title: Impaired growth and elevated fas receptor expression in PIGA(+) stem cells in primary paroxysmal nocturnal hemoglobinuria.
abstract: The genetic defect underlying paroxysmal nocturnal hemoglobinuria (PNH) has been shown to reside in PIGA, a gene that encodes an element required for the first step in glycophosphatidylinositol anchor assembly. Why PIGA-mutated cells are able to expand in PNH marrow, however, is as yet unclear. To address this question, we compared the growth of affected CD59(-)CD34(+) and unaffected CD59(+)CD34(+) cells from patients with that of normal CD59(+)CD34(+) cells in liquid culture. One hundred FACS-sorted cells were added per well into microtiter plates, and after 11 days at 37 degrees C the progeny were counted and were analyzed for their differentiation pattern. We found that CD59(-)CD34(+) cells from PNH patients proliferated to levels approaching those of normal cells, but that CD59(+)CD34(+) cells from the patients gave rise to 20- to 140-fold fewer cells. Prior to sorting, the patients' CD59(-) and CD59(+)CD34(+) cells were equivalent with respect to early differentiation markers, and following culture, the CD45 differentiation patterns were identical to those of control CD34(+) cells. Further analyses of the unsorted CD59(+)CD34(+) population, however, showed elevated levels of Fas receptor. Addition of agonist anti-Fas mAb to cultures reduced the CD59(+)CD34(+) cell yield by up to 78% but had a minimal effect on the CD59(-)CD34(+) cells, whereas antagonist anti-Fas mAb enhanced the yield by up to 250%. These results suggest that expansion of PIGA-mutated cells in PNH marrow is due to a growth defect in nonmutated cells, and that greater susceptibility to apoptosis is one factor involved in the growth impairment.
authors.affiliation: Institute of Pathology, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: R
authors.initials: R
authors.lastname: Chen
authors.forename: S
authors.initials: S
authors.lastname: Nagarajan
authors.forename: G M
authors.initials: GM
authors.lastname: Prince
authors.forename: U
authors.initials: U
authors.lastname: Maheshwari
authors.forename: L W
authors.initials: LW
authors.lastname: Terstappen
authors.forename: D R
authors.initials: DR
authors.lastname: Kaplan
authors.forename: S L
authors.initials: SL
authors.lastname: Gerson
authors.forename: J M
authors.initials: JM
authors.lastname: Albert
authors.forename: D E
authors.initials: DE
authors.lastname: Dunn
authors.forename: H M
authors.initials: HM
authors.lastname: Lazarus
authors.forename: M E
authors.initials: ME
authors.lastname: Medof
mesh_terms: D018952:Antigens, CD34; D001853:Bone Marrow / Q000601:surgery; D001854:Bone Marrow Cells / Q000166:cytology / Q000502:physiology*; D018957:CD59 Antigens; D002454:Cell Differentiation; D002469:Cell Separation; D016238:Cytapheresis; D017261:Glycosylphosphatidylinositols / Q000096:biosynthesis; D006457:Hemoglobinuria, Paroxysmal / Q000209:etiology / Q000235:genetics*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics*; D010641:Phenotype; D013234:Stem Cells / Q000166:cytology / Q000502:physiology*; D019014:fas Receptor / Q000378:metabolism*
chemical_list: D018952:Antigens, CD34; D018957:CD59 Antigens; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D019014:fas Receptor; C080086:phosphatidylinositol glycan-class A protein