title: TLR4 and TLR5 on corneal macrophages regulate Pseudomonas aeruginosa keratitis by signaling through MyD88-dependent and -independent pathways.
abstract: Pseudomonas aeruginosa is a major cause of blindness and visual impairment in the United States and worldwide. Using a murine model of keratitis in which abraded corneas are infected with P. aeruginosa parent and ΔfliC (aflagellar) strains 19660 and PAO1, we found that F4/80(+) macrophages were the predominant cell type in the cornea expressing TLR2, TLR4, and TLR5. Depletion of macrophages and dendritic cells using transgenic Mafia mice, in which Fas ligand is selectively activated in these cells, resulted in diminished cytokine production and cellular infiltration to the corneal stroma and unimpaired bacterial growth. TLR4(-/-) mice showed a similar phenotype postinfection with ΔfliC strains, whereas TLR4/5(-/-) mice were susceptible to corneal infection with parent strains. Bone marrow-derived macrophages stimulated with ΔfliC bacteria induced Toll/IL-1R intracellular domain (TIR)-containing adaptor inducing IFN-β (TRIF)-dependent phosphorylation of IFN regulatory factor 3 in addition to TIR-containing adaptor protein/MyD88-dependent phosphorylation of IκB and nuclear translocation of the p65 subunit of NFκB. Furthermore, TRIF(-/-) mice showed a similar phenotype as TLR4(-/-) mice in regulating only ΔfliC bacteria, whereas MyD88(-/-) mice were unable to clear parent or ΔfliC bacteria. Finally, IL-1R1(-/-) and IL-1α/β(-/-) mice were highly susceptible to infection. Taken together, these findings indicate that P. aeruginosa activates TLR4/5 on resident corneal macrophages, which signal through TRIF and TIR-containing adaptor protein/MyD88 pathways, leading to NF-κB translocation to the nucleus, transcription of CXCL1 and other CXC chemokines, recruitment of neutrophils to the corneal stroma, and subsequent bacterial killing and tissue damage. IL-1α and IL-1β are also produced, which activate an IL-1R1/MyD88-positive feedback loop in macrophages and IL-1R on other resident cells in the cornea.
authors.affiliation: Department of Ophthalmology and Visual Sciences, Case Western Reserve University, Cleveland, OH 44106, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Sun
authors.forename: Mausita
authors.initials: M
authors.lastname: Karmakar
authors.forename: Sanhita
authors.initials: S
authors.lastname: Roy
authors.forename: Raniyah T
authors.initials: RT
authors.lastname: Ramadan
authors.forename: Susan R
authors.initials: SR
authors.lastname: Williams
authors.forename: Scott
authors.initials: S
authors.lastname: Howell
authors.forename: Carey L
authors.initials: CL
authors.lastname: Shive
authors.forename: Yiping
authors.initials: Y
authors.lastname: Han
authors.forename: Charles M
authors.initials: CM
authors.lastname: Stopford
authors.forename: Arne
authors.initials: A
authors.lastname: Rietsch
authors.forename: Eric
authors.initials: E
authors.lastname: Pearlman
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000276:immunology / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002469:Cell Separation; D003315:Cornea / Q000276:immunology / Q000378:metabolism / Q000382:microbiology; D004195:Disease Models, Animal; D005434:Flow Cytometry; D007150:Immunohistochemistry; D007634:Keratitis / Q000276:immunology* / Q000382:microbiology; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D053594:Myeloid Differentiation Factor 88 / Q000276:immunology* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011552:Pseudomonas Infections / Q000276:immunology* / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000276:immunology* / Q000378:metabolism; D051198:Toll-Like Receptor 5 / Q000276:immunology* / Q000378:metabolism
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D053594:Myeloid Differentiation Factor 88; D051197:Toll-Like Receptor 4; D051198:Toll-Like Receptor 5; C507780:Myd88 protein, mouse; C476286:Ticam1 protein, mouse; C493487:Tlr4 protein, mouse
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