title: Technical advance: live-imaging analysis of human dendritic cell migrating behavior under the influence of immune-stimulating reagents in an organotypic model of lung.
abstract: This manuscript describes technical advances allowing manipulation and quantitative analyses of human DC migratory behavior in lung epithelial tissue. DCs are hematopoietic cells essential for the maintenance of tissue homeostasis and the induction of tissue-specific immune responses. Important functions include cytokine production and migration in response to infection for the induction of proper immune responses. To design appropriate strategies to exploit human DC functional properties in lung tissue for the purpose of clinical evaluation, e.g., candidate vaccination and immunotherapy strategies, we have developed a live-imaging assay based on our previously described organotypic model of the human lung. This assay allows provocations and subsequent quantitative investigations of DC functional properties under conditions mimicking morphological and functional features of the in vivo parental tissue. We present protocols to set up and prepare tissue models for 4D (x, y, z, time) fluorescence-imaging analysis that allow spatial and temporal studies of human DCs in live epithelial tissue, followed by flow cytometry analysis of DCs retrieved from digested tissue models. This model system can be useful for elucidating incompletely defined pathways controlling DC functional responses to infection and inflammation in lung epithelial tissue, as well as the efficacy of locally administered candidate interventions.
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and.
authors.forename: Anh Thu
authors.initials: AT
authors.lastname: Nguyen Hoang
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and.
authors.forename: Puran
authors.initials: P
authors.lastname: Chen
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and.
authors.forename: Sofia
authors.initials: S
authors.lastname: Björnfot
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and.
authors.forename: Kari
authors.initials: K
authors.lastname: Högstrand
authors.affiliation: Center for Biosciences, Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden; and.
authors.forename: John G
authors.initials: JG
authors.lastname: Lock
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and.
authors.forename: Alf
authors.initials: A
authors.lastname: Grandien
authors.affiliation: Centre for Immunology and Infection, Hull York Medical School and Department of Biology, University of York, United Kingdom.
authors.forename: Mark
authors.initials: M
authors.lastname: Coles
authors.affiliation: Center for Infectious Medicine, Department of Medicine, and mattias.svensson@ki.se.
authors.forename: Mattias
authors.initials: M
authors.lastname: Svensson
mesh_terms: D000276:Adjuvants, Immunologic / Q000494:pharmacology*; D002450:Cell Communication; D018929:Cell Culture Techniques; D002460:Cell Line; D002465:Cell Movement; D018932:Chemokine CCL2 / Q000494:pharmacology; D018920:Coculture Techniques; D017077:Culture Media, Conditioned; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D004847:Epithelial Cells / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D017930:Genes, Reporter; D006801:Humans; D007249:Inflammation / Q000139:chemically induced / Q000276:immunology; D055666:Lipopeptides / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008168:Lung / Q000166:cytology / Q000276:immunology*; D018448:Models, Immunological; D009000:Monocytes / Q000166:cytology; D011994:Recombinant Proteins / Q000494:pharmacology; D059008:Time-Lapse Imaging / Q000379:methods*
chemical_list: D000276:Adjuvants, Immunologic; C111835:CCL2 protein, human; D018932:Chemokine CCL2; D017077:Culture Media, Conditioned; D016207:Cytokines; D055666:Lipopeptides; D008070:Lipopolysaccharides; C508075:Pam(3)CSK(4) peptide; D011994:Recombinant Proteins
keywords: 3D-tissue models; Toll-like receptor agonists; epithelial tissue; inflammation