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NAMPT is essential for the G-CSF–induced myeloid differentiation via a NAD+–sirtuin-1–dependent pathway

Nature Medicine volume 15, pages 151–158 (2009)Cite this article

Abstract

We identified nicotinamide phosphoribosyltransferase (NAMPT), also known as pre-B cell colony enhancing factor (PBEF), as an essential enzyme mediating granulocyte colony-stimulating factor (G-CSF)-triggered granulopoiesis in healthy individuals and in individuals with severe congenital neutropenia. Intracellular NAMPT and NAD+ amounts in myeloid cells, as well as plasma NAMPT and NAD+ levels, were increased by G-CSF treatment of both healthy volunteers and individuals with congenital neutropenia. NAMPT administered both extracellularly and intracellularly induced granulocytic differentiation of CD34+ hematopoietic progenitor cells and of the promyelocytic leukemia cell line HL-60. Treatment of healthy individuals with high doses of vitamin B3 (nicotinamide), a substrate of NAMPT, induced neutrophilic granulocyte differentiation. The molecular events triggered by NAMPT include NAD+-dependent sirtuin-1 activation, subsequent induction of CCAAT/enhancer binding protein-α and CCAAT/enhancer binding protein-β, and, ultimately, upregulation of G-CSF synthesis and G-CSF receptor expression. G-CSF, in turn, further increases NAMPT levels. These results reveal a decisive role of the NAD+ metabolic pathway in G-CSF-triggered myelopoiesis.

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Figure 1: G-CSF induces NAMPT and NAD+ in CD33+ myeloid progenitors from healthy individuals and individuals with congenital neutropenia in vivo and in vitro.
Figure 2: NAMPT triggers myeloid differentiation of CD34+ cells from healthy individuals.
Figure 3: Nicotinamide induces in vitro myeloid differentiation of CD34+ cells from healthy individuals.
Figure 4: NAMPT is capable of G-CSF–dependent granulocytic differentiation.
Figure 5: SIRT1 is involved in NAMPT-C/EBP–dependent activation of G-CSF and G-CSFR.
Figure 6: Treatment of healthy individuals with vitamin B3 leads to increased numbers of neutrophils in peripheral blood.

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Acknowledgements

We thank M. Uenalan for helpful discussion, M. Morgan for critically reading the manuscript and M. Ballmaier and C. Reimer for assistance in cell sorting. We also thank C. Zeidler and the physicians within the Severe Chronic Neutropenia International Registry for providing subject material (bone marrow cells, peripheral blood neutrophils and plasma). Special thanks to study subjects and colleagues, especially M. Schatz, for their great cooperation. This work was supported by Elternverein Krebskranker Kinder Hannover e.V., Madeleine-Schickedanz-Kinderkrebsstiftung, Deutsche José Carreras Leukämia Stiftung e.V., REBIRTH Excellence Cluster (A.S.), Else Kröner Foundation (A.S.), an American Federation for Aging Research senior postdoctoral fellowship (F.W.) and the US Department of Agriculture and National Institutes of Health (Q.T.).

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Author notes

  1. Dan Lan

    Present address: Present address: Department of Pediatrics, the first affiliated Hospital of Guang Xi Medical University, Shuang Yong Road, Guang Xi 530021, China.,

  2. Julia Skokowa and Dan Lan: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Hematopoiesis, Hannover Medical School, Carl-Neuberg Strasse 1, Hannover, 30625, Germany

    Julia Skokowa, Dan Lan, Basant Kumar Thakur, Kshama Gupta, Annette Müller Brechlin, Lars Hinrichsen, Martin Stanulla & Karl Welte

  2. US Department of Agriculture–Agricultural Research Service, Children's Nutrition Research Center, Baylor College of Medicine, 1100 Bates Street, Houston, 77030, Texas, USA

    Fei Wang & Qiang Tong

  3. Department of Pediatrics, University Hospital Schleswig-Holstein, Campus Kiel, Arnold-Heller-Strasse 3, Kiel, 24105, Germany

    Gunnar Cario & Martin Stanulla

  4. Department of Experimental Hematology, Hannover Medical School, Carl-Neuberg Strasse 1, Hannover, 30625, Germany

    Axel Schambach

  5. Department of Biochemistry, Hannover Medical School, Carl-Neuberg Strasse 1, Hannover, 30625, Germany

    Gustav Meyer & Matthias Gaestel

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Contributions

K.W. and J.S. made initial observations, designed the main experiments, analyzed the data, supervised experimentation and wrote the manuscript; J.S. and D.L. performed the main experiments; B.K.T. performed western blotting and immunoprecipitation experiments; A.M.B. cloned and prepared plasmids for transfection experiments; K.G. introduced mutations into G-CSF and G-CSFR reporter gene constructs; L.H. cloned G-CSFR shRNA; F.W. and Q.T. provided plasmids for SIRT1, C/EBP-α and C/EBP-β, performed immunoprecipitation experiments in 293T cells and discussed the manuscript; A.S. designed NAMPT lv constructs and produced viral supernatants; G.C., M.G. and M.S. discussed the project and manuscript; G.M. and M.G. provided FK866 and helped to measure NAD+; K.W. coordinated the project and wrote the manuscript.

Corresponding authors

Correspondence to Julia Skokowa or Karl Welte.

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Skokowa, J., Lan, D., Thakur, B. et al. NAMPT is essential for the G-CSF–induced myeloid differentiation via a NAD+–sirtuin-1–dependent pathway. Nat Med 15, 151–158 (2009). https://doi.org/10.1038/nm.1913

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