Department of Anatomy and Center for Advanced Medical Education (BK21 Project), Inha University School of Medicine, Inchon, Republic of Korea.
Neurosci Lett. 2012 Sep 13;525(2):179-83. doi: 10.1016/j.neulet.2012.07.040. Epub 2012 Jul 31.
High mobility group box 1 (HMGB1) is an endogenous danger signal molecule. In a previous report, we showed that HMGB1 is massively released during NMDA-induced acute damaging process in the postischemic brain and triggers inflammatory processes and induces neuronal apoptosis. We have also reported a robust neuroprotection of intranasally delivered HMGB1 siRNA in the postischemic rat brain (middle cerebral artery occlusion (MCAO), 60 min). In the present study, we investigated the therapeutic efficacy of intranasally delivered HMGB1 binding heptamer peptide (HBHP; HMSKPVQ), which was selected using a phage display approach, in the same stroke animal model. A pull-down assay using biotin-labeled HBHP showed that HBHP binds directly to HMGB1, specifically to HMGB1 A box, confirming HMGB1/HBHP interaction. HBHP significantly suppressed HMGB1-mediated neuronal cell death in primary cortical cultures and HMGB1/HBHP binding was detected in NMDA-conditioned culture media. However, a heptamer peptide composed of a scrambled sequence of the seven amino acids in HBHP failed to bind HMGB1 and had no protective effect. Furthermore, HBHP (300 ng) delivered intranasally at 30 min before MCAO significantly suppressed infarct volume in the postischemic rat brain (maximal reduction by 41.8±5.4%) and ameliorated neurological and behavioral deficits. In contrast, scrambled heptamer peptide had no protective effect at the same dose. Together these results suggest that intranasal HBHP ameliorates neuronal damage in the ischemic brain by binding HMGB1, which might inhibit the function of HMGB1 as an endogenous danger signal molecule.
高迁移率族蛋白 B1（HMGB1）是一种内源性危险信号分子。在之前的报告中，我们表明 HMGB1 在 NMDA 诱导的缺血后大脑急性损伤过程中大量释放，并引发炎症过程和诱导神经元凋亡。我们还报告了在缺血后大鼠大脑（大脑中动脉闭塞（MCAO），60 分钟）中，鼻内给予 HMGB1 siRNA 具有强大的神经保护作用。在本研究中，我们使用噬菌体展示方法选择了高迁移率族蛋白 B1 结合七肽（HBHP；HMSKPVQ），研究了其在相同的卒中动物模型中的治疗效果。使用生物素标记的 HBHP 进行的下拉测定表明，HBHP 直接与 HMGB1 结合，特异性地与 HMGB1 A 盒结合，证实了 HMGB1/HBHP 相互作用。HBHP 显著抑制原代皮质培养物中 HMGB1 介导的神经元细胞死亡，并且在 NMDA 条件培养基中检测到 HMGB1/HBHP 结合。然而，由 HBHP 中七个氨基酸的七肽组成的七肽与 HMGB1 结合，没有保护作用。此外，在 MCAO 前 30 分钟鼻内给予 300 ng HBHP 可显著抑制缺血后大鼠大脑中的梗死体积（最大减少 41.8±5.4%），并改善神经功能和行为缺陷。相比之下，相同剂量的 scrambled 七肽没有保护作用。这些结果表明，鼻内 HBHP 通过结合 HMGB1 改善缺血性脑损伤中的神经元损伤，这可能抑制 HMGB1 作为内源性危险信号分子的功能。
