The NADPH oxidase enzyme as a therapeutic target after Spinal Cord Injury

描述（由申请人提供）：目前美国有超过25万人患有脊髓损伤（SCI）引起的残疾，每年报告约12，000例新损伤。炎症，包括小胶质细胞的活化和巨噬细胞的侵入，在SCI后观察到的继发性损伤中起着核心作用。SCI后，NOX家族的酶长期上调，并可能通过产生活性氧（ROS）促进小胶质细胞/巨噬细胞活化、炎症和组织损伤。这种表达谱与巨噬细胞的侵袭和小胶质细胞活化成促炎或M1表型相一致，但与M2或抗炎表型不一致。NOX 2亚型的敲除可以减少ROS的产生。此外，急性药理学抑制NOX 2减少了大鼠SCI后的病变体积。然而，该酶家族的表达谱目前尚不清楚，并且NOX 2抑制对SCI后慢性炎症、轴突保留或运动或自主神经功能的影响尚不清楚。因此，我们假设NOX 2是脊髓损伤后M1小胶质细胞/巨噬细胞中表达的主要NOX亚型，并且抑制NOX 2将减少慢性炎症并改善恢复，包括轴突发芽/保留以及运动和自主神经功能。为了验证这一假设，我们提出了三个具体目标。目的1：明确脊髓损伤后NOX亚型的表达谱及其与M1和M2表型的相关性。在这个目标中，我们将使用免疫组织化学来确定NOX亚型的表达谱及其与M1和M2活化标志物的共定位。在目标2中，我们将证明NOX 2抑制诱导小胶质细胞和巨噬细胞中的M2表型，并减少损伤脊髓中的炎症。为此，我们将使用基因敲除（gp 91 PHOX）和药理学抑制方法评估中度SCI后急性、亚急性和慢性时间点的炎症。最后，在目标3中，我们将表明延迟给予N 0X 2抑制剂改善脊髓损伤后的轴突发芽和保留，并恢复运动和自主神经功能。利用NOX 2特异性抑制剂，我们将评估运动和自主神经功能，以及轴突保留和发芽。拟议的研究将阐明SCI后NOX的作用，并展示NOX抑制的治疗应用。这些研究的数据将为炎症（包括慢性炎症）在SCI恢复中的作用提供深入了解。

DESCRIPTION (provided by applicant): Over 250,000 people in the United States are currently living with disabilities caused by spinal cord injury (SCI), and approximately 12,000 new injuries are reported each year. Inflammation, including activation of microglia and invasion of macrophages, plays a central role in the secondary injury observed after SCI. After SCI, the NOX family of enzymes is chronically up-regulated and may contribute to microglial/macrophage activation, inflammation and tissue damage through the production of reactive oxygen species (ROS). This expression profile coincides with the invasion of macrophages and activation of microglia into a pro-inflammatory, or M1, phenotype, but not with the M2, or anti-inflammatory, phenotype. Knockout of the NOX2 isoform can reduce ROS production. Further, acute pharmacological inhibition of NOX2 reduces lesion volume following SCI in rats. However, the expression profile of the family of enzymes is currently unclear and the effect of NOX2 inhibition on chronic inflammation, axonal sparing, or motor or autonomic function after SCI is unknown. Therefore, we hypothesize that NOX2 is the primary NOX isoform expressed in M1 microglia/macrophages after spinal cord injury and that inhibition of NOX2 will reduce chronic inflammation and improve recovery, including axonal sprouting/sparing and motor and autonomic function. To test this hypothesis, we propose three specific aims. In aim 1, we will identify the NOX isoform expression profile and correlation with M1 and M2 phenotype after spinal cord injury. In this aim, we will use immunohistochemistry to identify the expression profile of the NOX isoforms and their co-localization with M1 and M2 activation markers. In aim 2, we will demonstrate that NOX2 inhibition induces an M2 phenotype in microglia and macrophages and reduces inflammation in the injured spinal cord. In this aim, we will assess inflammation after moderate SCI at acute, sub-acute and chronic time points using both genetic knockout (gp91PHOX) and pharmacological inhibition approaches. Finally, in aim 3 we will show that delayed administration of a NOX2 inhibitor improves axonal sprouting and sparing and restores motor and autonomic function after spinal cord injury. Utilizing a NOX2 specific inhibitor, we will assess motor and autonomic function, as well as axonal sparing and sprouting. The proposed research study will clarify the role of NOX after SCI and demonstrate the therapeutic applications of NOX inhibition. The data from these studies will provide insight into the role of inflammation, including chronic inflammation, in recovery from SCI.