Regulation of hematopoiesis by the Smad signaling pathway

University dissertation from Department of Molecular Medicine and Gene Therapy

Abstract: Hematopoietic stem cells (HSCs) reside in the bone marrow (BM) of adult individuals and are ultimately responsible for the continuous production of blood cells throughout life. The regulation of HSCs in vivo is tightly regulated by both intrinsic and extrinsic factors. The Smad-signaling pathway is an evolutionary conserved signaling circuitry with critical functions during embryogenesis and throughout adult life, regulating diverse biological processes. The transforming growth factor-? (TGF-?) superfamily of ligands transduce their signals intracellularly through the Smad pathway. A large number of studies, the majority of which have been carried out in vitro, have cataloged TGF-? as a potent negative regulator of HSC proliferation. However, due to embryonic lethality of knockout mice, in vivo investigations of the role of TGF-? and the downstream Smad pathway in the context of adult hematopoiesis have been hampered. To address this, we made use of the Cre/loxP system for inducible gene deletion of two different components of the TGF-? signaling pathway, the type I TGF-? receptor and Smad4 respectively. In addition, retroviral mediated gene transfer to HSCs was used as a tool to block the entire Smad-signaling pathway, by overexpression of the inhibitory Smad7. Induced disruption of the type I TGF-? receptor in adult mice resulted in an inflammatory disorder with a lethal outcome 8-10 weeks post induction. However, all hematopoietic parameters were normal under steady state conditions as well as the regenerative- and self-renewal capacity of mutant HSCs as assessed by transplantation. Smad4 null HSCs exhibited impaired repopulative capacity in a competitive repopulation assay, a behavior that was exacerbated upon secondary transplantation. Overexpression of Smad7 in HSCs resulted in increased regenerative capacity upon secondary transplantation, with a normal lineage distribution. Taken together, our data suggests that the Smad pathway is a critical regulator of HSC self-renewal in vivo.

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