title: Antibody repertoires in humanized NOD-scid-IL2Rγ(null) mice and human B cells reveals human-like diversification and tolerance checkpoints in the mouse.
abstract: Immunodeficient mice reconstituted with human hematopoietic stem cells enable the in vivo study of human hematopoiesis. In particular, NOD-scid-IL2Rγ(null) engrafted mice have been shown to have reasonable levels of T and B cell repopulation and can mount T-cell dependent responses; however, antigen-specific B-cell responses in this model are generally poor. We explored whether developmental defects in the immunoglobulin gene repertoire might be partly responsible for the low level of antibody responses in this model. Roche 454 sequencing was used to obtain over 685,000 reads from cDNA encoding immunoglobulin heavy (IGH) and light (IGK and IGL) genes isolated from immature, naïve, or total splenic B cells in engrafted NOD-scid-IL2Rγ(null) mice, and compared with over 940,000 reads from peripheral B cells of two healthy volunteers. We find that while naïve B-cell repertoires in humanized mice are chiefly indistinguishable from those in human blood B cells, and display highly correlated patterns of immunoglobulin gene segment use, the complementarity-determining region H3 (CDR-H3) repertoires are nevertheless extremely diverse and are specific for each individual. Despite this diversity, preferential D(H)-J(H) pairings repeatedly occur within the CDR-H3 interval that are strikingly similar across all repertoires examined, implying a genetic constraint imposed on repertoire generation. Moreover, CDR-H3 length, charged amino-acid content, and hydropathy are indistinguishable between humans and humanized mice, with no evidence of global autoimmune signatures. Importantly, however, a statistically greater usage of the inherently autoreactive IGHV4-34 and IGKV4-1 genes was observed in the newly formed immature B cells relative to naïve B or total splenic B cells in the humanized mice, a finding consistent with the deletion of autoreactive B cells in humans. Overall, our results provide evidence that key features of the primary repertoire are shaped by genetic factors intrinsic to human B cells and are principally unaltered by differences between mouse and human stromal microenvironments.
authors.affiliation: Section of Molecular Genetics and Microbiology, University of Texas at Austin, Austin, Texas, United States of America.
authors.forename: Gregory C
authors.initials: GC
authors.lastname: Ippolito
authors.forename: Kam Hon
authors.initials: KH
authors.lastname: Hoi
authors.forename: Sai T
authors.initials: ST
authors.lastname: Reddy
authors.forename: Sean M
authors.initials: SM
authors.lastname: Carroll
authors.forename: Xin
authors.initials: X
authors.lastname: Ge
authors.forename: Tobias
authors.initials: T
authors.lastname: Rogosch
authors.forename: Michael
authors.initials: M
authors.lastname: Zemlin
authors.forename: Leonard D
authors.initials: LD
authors.lastname: Shultz
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Ellington
authors.forename: Carla L
authors.initials: CL
authors.lastname: Vandenberg
authors.forename: George
authors.initials: G
authors.lastname: Georgiou
mesh_terms: D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000235:genetics* / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology*; D001483:Base Sequence; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D014644:Genetic Variation; D006410:Hematopoiesis / Q000276:immunology*; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D050440:Immunoglobulin Subunits / Q000235:genetics; D053631:Interleukin Receptor Common gamma Subunit / Q000235:genetics; D051379:Mice; D016688:Mice, Inbred NOD / Q000235:genetics / Q000276:immunology*; D016513:Mice, SCID / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA; D018709:Statistics, Nonparametric
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D017931:DNA Primers; D018076:DNA, Complementary; D005456:Fluorescent Dyes; C507942:Il2rg protein, mouse; D050440:Immunoglobulin Subunits; D053631:Interleukin Receptor Common gamma Subunit