Extracellular HMGB1 released by NMDA treatment confers neuronal apoptosis via RAGE-p38 MAPK/ERK signaling pathway.
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.
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.
Seung-Woo
SW
Kim
Chae-Moon
CM
Lim
Jung-Bin
JB
Kim
Joo-Hyun
JH
Shin
Sanghyun
S
Lee
Minhyung
M
Lee
Ja-Kyeong
JK
Lee
D000704:Analysis of Variance; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002540:Cerebral Cortex / Q000166:cytology; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D018690:Excitatory Amino Acid Agonists; D045604:Extracellular Fluid / Q000187:drug effects*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D024243:HMGB1 Protein / Q000378:metabolism* / Q000494:pharmacology; D047468:Immunoprecipitation; D020287:In Situ Nick-End Labeling / Q000379:methods; D051379:Mice; D016202:N-Methylaspartate / Q000494:pharmacology*; D009457:Neuroglia / Q000187:drug effects / Q000502:physiology; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014162:Transfection; D015032:Zinc / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D005904:Glial Fibrillary Acidic Protein; D024243:HMGB1 Protein; D034741:RNA, Small Interfering; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; D016202:N-Methylaspartate; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D053148:Caspase 3; D010751:Phosphopyruvate Hydratase; D015032:Zinc