Impaired growth and elevated Fas receptor expression in PIGA(+) stem cellsin 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-mutat ed cells are able to expand in PNH marrow, however, is as yet unclear. To a ddress this question, we compared the growth of affected CD59(-)CD34(+) and unaffected CD59(+)CD34(+) cells from patients with that of normal CD59(+)C D34(+) cells in liquid culture. One hundred FAGS-sorted cells were added pe r well into microtiter plates, and after 11 days at 37 degrees C the progen y were counted and were analyzed for their differentiation pattern. We foun d that CD59-CD34(+) cells from PNH patients proliferated to levels approach ing 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' C D59(-) and CD59(+)CD34(+) cells were equivalent with respect to early diffe rentiation markers, and following culture, the CD45 differentiation pattern s 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 susceptibil ity to apoptosis is one factor involved in the growth impairment.