title: Human trophoblast-derived exosomal fibronectin induces pro-inflammatory IL-1β production by macrophages.
abstract: PROBLEM  Our previous studies demonstrated that trophoblast-derived exosomes induced synthesis and release of pro-inflammatory cytokines, including interleukin-1β (IL-1β) by macrophages. The objective of this study was to characterize the mechanism and receptors associated with this induction. METHOD OF STUDY  Exosomes were isolated from Sw71 trophoblast-conditioned media by ultrafiltration and ultracentrifugation. Using macrophages isolated from normal donors, cytochalasin D was used to block exosome uptake. Induction of IL-1β mRNA was investigated by qRT-PCR, pro-IL-1β protein by western immunoblotting, and mature IL-1β release by ELISA. RGD peptides were used to block fibronectin binding by macrophage α5β1 integrin. RESULTS  Uptake of exosomes by macrophages was completely blocked by pre-treatment with cytochalasin D. Although induction of some cytokines (such as C4A and CCL11) requires uptake, induction of IL-1β occurred without exosome internalization. Cytochalasin D treatment did not inhibit exosome-mediated induction of IL-1β mRNA, production of the pro-protein, or release of mature IL-1β. Blocking of fibronectin binding using RGD peptides demonstrated the abrogation of exosome-mediated IL-1β production. CONCLUSION  Although trophoblast-derived exosomes have been demonstrated to induce IL-1β, this is the first demonstration of IL-1β induction by exosome-associated fibronectin. Based on this pro-inflammatory role of exosome-associated fibronectin, it may represent an important general immunoregulatory mechanism.
authors.affiliation: Department of Microbiology & Immunology, University of Louisville School of Medicine, KY, USA.
authors.forename: Safinur
authors.initials: S
authors.lastname: Atay
authors.forename: Cicek
authors.initials: C
authors.lastname: Gercel-Taylor
authors.forename: Douglas D
authors.initials: DD
authors.lastname: Taylor
mesh_terms: D002460:Cell Line; D054413:Chemokine CCL11 / Q000096:biosynthesis; D017077:Culture Media, Conditioned / Q000737:chemistry; D015638:Cytochalasin D / Q000494:pharmacology; D004705:Endocytosis / Q000187:drug effects* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D055354:Exosomes / Q000276:immunology / Q000378:metabolism*; D005260:Female; D005353:Fibronectins / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D007249:Inflammation / Q000276:immunology / Q000378:metabolism; D053583:Interleukin-1beta / Q000032:analysis / Q000096:biosynthesis*; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D011247:Pregnancy; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D060888:Real-Time Polymerase Chain Reaction; D011971:Receptors, Immunologic / Q000037:antagonists & inhibitors; D018000:Receptors, Peptide / Q000037:antagonists & inhibitors; D014327:Trophoblasts / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D014461:Ultracentrifugation
chemical_list: D054413:Chemokine CCL11; D017077:Culture Media, Conditioned; D005353:Fibronectins; D053583:Interleukin-1beta; D009842:Oligopeptides; D012333:RNA, Messenger; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; C051382:arginyl-glycyl-aspartic acid directed cell adhesion receptor; D015638:Cytochalasin D; C047981:arginyl-glycyl-aspartic acid