Department of Anesthesiology & Critical Care Medicine, Medical Center, University of Freiburg, Germany.
Faculty of Medicine, University of Freiburg, Germany.
Glia. 2020 Nov;68(11):2427-2445. doi: 10.1002/glia.23864. Epub 2020 May 31.
Microglial erythrophagocytosis is crucial in injury response to hemorrhagic stroke. We hypothesized that regulation of microglial erythrophagocytosis via HO-1/CO depends on a pathway involving reactive oxygen species (ROS) and CD36 surface-expression. The microglial BV-2 cell line and primary microglia (PMG) were incubated +/-blood and +/-CO-exposure. PMG isolated from tissue-specific HO-1-deficient (LyzM-Cre-Hmox1 ) and CD36 mice or siRNA against AMPK (AMP-activated protein kinase) were used to test our hypothesis. In a murine subarachnoid hemorrhage (SAH) model, we compared neuronal injury in wild-type and CD36 mice. Readouts included vasospasm, microglia activation, neuronal apoptosis, and spatial memory. We observed increased microglial HO-1-expression after blood-exposure. A burst in ROS-production was seen after CO-exposure, which led to increased amounts of phosphorylated AMPK with subsequently enhanced CD36 surface-expression. Naïve PMG from LyzM-Cre-Hmox1 mice showed reduced ROS-production and CD36 surface-expression and failed to respond to CO with increased CD36 surface-expression. Lack of HO-1 and CD36 resulted in reduced erythrophagocytosis that could not be rescued with CO. Erythrophagocytosis was enhanced in BV-2 cells in the presence of exogenous CO, which was abolished in cells treated with siRNA to AMPK. CD36 mice subjected to SAH showed enhanced neuronal cell death, which resulted in impaired spatial memory function. We demonstrate that microglial phagocytic function partly depends on a pathway involving HO-1 with changes in ROS-production, phosphorylated AMPK, and surface expression of CD36. CD36 was identified as a crucial component in blood clearance after hemorrhage that ultimately determines neuronal outcome. These results demand further investigations studying the potential neuroprotective properties of CO.
小胶质细胞的红细胞吞噬作用对于出血性中风的损伤反应至关重要。我们假设，通过 HO-1/CO 调节小胶质细胞的红细胞吞噬作用依赖于一条涉及活性氧 (ROS) 和 CD36 表面表达的途径。BV-2 细胞系和原代小胶质细胞 (PMG) 在孵育 +/-血和 +/-CO 暴露。使用组织特异性 HO-1 缺陷型 (LyzM-Cre-Hmox1 ) 和 CD36 小鼠或针对 AMPK (AMP 激活蛋白激酶) 的 siRNA 的 PMG 来测试我们的假设。在小鼠蛛网膜下腔出血 (SAH) 模型中，我们比较了野生型和 CD36 小鼠的神经元损伤。检测指标包括血管痉挛、小胶质细胞激活、神经元凋亡和空间记忆。我们观察到血液暴露后小胶质细胞 HO-1 表达增加。CO 暴露后观察到 ROS 产生爆发，导致磷酸化 AMPK 增加，随后 CD36 表面表达增强。LyzM-Cre-Hmox1 小鼠的幼稚 PMG 显示 ROS 产生和 CD36 表面表达减少，并且无法对 CO 做出反应，增加 CD36 表面表达。缺乏 HO-1 和 CD36 导致红细胞吞噬作用减少，而 CO 无法挽救。在存在外源性 CO 的情况下，BV-2 细胞中的红细胞吞噬作用增强，而在用 AMPK 的 siRNA 处理的细胞中则被消除。在发生 SAH 的 CD36 小鼠中，神经元细胞死亡增加，导致空间记忆功能受损。我们证明小胶质细胞吞噬功能部分依赖于一条涉及 HO-1 的途径，该途径涉及 ROS 产生、磷酸化 AMPK 和 CD36 表面表达的变化。CD36 被确定为出血后血液清除的关键组成部分，最终决定神经元结局。这些结果需要进一步研究，以研究 CO 的潜在神经保护特性。
