Nitric oxide depletion alters hematopoietic stem cell commitment toward immunogenic dendritic cells

BACKGROUND: NO is a key molecule involved in the regulation of cell survival, proliferation and differentiation in many cell types. In this study we investigated the contribution of NO() during the differentiation of human peripheral blood hemopoietic stem cells (CD34(+)HSCs) toward immunogenic dendritic cells (i-DCs). METHODS: We depleted autocrine NO() production, using N(G)-monomethyl-l-arginine monoacetate (l-NMMA) and paracrine NO(), using oxy-hemoglobin (HbO(2)) as a NO() scavenger during in vitro differentiation of CD34(+)HSCs to i-DCs. We monitored the NO() level, cell proliferation, phenotype and differentiation potential. RESULTS: We found that the depletion of paracrine or autocrine NO() correlated with (I) an active proliferation state at the end of differentiation, when control cells were not proliferating; (II) a significant reduction in the expression levels of differentiative markers (CD1a and HLA-DR) with a parallel high expression of the CD34 marker (III) with a retrieved clonogenic ability compared to control cells. CONCLUSIONS: On the whole, our data indicate that the depletion of NO() during the commitment stage blocks CD34(+)HSC differentiation into i-DCs and maintains an undifferentiated, highly proliferating cell population, indicating/revealing a novel role for NO() in the commitment of CD34(+)HSCs into i-DCs. GENERAL SIGNIFICANCE: The essential finding of the present study is that NO(), produced in HSCs by NOS enzymes, may act as autocrine and paracrine effectors regulating the in vitro differentiation process of CD34(+)-HSCs toward i-DCs.