title: NOD2/RICK-dependent β-defensin 2 regulation is protective for nontypeable Haemophilus influenzae-induced middle ear infection.
abstract: Middle ear infection, otitis media (OM), is clinically important due to the high incidence in children and its impact on the development of language and motor coordination. Previously, we have demonstrated that the human middle ear epithelial cells up-regulate β-defensin 2, a model innate immune molecule, in response to nontypeable Haemophilus influenzae (NTHi), the most common OM pathogen, via TLR2 signaling. NTHi does internalize into the epithelial cells, but its intracellular trafficking and host responses to the internalized NTHi are poorly understood. Here we aimed to determine a role of cytoplasmic pathogen recognition receptors in NTHi-induced β-defensin 2 regulation and NTHi clearance from the middle ear. Notably, we observed that the internalized NTHi is able to exist freely in the cytoplasm of the human epithelial cells after rupturing the surrounding membrane. The human middle ear epithelial cells inhibited NTHi-induced β-defensin 2 production by NOD2 silencing but augmented it by NOD2 over-expression. NTHi-induced β-defensin 2 up-regulation was attenuated by cytochalasin D, an inhibitor of actin polymerization and was enhanced by α-hemolysin, a pore-forming toxin. NOD2 silencing was found to block α-hemolysin-mediated enhancement of NTHi-induced β-defensin 2 up-regulation. NOD2 deficiency appeared to reduce inflammatory reactions in response to intratympanic inoculation of NTHi and inhibit NTHi clearance from the middle ear. Taken together, our findings suggest that a cytoplasmic release of internalized NTHi is involved in the pathogenesis of NTHi infections, and NOD2-mediated β-defensin 2 regulation contributes to the protection against NTHi-induced otitis media.
authors.affiliation: Department of Head and Neck Surgery, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, California, United States of America.
authors.forename: Jeong-Im
authors.initials: JI
authors.lastname: Woo
authors.affiliation: Division of Clinical and Translational Research, House Research Institute, Los Angeles, California, United States of America.
authors.forename: Sejo
authors.initials: S
authors.lastname: Oh
authors.affiliation: Center for Electron Microscopy and Microanalysis, University of Southern California, Los Angeles, California, United States of America.
authors.forename: Paul
authors.initials: P
authors.lastname: Webster
authors.affiliation: Department of Head and Neck Surgery, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, California, United States of America.
authors.forename: Yoo Jin
authors.initials: YJ
authors.lastname: Lee
authors.affiliation: Department of Head and Neck Surgery, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, California, United States of America.
authors.forename: David J
authors.initials: DJ
authors.lastname: Lim
authors.affiliation: Department of Head and Neck Surgery, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, California, United States of America.
authors.forename: Sung K
authors.initials: SK
authors.lastname: Moon
mesh_terms: D000818:Animals; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D004432:Ear, Middle / Q000378:metabolism; D004705:Endocytosis; D005786:Gene Expression Regulation; D020868:Gene Silencing; D005839:Gentamicins / Q000737:chemistry; D006192:Haemophilus Infections / Q000276:immunology* / Q000382:microbiology; D006801:Humans; D007113:Immunity, Innate; D007249:Inflammation / Q000276:immunology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D046529:Microscopy, Electron, Transmission; D053473:Nod2 Signaling Adaptor Protein / Q000378:metabolism*; D010033:Otitis Media / Q000276:immunology* / Q000382:microbiology; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction; D051195:Toll-Like Receptor 2 / Q000378:metabolism; D023083:beta-Defensins / Q000378:metabolism*
chemical_list: C117047:Defb2 protein, mouse; D005839:Gentamicins; D053473:Nod2 Signaling Adaptor Protein; C495945:Nod2 protein, mouse; C493486:Tlr2 protein, mouse; D051195:Toll-Like Receptor 2; D023083:beta-Defensins; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C506255:Ripk2 protein, mouse