Astrocyte over-activation and extensive neuron loss are the main characteristic pathological features of spinal cord ischemia–reperfusion injury (SCII). Prior studies have placed substantial emphasis on the role of heat shock protein family A member 8 (HSPA8) on postischemic myocardial inflammation and cardiac dysfunction. However, it has never been determined whether HSPA8 participates in astrocyte activation and thus mediated neuroinflammation associated with SCII. The left renal artery ligation-induced SCII rat models and oxygen–glucose deprivation and reoxygenation (OGD/R)-induced rat primary cultured astrocytes were established. The lentiviral vector encoding short hairpin RNA targeting HSPA8 was delivered to the spinal cord by intrathecal administration or to culture astrocytes. Then, the spinal neuron survival, gliosis, and nod-like receptor pyrin domain-containing 3 (NLRP3) inflammasome and its related pro-inflammatory cytokines were analyzed. SCII significantly enhanced the GFAP and HSPA8 expression in the spinal cord, resulting in blood–brain barrier breakdown and the dramatical loss of spinal neuron and motor function. Moreover, injury also increased spinal nuclear factor-kappa B (NF-κB) p65 phosphorylation, NLRP3 inflammasome-mediated caspase-1 activation, and subsequent interleukin (IL)-1β as well as IL-18 secretion. Silencing the HSPA8 expression efficiently ameliorated the spinal cord tissue damage and promoted motor function recovery after SCII, through blockade of the astrocyte activation and levels of phosphorylated NF-κB, NLRP3, caspase-1, IL-1β, and IL-18. Further in vitro studies confirmed that HSPA8 knockdown protected astrocytes from OGD/R-induced injury via the blockade of NF-κB and NLRP3 inflammasome activation. Our findings indicate that knockdown of HSPA8 inhibits spinal astrocytic damage after SCII, which may provide a promising therapeutic strategy for SCII treatment.

中文翻译：

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抑制热休克蛋白家族 A 成员 8 通过星形胶质细胞 NF-κB/NLRP3 炎性体通路减轻脊髓缺血再灌注损伤

星形胶质细胞过度激活和广泛的神经元丢失是脊髓缺血再灌注损伤（SCII）的主要病理特征。先前的研究非常重视热休克蛋白家族 A 成员 8 (HSPA8) 在缺血后心肌炎症和心脏功能障碍中的作用。然而，从未确定 HSPA8 是否参与星形胶质细胞激活，从而介导与 SCII 相关的神经炎症。建立了左肾动脉结扎诱导的SCII大鼠模型和氧-葡萄糖剥夺和复氧(OGD/R)诱导的大鼠原代培养星形胶质细胞。编码靶向 HSPA8 的短发夹 RNA 的慢病毒载体通过鞘内给药或培养星形胶质细胞被递送至脊髓。然后，脊髓神经元存活，神经胶质增生，并分析了含有点头样受体pyrin结构域的3（NLRP3）炎性体及其相关的促炎细胞因子。SCII 显着增强脊髓中 GFAP 和 HSPA8 的表达，导致血脑屏障破坏以及脊髓神经元和运动功能的显着丧失。此外，损伤还增加了脊髓核因子-kappa B (NF-κB) p65 磷酸化、NLRP3 炎症小体介导的 caspase-1 活化，以及随后的白细胞介素 (IL)-1β 和 IL-18 分泌。通过阻断星形胶质细胞的活化和磷酸化 NF-κB、NLRP3、caspase-1、IL-1β 和 IL-18 的水平，沉默 HSPA8 表达可有效改善脊髓组织损伤并促进 SCII 后的运动功能恢复。进一步的体外研究证实，HSPA8 敲低通过阻断 NF-κB 和 NLRP3 炎性体激活来保护星形胶质细胞免受 OGD/R 诱导的损伤。我们的研究结果表明，HSPA8 的敲低可抑制 SCII 后的脊髓星形胶质细胞损伤，这可能为 SCII 治疗提供有希望的治疗策略。