化为 M2 表型并降低 BBB 通透性。PD-1 的过表达通过 ERK 和 p38 MAPK 信号通路促进 M1 表型向 M2 表型的转变并降低 BBB 通透性。PD-1 减少了体内炎症渗出、BBB 通透性、细胞凋亡和脑损伤。
本研究证实 PD-1 通过将小胶质细胞 M1 表型转化为 M2 表型、降低 BBB 通透性来发挥抗炎作用，从而减轻脑缺血引起的脑损伤。PD-1 是脑缺血引起的脑损伤的潜在治疗靶点。
