title: Receptor cross-linking on human plasmacytoid dendritic cells leads to the regulation of IFN-alpha production.
abstract: Plasmacytoid dendritic cells (PDC) are the natural type I IFN-producing cells that produce large amounts of IFN-alpha in response to viral stimulation. During attempts to isolate PDC from human PBMC, we observed that cross-linking a variety of cell surface receptors, including blood DC Ag (BDCA)-2, BDCA-4, CD4, or CD123 with Abs and immunobeads on PDC leads to inhibition of IFN-alpha production in response to HSV. To understand the mechanisms involved, a number of parameters were investigated. Cross-linking did not inhibit endocytosis of soluble Ag by PDC. Flow cytometry for annexin V and activated caspase-3 indicated that PDC are not undergoing apoptosis after receptor cross-linking. Cross-linking of CD123, but not the other receptors, caused the up-regulation of costimulatory molecules CD80 and CD86, as well as the down-regulation of CD62L, indicating PDC maturation. Thus, anti-CD123 Ab may be acting similar to the natural ligand, IL-3. Anti-phosphotyrosine Ab, as well as Ab to the IFN regulatory factor, IRF-7, was used in intracellular flow cytometry to elucidate the signaling pathways involved. Tyrosine phosphorylation occurred after cross-linking BDCA-2 and BDCA-4, but not CD4. Cross-linking did not affect IRF-7 levels in PDC, however, cross-linking BDCA-2, BDCA-4, and CD4, but not CD123, inhibited the ability of IRF-7 to translocate to the nucleus. Taken together, these results suggest that cross-linking BDCA-2, BDCA-4, and CD4 on PDC regulates IFN-alpha production at the level of IRF-7, while the decrease in IFN-alpha production after CD123 cross-linking is due to stimulation of the IL-3R and induction of PDC maturation.
authors.affiliation: New Jersey Medical School, University of Medicine and Dentistry of New Jersey, 185 South Orange Avenue, Newark, NJ 07103, USA.
authors.forename: Stacey L
authors.initials: SL
authors.lastname: Fanning
authors.forename: Thaddeus C
authors.initials: TC
authors.lastname: George
authors.forename: Di
authors.initials: D
authors.lastname: Feng
authors.forename: Steven B
authors.initials: SB
authors.lastname: Feldman
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Megjugorac
authors.forename: Alexander G
authors.initials: AG
authors.lastname: Izaguirre
authors.forename: Patricia
authors.initials: P
authors.lastname: Fitzgerald-Bocarsly
mesh_terms: D000906:Antibodies / Q000494:pharmacology; D000954:Antigens, Surface / Q000187:drug effects; D017209:Apoptosis; D015704:CD4 Antigens / Q000187:drug effects; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology*; D004705:Endocytosis / Q000187:drug effects; D018259:Herpesvirus 1, Human / Q000502:physiology*; D006801:Humans; D050839:Interferon Regulatory Factor-7 / Q000378:metabolism; D016898:Interferon-alpha / Q000096:biosynthesis* / Q000235:genetics; D053650:Interleukin-3 Receptor alpha Subunit / Q000037:antagonists & inhibitors / Q000187:drug effects; D037181:Lectins, C-Type / Q000037:antagonists & inhibitors / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000187:drug effects; D008863:Microspheres; D010766:Phosphorylation; D011971:Receptors, Immunologic / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D016185:Receptors, Interleukin-3 / Q000037:antagonists & inhibitors / Q000187:drug effects; D014443:Tyrosine / Q000378:metabolism
chemical_list: D000906:Antibodies; D000954:Antigens, Surface; D015704:CD4 Antigens; C443299:CLEC4C protein, human; D003432:Cross-Linking Reagents; C115022:IL3RA protein, human; D050839:Interferon Regulatory Factor-7; D016898:Interferon-alpha; D053650:Interleukin-3 Receptor alpha Subunit; D037181:Lectins, C-Type; D008562:Membrane Glycoproteins; D011971:Receptors, Immunologic; D016185:Receptors, Interleukin-3; C478196:blood dendritic cell antigen 4, human; D014443:Tyrosine