title: T-cell hybridoma specific for myelin oligodendrocyte glycoprotein-35-55 peptide produced from HLA-DRB1*1501-transgenic mice.
abstract: The goal of this study was to establish an unlimited and standardized source of humanized myelin peptide-specific T cells for in vitro testing of biological function. Thus, we perpetuated myelin oligodendrocyte glycoprotein (MOG)-35-55 peptide-specific T cells obtained from immunized HLA-DRB1*1501-transgenic (Tg) mice by somatic fusions with BW5147 thymoma cells or BW5147 T-cell receptor (TCR) alpha(-)beta(-) variant (BW5147 variant) cells. The resulting T-cell hybridomas responded strongly to both mouse MOG-35-55 (42S) and human MOG-35-55 peptide (42P), regardless of which peptide was used for initial immunization, and were DRB1*1501 restricted. The MOG-35-55-reactive T-cell hybridomas were CD3(+)CD4(+)CD8(-) and expressed intracellular Th1 cytokines upon concanavalin A stimulation. Clones from either human MOG-35-55- or mouse MOG-35-55-selected hybridomas uniquely expressed the TCR BV8 gene in combination with AV17 and AV11 genes. V gene analyses confirmed the expression of TCR AV1, AV11, AV16, BV1, and BV5 gene segments in the widely used fusion partner BW5147 and demonstrated deletion of TCR AV1, AV11, and BV1 in the BW5147 variant. T-cell hybridomas were positively stained with anti-TCR beta-chain antibody on the cell surface, whereas neither BW5147 nor its variant had positive TCR surface expression. For functional application, we found that a monomeric form of the human HLA-DR2-derived recombinant T-cell receptor ligand (RTL) covalently linked to human MOG-35-55 peptide specifically inhibited proliferation of a hybridoma clone selected with human MOG-35-55 but not a different hybridoma clone selected with myelin basic protein. The RTL-induced inhibition in vitro of the human MOG-35-55 peptide-specific hybridoma reflected the ability of the RTL to inhibit experimental autoimmune encephalomyelitis induced by human MOG-35-55 peptide in HLA-DR2 transgenic mice. Thus, the MOG-35-55 peptide-specific T-cell hybridoma from DR2-Tg mice represents a novel humanized T-cell reagent useful for standardized biological screening of both DR2-restricted stimulation and RTL-dependent inhibition of response to human MOG-35-55 peptide.
authors.affiliation: Neuroimmunology Research and Tykeson Multiple Sclerosis Research Laboratory, Veterans Affairs Medical Center, and Oregon Health and Science University, Portland, Oregon 97239, USA. chouy@ohsu.edu
authors.forename: Yuan K
authors.initials: YK
authors.lastname: Chou
authors.forename: Nicole
authors.initials: N
authors.lastname: Culbertson
authors.forename: Cathleen
authors.initials: C
authors.lastname: Rich
authors.forename: Dorian
authors.initials: D
authors.lastname: LaTocha
authors.forename: Abigail C
authors.initials: AC
authors.lastname: Buenafe
authors.forename: Jianya
authors.initials: J
authors.lastname: Huan
authors.forename: Jason
authors.initials: J
authors.lastname: Link
authors.forename: J M
authors.initials: JM
authors.lastname: Wands
authors.forename: Willi K
authors.initials: WK
authors.lastname: Born
authors.forename: Halina
authors.initials: H
authors.lastname: Offner
authors.forename: Dennis N
authors.initials: DN
authors.lastname: Bourdette
authors.forename: Gregory G
authors.initials: GG
authors.lastname: Burrows
authors.forename: Arthur A
authors.initials: AA
authors.lastname: Vandenbark
mesh_terms: D000818:Animals; D000917:Antibody Formation / Q000502:physiology; D015139:Blotting, Southern / Q000379:methods; D045744:Cell Line, Tumor; D018920:Coculture Techniques / Q000379:methods; D004306:Dose-Response Relationship, Immunologic; D005434:Flow Cytometry / Q000379:methods; D006023:Glycoproteins / Q000276:immunology*; D006684:HLA-DR Antigens / Q000235:genetics / Q000378:metabolism*; D059811:HLA-DRB1 Chains; D006801:Humans; D006825:Hybridomas / Q000378:metabolism*; D007376:Interleukin-2 / Q000378:metabolism; D051379:Mice; D008815:Mice, Inbred Strains; D008822:Mice, Transgenic; D063308:Myelin-Oligodendrocyte Glycoprotein; D010446:Peptide Fragments / Q000276:immunology*; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D037182:T-Cell Antigen Receptor Specificity / Q000502:physiology; D013601:T-Lymphocytes / Q000378:metabolism*; D013997:Time Factors
chemical_list: D006023:Glycoproteins; D006684:HLA-DR Antigens; D059811:HLA-DRB1 Chains; D007376:Interleukin-2; D063308:Myelin-Oligodendrocyte Glycoprotein; D010446:Peptide Fragments; D012333:RNA, Messenger; C445081:myelin oligodendrocyte glycoprotein (35-55)