Constitutively Active β-Catenin Promotes Expansion of Multipotent Hematopoietic Progenitors in Culture

Abstract

This study was designed to investigate one component of the Wnt/β-catenin signaling pathway that has been implicated in stem cell self-renewal. Retroviral-mediated introduction of stable β-catenin to primitive murine bone marrow cells allowed the expansion of multipotential c-KitlowSca-1low/−CD19− CD11b/Mac-1−Flk-2−CD43+AA4.1+NK1.1−CD3−CD11c−Gr-1−CD45R/B220+ cells in the presence of stromal cells and cytokines. They generated myeloid, T, and B lineage lymphoid cells in culture, but had no T lymphopoietic potential when transplanted. Stem cell factor and IL-6 were found to be minimal requirements for long-term, stromal-free propagation, and a β-catenin-transduced cell line was maintained for 5 mo with these defined conditions. Although multipotential and responsive to many normal stimuli in culture, it was unable to engraft several types of irradiated recipients. These findings support previous studies that have implicated the canonical Wnt pathway signaling in regulation of multipotent progenitors. In addition, we demonstrate how it may be experimentally manipulated to generate valuable cell lines.

Wnt3a deficiency irreversibly impairs hematopoietic stem cell self-renewal and leads to defects in progenitor cell differentiation

“…Indeed different ␤-catenin gain-of-function studies yielded different results probably reflecting the effect of different dosages of activation of the pathway. 6,8,9 The notion that correct levels of Wnt signaling are critical in inducing a self-renewal type of expansion in HSC 45 is also supported by experiments from Baba et al, 46,47 who showed that it is possible to induce some stem cell characteristics in committed progenitors by ectopic expression of stable ␤-catenin. However, such progenitors did not reconstitute T cells when transplanted to immunodeficient mice, showing again that a very strong signal that bypasses all negative regulatory steps of the Wnt-signaling cascade such as given by stabilized ␤-catenin cannot mimic events triggered by tightly regulated physiological Wnt signals.…”

Pivotal role for glycogen synthase kinase–3 in hematopoietic stem cell homeostasis in mice

“…The serial BM from WT and β-catenin CKO transplantation data (summarized in Figure 5C) demonstrate that β-catenin contributes to HSC maintenance throughout successive transplants of Gsk3-deficient BM cells. These data, taken together with previously published observations from others (11,(20)(21)(22)(23)(24)(25)(26)(27)(28), support a positive role for the Wnt/β-catenin pathway in HSC maintenance in the context of reduced GSK-3 activity, but also suggest that an additional GSK-3-regulated pathway plays a distinct and possibly antagonistic role in HSC maintenance.…”

“…A role for Wnt signaling in hematopoiesis is supported by observations that Wnt ligands enhance proliferation of HSCs ex vivo (20)(21)(22) and that Wnt antagonists inhibit HSC proliferation and reconstitution (23,24). In addition, overexpression of stabilized β-catenin promotes HSC self renewal and proliferation ex vivo under certain conditions (20)(21)(22)(23)25), and conditional deletion of β-catenin using vav-cre impairs HSC function in competitive repopulation assays (26). Furthermore, long-term reconstituting capacity in serial transplants is impaired in HSCs recovered from fetal liver of Wnt3a KO embryos (20-23, 25, 27) or from adults overexpressing the Wnt inhibitor Dkk in the hematopoietic niche (28), which suggests that Wnt signaling is required to maintain the long-term repopulating activity of HSCs.…”

“…Although the requirement for Wnt signaling in basal hematopoiesis remains controversial, activation of Wnt signaling can enhance HSC self renewal and HPC function in vivo and ex vivo (10,11,(20)(21)(22)(23)25). We therefore tested whether β-catenin is required for the effects of Gsk3 depletion on hematopoiesis using a conditional β-catenin loss-of-function allele (β-catenin fl/fl ) crossed to interferon-inducible Mx-cre mice (29).…”

Activation of Wnt/β-Catenin Protein Signaling Induces Mitochondria-mediated Apoptosis in Hematopoietic Progenitor Cells

“…However, others reported that purified Wnt protein in vitro (5) or inhibition of GSK3␤ in vivo (6) promotes the self-renewal of HSCs. Baba et al (7,8) showed that constitutive activation of ␤-catenin impaired multilineage differentiation, whereas others found that expression of constitutively active ␤-catenin in hematopoietic cells in vivo leads to loss of HSC repopulation and multilineage differentiation block (9,10). The role of ␤-catenin in the regulation of cell proliferation and apoptosis is cell context-dependent.…”