title: A knock-out model of paroxysmal nocturnal hemoglobinuria: Pig-a(-) hematopoiesis is reconstituted following intercellular transfer of GPI-anchored proteins.
abstract: We created a "knockout" embryonic stem cell via targeted disruption of the phosphatidylinositol glycan class A (Pig-a) gene, resulting in loss of expression of cell surface glycosyl phosphatidylinositol-anchored proteins and reproducing the mutant phenotype of the human disease paroxysmal nocturnal hemoglobinuria. Morphogenesis of Pig-a- embryoid bodies (EB) in vitro was grossly aberrant and, unlike EB derived from normal embryonic stem cells, Pig-A EB produced no secondary hematopoietic colonies. Chimeric EB composed of control plus Pig-A- cells, however, appeared normal, and hematopoiesis from knock-out cells was reconstituted. Transfer in situ of glycosyl phosphatidylinositol-anchored proteins from normal to knock-out cells was demonstrated by two-color fluorescent analysis, suggesting a possible mechanism for these functional effects. Hematopoietic cells with mutated PIG-A genes in humans with paroxysmal nocturnal hemoglobinuria may be subject to comparable pathophysiologic processes and amenable to similar therapeutic protein transfer.
authors: Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892-1652, USA.
authors: D E
authors: DE
authors: Dunn
authors: J
authors: J
authors: Yu
authors: S
authors: S
authors: Nagarajan
authors: M
authors: M
authors: Devetten
authors: F F
authors: FF
authors: Weichold
authors: M E
authors: ME
authors: Medof
authors: N S
authors: NS
authors: Young
authors: J M
authors: JM
authors: Liu
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D018920:Coculture Techniques; D004622:Embryo, Mammalian; D017261:Glycosylphosphatidylinositols / Q000235:genetics* / Q000378:metabolism*; D006457:Hemoglobinuria, Paroxysmal / Q000235:genetics* / Q000473:pathology*; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D016254:Mutagenesis, Insertional; D013234:Stem Cells / Q000166:cytology*
chemical_list: D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins