title: Molecular analysis of the heavy chain of antibodies that recognize the capsular polysaccharide of Neisseria meningitidis in hu-PBMC reconstituted SCID mice and in the immunized human donor.
abstract: The severe combined immunodeficient (SCID) mouse model, engrafted with human peripheral blood mononuclear cells (hu-PBMC) has proven to be useful in studying the human immune response. A major limitation of the hu-PBMC-SCID model has been the failure to consistently demonstrate a primary human immune response. Previously we developed a hu-PBMC-SCID mouse model in which we addressed both issues of adequate human lymphocyte engraftment and impaired differentiation. We demonstrated that a primary human immune response to the T-independent (TI-2) meningococcal group C capsular polysaccharide (MCPS) can be obtained in hu-PBMC-SCID mice by the administration of human cytokines. In this study we compared the V(H) sequence of the MCPS response generated by B cells derived from a volunteer in the SCID mouse model to those generated by the donors' B cells in vivo. Human peripheral blood mononuclear cells were recovered from MCPS immunized hu-PBMC-SCID mice and immunized donor. B cells with specificity for MCPS were isolated from these cell preparations using an anti-idiotypic monoclonal antibody which mimics MCPS. Immunoglobulin mRNA was isolated from single cells, amplified by the polymerase chain reaction, cloned and sequenced. We analysed a total of 15 V(H) regions from B cells obtained from SCID mice and a total of 13 V(H) regions from B cells obtained from the immunized donor. The response differed between SCID and in vivo cells, when studied at the genetric level. V, D and J gene usage was markedly different, however canonical structures of the hypervariable loops were conserved. The complementary determining region 3 (CDR3) varied, such that SCID-derived sequences encoded longer CDR3 s than those of the donor. However all CDR3 s were rich in hydrophobic amino acids, most notably tyrosine and tryptophan, a characteristic common to many carbohydrate binding antibodies.
authors.affiliation: Department of Medicine, Medical College of Ohio, Toledo 43614, USA.
authors.forename: S L
authors.initials: SL
authors.lastname: Smithson
authors.forename: N
authors.initials: N
authors.lastname: Srivastava
authors.forename: W A
authors.initials: WA
authors.lastname: Hutchins
authors.forename: M A
authors.initials: MA
authors.lastname: Westerink
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000235:genetics / Q000276:immunology*; D000942:Antigens, Bacterial / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D006801:Humans; D007114:Immunization; D007143:Immunoglobulin Heavy Chains / Q000235:genetics* / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000276:immunology* / Q000637:transplantation; D051379:Mice; D016513:Mice, SCID; D008969:Molecular Sequence Data; D009345:Neisseria meningitidis / Q000276:immunology*; D011135:Polysaccharides, Bacterial / Q000276:immunology*; D016415:Sequence Alignment
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D007143:Immunoglobulin Heavy Chains; D011135:Polysaccharides, Bacterial