Impaired growth and elevated fas receptor expression in PIGA(+) stem cells in primary paroxysmal nocturnal hemoglobinuria.
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.
Chen
R
R
Institute of Pathology, Case Western Reserve University, Cleveland, Ohio, USA.
Nagarajan
S
S
Prince
G M
GM
Maheshwari
U
U
Terstappen
L W
LW
Kaplan
D R
DR
Gerson
S L
SL
Albert
J M
JM
Dunn
D E
DE
Lazarus
H M
HM
Medof
M E
ME
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*
D018952:Antigens, CD34; D018957:CD59 Antigens; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D019014:fas Receptor; C080086:phosphatidylinositol glycan-class A protein