The Role of Neuroglobin in the Neuroprotection of Limb Ischemic Preconditioning in Rats

Recent evidence suggests that limb ischemic preconditioning (LIP) protects neurons against cerebral ischemia-reperfusion injury. However, the mechanisms of LIP are not well understood. Neuroglobin (Ngb) is a recently discovered globin that affords protection against hypoxic/ischemic brain injury. This study was performed to investigate the role of Ngb in the neuroprotection of LIP against brain ischemia and the involvements of mitochondria in the process. The rat global brain ischemic model was used, and the CA1 hippocampus was selected as the observational target. Ngb expression was investigated by RT-PCR and Western blot. Neuropathological evaluation was performed by thionin staining. Mitochondrial membrane potential (Delta psi m), Na+-K+-ATPase activity, and ultrastructure were examined by flow cytometry, spectrophotometry, and transmission electron microscopy, respectively. We also used Ngb antisense oligodeoxynucleotides (AS-ODNs) and Ngb inducer hemin to inhibit or mimic the effect of LIP. We found that LIP significantly up-regulated Ngb expression and protected neurons against ischemia. Furthermore, LIP effectively improved deterioration in the Delta psi m, mitochondrial Na+-K+-ATPase activity, and ultrastructure induced by cerebral ischemia. These effects of LIP were inhibited partly by Ngb AS-ODNs and mimicked by hemin. It could be concluded that up-regulation of Ngb expression played an important role in the neuroprotection induced by LIP, and the Ngb-mediated neuroprotection of LIP was, at least partly, associated with mitochondria.

近期证据表明，肢体缺血预处理（LIP）可保护神经元免受脑缺血再灌注损伤。然而，LIP的机制尚未完全明确。脑红蛋白（Ngb）是一种新近发现的珠蛋白，对缺氧/缺血性脑损伤具有保护作用。本研究旨在探讨Ngb在LIP对脑缺血的神经保护中的作用以及线粒体在该过程中的参与情况。采用大鼠全脑缺血模型，并选择CA1海马作为观察靶点。通过逆转录聚合酶链反应（RT - PCR）和蛋白质印迹法（Western blot）研究Ngb的表达。通过硫堇染色进行神经病理学评估。分别采用流式细胞术、分光光度法和透射电子显微镜检测线粒体膜电位（ΔΨm）、Na⁺ - K⁺ - ATP酶活性和超微结构。我们还使用Ngb反义寡脱氧核苷酸（AS - ODNs）和Ngb诱导剂氯化血红素抑制或模拟LIP的作用。我们发现LIP显著上调Ngb表达并保护神经元免受缺血损伤。此外，LIP有效改善了脑缺血引起的ΔΨm、线粒体Na⁺ - K⁺ - ATP酶活性和超微结构的恶化。LIP的这些作用部分被Ngb AS - ODNs抑制，并被氯化血红素模拟。可以得出结论，Ngb表达的上调在LIP诱导的神经保护中起重要作用，并且LIP的Ngb介导的神经保护至少部分与线粒体有关。