BACKGROUND: Naturally occurring antibodies (Nabs) that bind to terminal galactose alpha1,3-galactose carbohydrate structures (Gal) are present in humans and Old World monkeys but are negatively regulated in other mammalian species because they express Gal epitopes on their cell surfaces. A Gal knockout mouse (Gal-/-) model, generated by homologous disruption of alpha1,3-galactosyltransferase gene, is capable of producing natural anti-Gal Abs. METHODS: To study the genetic control of the anti-Gal response, we have generated anti-Gal hybridomas from Gal-/- mice and analyzed VH genes of anti-Gal Abs from naïve animals and from mice stimulated by rat heterotopic heart transplantation. RESULTS: Six immunoglobulin (Ig)M anti-Gal hybridomas derived from naïve Gal-/- mice exhibited anti-Gal binding activity with some cross-reactivity to related carbohydrate structures. These naïve anti-Gal Abs used five different VH genes in a germline configuration. Anti-Gal IgM hybridomas isolated after a rat heterotopic heart xenograft (4 days) utilized germline VH gene segments from the VH7183 family and exhibited less cross-reactivity. In contrast to mice 4 days after xenograft, we have predominantly isolated IgG anti-Gal hybridomas from mice 21 days after rat heterotopic heart xenografts, indicating an isotype switch. Nine of the IgG anti-Gal hybridomas secreted IgG3 subclass and one produced IgG1. Sequence analysis of the VH gene usage from the induced anti-Gal IgG antibodies demonstrated a restricted gene utilization (VHJ606-V14A). CONCLUSION: Our results demonstrate that the anti-Gal response in naïve Gal-/- mice is encoded by multiple germline progenitors. In response to a xenograft, the induced anti-Gal Abs exhibited a restricted gene usage and somatic mutations, indicating a positive selection.
BACKGROUND: Naturally occurring antibodies (Nabs) that bind to terminal galactose alpha1,3-galactose carbohydrate structures (Gal) are present in humans and Old World monkeys but are negatively regulated in other mammalian species because they express Gal epitopes on their cell surfaces. A Gal knockout mouse (Gal-/-) model, generated by homologous disruption of alpha1,3-galactosyltransferase gene, is capable of producing natural anti-Gal Abs. METHODS: To study the genetic control of the anti-Gal response, we have generated anti-Gal hybridomas from Gal-/- mice and analyzed VH genes of anti-Gal Abs from naïve animals and from mice stimulated by rat heterotopic heart transplantation. RESULTS: Six immunoglobulin (Ig)M anti-Gal hybridomas derived from naïve Gal-/- mice exhibited anti-Gal binding activity with some cross-reactivity to related carbohydrate structures. These naïve anti-Gal Abs used five different VH genes in a germline configuration. Anti-Gal IgM hybridomas isolated after a rat heterotopic heart xenograft (4 days) utilized germline VH gene segments from the VH7183 family and exhibited less cross-reactivity. In contrast to mice 4 days after xenograft, we have predominantly isolated IgG anti-Gal hybridomas from mice 21 days after rat heterotopic heart xenografts, indicating an isotype switch. Nine of the IgG anti-Gal hybridomas secreted IgG3 subclass and one produced IgG1. Sequence analysis of the VH gene usage from the induced anti-Gal IgG antibodies demonstrated a restricted gene utilization (VHJ606-V14A). CONCLUSION: Our results demonstrate that the anti-Gal response in naïve Gal-/- mice is encoded by multiple germline progenitors. In response to a xenograft, the induced anti-Gal Abs exhibited a restricted gene usage and somatic mutations, indicating a positive selection.
Authors: Annette Kleihauer; Clare R Gregory; Dominic C Borie; Andrew E Kyles; Irina Shulkin; Insiyyah Patanwala; Joanne Zahorsky-Reeves; Vaughn A Starnes; Yoko Mullen; Ivan T Todorov; Mary Kearns-Jonker Journal: Immunology Date: 2005-09 Impact factor: 7.397
Authors: Mary Kearns-Jonker; Natasha Barteneva; Robert Mencel; Namath Hussain; Irina Shulkin; Alan Xu; Margaret Yew; Donald V Cramer Journal: BMC Immunol Date: 2007-03-12 Impact factor: 3.615