肼苯哒嗪通过PI3K/Akt通路调控巨噬/小胶质细胞M1/M2亚型转换保护小鼠脊髓损伤后焦虑、抑郁的机制研究

Spinal cord injury (SCI) is a common central nervous system injury, and the incidence of anxiety or depression concomitant with SCI is high. Oxidative stress is an important factor in secondary spinal cord injury. As a kind of highly active α, β- unsaturated aldehydes, acrolein can cause extensive oxidative stress and inflammation in the spinal cord. The M1 and M2 subtypes of macrophages/microglia cells transformation after SCI are an important part of inflammatory response. Although acrolein and its scavenger hydralazine can worsen or improve SCI, but whether it could regulate and control M1 and M2 subtype of macrophage/microglia cells transformation, thereby to play a protective role after SCI, and its mechanism are still unknown. The results of our preliminary study showed that the application of hydralazine can increase the M2 subtype of macrophages in SCI and improve anxiety and depression after SCI. Based on the preliminary results, this subject proposed to study the mechanism of acrolein scavenger hydralazine on inflammation control, reveal its protective mechanism through regulating macrophage/microglia subtype transformation, and thereby improving anxiety or depression after SCI. These studies can expand the understanding of acrolein and its scavenger, hydralazine, and find its protective mechanism after SCI, and provide important experimental evidence for clinical application.

脊髓损伤是临床常见的中枢神经系统损伤，脊髓损伤后焦虑、抑郁等不良情绪发生率高。氧化应激是继发性脊髓损伤的重要因素。以丙烯醛（acrolein）为代表的一类高度活性α, β-不饱和醛类可引起广泛的脊髓氧化应激损伤和炎症反应。而脊髓损伤后巨噬/小胶质细胞的M1、M2亚型转换是炎症反应的重要环节。虽然丙烯醛及其清除剂肼苯哒嗪可引起或改善脊髓损伤反应，但其是否可通过调控巨噬/小胶质细胞的M1、M2亚型转换发挥保护作用，其机制如何，目前尚不明确。我们前期研究结果显示，应用肼苯哒嗪可增加脊髓损伤部位M2型巨噬细胞数量，改善脊髓损伤后焦虑、抑郁程度。本课题拟在此基础上，研究肼苯哒嗪控制炎症反应的机制，揭示其调控巨噬/小胶质细胞亚型转换，保护脊髓损伤，改善损伤后焦虑、抑郁的机理，阐明其保护作用机制。这些研究可拓展对丙烯醛及其清除剂的认识，发现肼苯哒嗪保护脊髓损伤的作用机制，为临床应用提供重要实验依据。

脊髓损伤是临床常见的中枢神经系统损伤，脊髓损伤后焦虑、抑郁等不良情绪发生率高。氧化应激是继发性脊髓损伤的重要因素。以丙烯醛（acrolein）为代表的一类高度活性α,β-不饱和醛类可引起广泛的脊髓氧化应激损伤和炎症反应。而脊髓损伤后巨噬/小胶质细胞的M1、M2亚型转换是炎症反应的重要环节。虽然丙烯醛及其清除剂肼苯哒嗪可引起或改善脊髓损伤反应，但其是否可通过调控巨噬/小胶质细胞的M1、M2亚型转换发挥保护作用，改善脊髓损伤后焦虑抑郁样行为，其作用机制如何，目前尚不明确。该基金项目通过在脊髓损伤小鼠模型中应用肼苯哒嗪，系统对比观察了（1）脊髓损伤应用肼苯哒嗪小鼠运动、感觉功能变化及焦虑、抑郁等情绪行为变化；（2）脊髓损伤应用肼苯哒嗪小鼠脊髓、DRG 组织丙烯醛表达与巨噬/小胶质细胞M1、M2 表型变化及局部炎症因子之间的关系；（3）脊髓损伤应用肼苯哒嗪对小鼠脊髓和焦虑、抑郁相关脑区（杏仁核、海马）PI3K/Akt 通路活化的分子生物学检测；（4）检测脊髓损伤应用肼苯哒嗪对小鼠焦虑、抑郁相关脑区（杏仁核、海马）的小胶质细胞亚型转换、神经纤维及兴奋性和抑制性神经递质变化情况的影响；（5）并通过体外验证肼苯哒嗪处理的巨噬细胞丙烯醛表达及对脊髓神经元细胞生长的影响。结果发现应用肼苯哒嗪可显著减少损伤后丙烯醛表达、抑制脊髓损伤区及焦虑抑郁相关脑区巨噬/小胶质细胞激活浸润，增加M2型亚型巨噬细胞浸润，改变相关神经递质表达，改善损伤后焦虑抑郁样行为，而其背后机制则和PI3K/Akt 通路磷酸化激活有关，体外研究也证实了肼苯哒嗪对巨噬细胞及神经元细胞生长的良性促进作用。本研究阐明了肼苯哒嗪通过清除氧自由基acrolein来调控脊髓损伤后炎症反应的机制，揭示了其调控巨噬/小胶质细胞亚型转换、保护脊髓损伤、改善损伤后焦虑、抑郁样行为的机理，阐明了其脊髓损伤保护作用机制。这些研究可拓展对丙烯醛及其清除剂的认识，发现肼苯哒嗪保护脊髓损伤的作用机理，为进一步临床应用提供重要实验依据。