Effect of proinflammatory cytokines on PIGA− hematopoiesis

Objective. Blood cells from patients with paroxysmal nocturnal hemoglobinuria lack glycosyl phosphatidylinositol (GPI)-linked proteins, due to a somatic mutation in the X-linked PIGA gene. It is believed that clonal expansion of PIGA− blood cells is due to a survival advantage in the hostile marrow environment of aplastic anemia. Here we investigated the effects of inhibitory cytokines in mice genetically engineered to have blood cells deficient in GPI-linked proteins. Materials and Methods. The effect of inhibitory cytokines (tumor necrosis factor-α [TNF-α], interferon-γ [IFN-γ], macrophage inflammatory protein-1 alpha [MIP-1α], and transforming growth factor-β1 [TGF-β1]) was investigated, using clonogenic assays, competitive repopulation, and in vivo induction of proinflammatory cytokines by double-stranded RNA. The expression of Fas on progenitor cells and its up-regulation by inhibitory cytokines were analyzed by flow cytometry. Results. TNF-α, IFN-γ, MIP-1α, and TGF-β1 suppressed colony formation in a dose-dependent fashion that was similar for PIGA+ and PIGA− blood bone marrow cells. Competitive repopulation of bone marrow cells cultured in IFN-γ and TNF-α resulted in a comparable ability of PIGA+ and PIGA− hematopoietic stem cells to reconstitute hematopoiesis. Fas expression was minimal on PIGA+ and PIGA− progenitor cells and was up-regulated to the same extent in response to IFN-γ and TNF-α as assessed by Fas antibody-mediated apoptosis. Similarly, in vivo induction of proinflammatory cytokines by double-stranded RNA had no effect on the proportion of circulating PIGA− blood cells. Conclusions. These results indicate that PIGA+ and PIGA− hematopoietic progenitor cells respond similarly to inhibitory cytokines, suggesting that other factors are responsible for the clonal expansion of paroxysmal nocturnal hemoglobinuria cells.