MOUSE MODEL OF IGA DEFICIENCY USING GENE TARGETING

IgA缺乏症是人类最常见的体液性免疫缺陷，

IgA deficiency is the most common humoral immunodeficiency in humans, affecting approximately 1 in 600 people. This deficiency results in the virtual complete absence of detectable IgA in serum or mucosal secretions and is associated with an increased incidence of infections and autoimmune diseases. No small animal model currently exists for selective IgA deficiency. The immediate objective of this project is to develop a mouse model of IgA deficiency using gene targeting to functionally delete the immunoglobulin heavy chain (Igh) IgA locus from mice. The more long-term goal of this project is to gain insight into IgA B cell ontogeny and the role of IgA in systemic and mucosal immune responses. Such knowledge will likely enhance our understanding of humoral immunodeficiencies, particularly IgA deficiency, and should enhance our ability to design better oral and systemic vaccines against various pathogens, such as HIV. In addition, the production of an IgA deficient mouse by gene targeting will further expand the experience using gene targeting to develop animal models of human diseases. The immediate objective of this project will be achieved as follows. Embryonic stem cells derived from the murine strain 129 (agouti coat color) will have the Igh alpha constant region exons deleted using homologous recombination techniques. To produce recombinant mice containing this deletion, the mutated embryonic stem cells will be introduced into C57BL/6 (black coat color) blastocysts which will then be inserted into foster mothers. Chimeric male offspring with the largest contribution from the mutated embryonic stem cells, as determined by extent of coat color chimerism, will be bred with C57BL/6 female mice. Germline transmission of the mutation into the F1 generation will be assessed by Southern blotting of offspring tail DNA. Heterozygote offspring will be bred to obtain homozygous mice lacking the Igh IgA locus. Confirmation that mutation of the Igh IgA locus has resulted in loss of IgA expression will be obtained by measuring serum IgA levels in the mutant mice with an IgA isotype-specific ELISA and by measuring expression of surface IgA on Peyer's patch or spleen B cells with flow cytometry. The IgA deficient mice will be bred to establish a colony which will then be available for future studies intended to evaluate B cell ontogeny and immunoglobulin isotype expression in mucosal tissues and the role of IgA in mucosal immune responses against microbial pathogens.