title: Botulinum neurotoxin type A induces TLR2-mediated inflammatory responses in macrophages.
abstract: Botulinum neurotoxin type A (BoNT/A) is the most potent protein toxin and causes fatal flaccid muscle paralysis by blocking neurotransmission. Application of BoNT/A has been extended to the fields of therapeutics and biodefense. Nevertheless, the global response of host immune cells to authentic BoNT/A has not been reported. Employing microarray analysis, we performed global transcriptional profiling of RAW264.7 cells, a murine alveolar macrophage cell line. We identified 70 genes that were modulated following 1 nM BoNT/A treatment. The altered genes were mainly involved in signal transduction, immunity and defense, protein metabolism and modification, neuronal activities, intracellular protein trafficking, and muscle contraction. Microarray data were validated with real-time RT-PCR for seven selected genes including tlr2, tnf, inos, ccl4, slpi, stx11, and irg1. Proinflammatory mediators such as nitric oxide (NO) and tumor necrosis factor alpha (TNFα) were induced in a dose-dependent manner in BoNT/A-stimulated RAW264.7 cells. Increased expression of these factors was inhibited by monoclonal anti-Toll-like receptor 2 (TLR2) and inhibitors specific to intracellular proteins such as c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p38 mitogen-activated protein kinase (MAPK). BoNT/A also suppressed lipopolysaccharide-induced NO and TNFα production from RAW264.7 macrophages at the transcription level by blocking activation of JNK, ERK, and p38 MAPK. As confirmed by TLR2-/- knock out experiments, these results suggest that BoNT/A induces global gene expression changes in host immune cells and that host responses to BoNT/A proceed through a TLR2-dependent pathway, which is modulated by JNK, ERK, and p38 MAPK.
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Yun Jeong
authors: YJ
authors: Kim
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Jeong-Hee
authors: JH
authors: Kim
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Kwang-Jun
authors: KJ
authors: Lee
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Myung-Min
authors: MM
authors: Choi
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Yeon Hee
authors: YH
authors: Kim
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Gi-Eun
authors: GE
authors: Rhie
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Cheon-Kwon
authors: CK
authors: Yoo
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Kiweon
authors: K
authors: Cha
authors: Division of High-risk Pathogen Research, Center for Infectious Diseases, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, Korea.
authors: Na-Ri
authors: NR
authors: Shin
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D019274:Botulinum Toxins, Type A / Q000633:toxicity*; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D055786:Gene Knockout Techniques; D007113:Immunity, Innate / Q000187:drug effects; D007249:Inflammation / Q000139:chemically induced / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009569:Nitric Oxide / Q000096:biosynthesis / Q000378:metabolism; D000067996:RAW 264.7 Cells; D015398:Signal Transduction / Q000187:drug effects; D051195:Toll-Like Receptor 2 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D051195:Toll-Like Receptor 2; D009569:Nitric Oxide; D019274:Botulinum Toxins, Type A