Phenotypic and functional characterization of a mouse model of targeted Pig-a deletion in hematopoietic cells.
BACKGROUND
Somatic mutation in the X-linked phosphatidylinositol glycan class A gene (PIG-A) causes glycosyl phosphatidylinositol anchor deficiency in human patients with paroxysmal nocturnal hemoglobinuria.


METHODS
We produced an animal model of paroxysmal nocturnal hemoglobinuria by conditional Pig-a gene inactivation (Pig-a(-/-)) in hematopoietic cells; mice carrying two lox sites flanking exon 6 of the Pig-a gene were bred with mice carrying the transgene Cre-recombinase under the human c-fes promoter. We characterized the phenotypic and functional properties of glycosyl phosphatidylinositol-deficient and glycosyl phosphatidylinositol-normal hematopoietic cells from these Pig-a(-/-) mice using gene expression microarray, flow cytometry, bone marrow transplantation, spectratyping, and immunoblotting.


RESULTS
In comparison to glycosyl phosphatidylinositol-normal bone marrow cells, glycosyl phosphatidylinositol-deficient bone marrow cells from the same Pig-a(-/-) animals showed up-regulation of the expression of immune function genes and contained a significantly higher proportion of CD8 T cells. Both characteristics were maintained when glycosyl phosphatidylinositol-deficient cells were transplanted into lethally-irradiated recipients. Glycosyl phosphatidylinositol-deficient T cells were inactive, showed pronounced Vbeta5.1/5.2 skewing, had fewer gamma-interferon-producing cells after lectin stimulation, and contained fewer CD4(+)CD25(+)FoxP3(+) regulatory T cells. However, the levels of T-cell receptor signaling proteins from glycosyl phosphatidylinositol-deficient cells were normal relative to glycosyl phosphatidylinositol-normal cells from wild type animals, and cells were capable of inducing target cell apoptosis in vitro.


CONCLUSIONS
Deletion of the Pig-a gene in hematopoietic cells does not cause frank marrow failure but leads to the appearance of clonally-restricted, inactive yet functionally competent CD8 T cells.
Visconte
Valeria
V
Hematology Branch, NHLBI, National Institutes of Health, Bethesda, MD 20892-1202 USA. viscontev@mail.nih.gov
Raghavachari
Nalini
N
Liu
Delong
D
Keyvanfar
Keyvan
K
Desierto
Marie J
MJ
Chen
Jichun
J
Young
Neal S
NS
D000818:Animals; D001854:Bone Marrow Cells / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D004195:Disease Models, Animal; D005260:Female; D017353:Gene Deletion; D006457:Hemoglobinuria, Paroxysmal / Q000235:genetics*; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D010641:Phenotype; D015854:Up-Regulation
D008565:Membrane Proteins; C080086:phosphatidylinositol glycan-class A protein
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