Department of Anatomy and Center for Advanced Medical Education (BK21 Project), Inha University School of Medicine, 7-241 Shinheung-dong, Jung-Gu, Inchon, 400-712, Republic of Korea.
Neurotox Res. 2011 Aug;20(2):159-69. doi: 10.1007/s12640-010-9231-x. Epub 2010 Nov 30.
High mobility group box 1 (HMGB1) was originally identified as ubiquitously expressed nonhistone DNA-binding protein, but recently, it was found to act as an endogenous danger molecule, which signals danger and traumatic cell death. Previously, the authors showed that HMGB1 is massively released immediately after an ischemic insult and that it subsequently activates microglia and induces inflammation in the postischemic brain. Here, we showed the endogenous danger molecule-like function of HMGB1 in primary cortical cultures. HMGB1 was found to be accumulated in NMDA-treated primary cortical culture media, and media collected from these cultures were able to induce neuronal cell death when added to fresh primary cortical cultures. However, HMGB1-depleted NMDA-conditioned media produced by HMGB1 siRNA transfection or by preincubation with anti-HMGB1 antibody or with HMGB1 A box failed to induce neuronal cell death. Furthermore, siRNA-mediated HMGB1 knockdown substantially suppressed NMDA- or Zn(2+)-induced cell death. It was interesting to find that extracellular HMGB1-induced neuronal apoptosis, as evidenced by TUNEL staining and caspase 3 assay in combination with double immunofluorescence staining. A series of RAGE and HMGB1 co-immunoprecipitation experiments in the presence of SB203580 and PD98059 (p38 MAPK and ERK inhibitors, respectively) demonstrated that RAGE-p38 MAPK and RAGE-ERK pathway might underlie extracellular HMGB1-mediated neuronal apoptosis. These results together with our previous reports regarding microglial activation by extracellular HMGB1 indicate that HMGB1 functions as a novel danger signal, which aggravates brain damage via autocrine and paracrine manners.
高迁移率族蛋白 B1（HMGB1）最初被鉴定为广泛表达的非组蛋白 DNA 结合蛋白，但最近发现它作为一种内源性危险分子，可发出危险和创伤性细胞死亡的信号。作者先前表明，HMGB1 在缺血性损伤后立即大量释放，随后激活小胶质细胞并在缺血后大脑中引发炎症。在这里，我们展示了 HMGB1 在原代皮质培养物中的内源性危险分子样功能。发现 HMGB1 在 NMDA 处理的原代皮质培养物培养基中积累，并且从这些培养物收集的培养基在添加到新鲜的原代皮质培养物时能够诱导神经元细胞死亡。然而，通过 HMGB1 siRNA 转染或通过与抗 HMGB1 抗体或 HMGB1 A 盒预孵育产生的 HMGB1 耗尽的 NMDA 条件培养基均不能诱导神经元细胞死亡。此外，siRNA 介导的 HMGB1 敲低可显著抑制 NMDA 或 Zn(2+)诱导的细胞死亡。有趣的是发现，细胞外 HMGB1 诱导的神经元凋亡，如 TUNEL 染色和 caspase 3 测定与双免疫荧光染色结合所证明的那样。在存在 SB203580 和 PD98059（p38 MAPK 和 ERK 抑制剂，分别）的情况下进行的一系列 RAGE 和 HMGB1 共免疫沉淀实验表明，RAGE-p38 MAPK 和 RAGE-ERK 途径可能是细胞外 HMGB1 介导的神经元凋亡的基础。这些结果以及我们以前关于细胞外 HMGB1 激活小胶质细胞的报告表明，HMGB1 作为一种新型危险信号，通过自分泌和旁分泌方式加重脑损伤。
