Search

Bone marrow niches for hematopoietic progenitor cells are not well defined despite their critical role in blood homeostasis. We previously found that cells expressing osteocalcin, a marker of mature osteolineage cells, regulate the production of thymic-seeding T lymphoid progenitors. Here, using a selective cell deletion strategy, we demonstrate that a subset of mesenchymal cells expressing osterix, a marker of bone precursors in the adult, serve to regulate the maturation of early B lymphoid precursors by promoting pro-B to pre-B cell transition through insulin-like growth factor 1 (IGF-1) production. Loss of Osx+ cells or Osx-specific deletion of IGF-1 led to a failure of B cell maturation and the impaired adaptive immune response. These data highlight the notion that bone marrow is a composite of specialized niches formed by pairings of specific mesenchymal cells with parenchymal stem or lineage committed progenitor cells, thereby providing distinctive functional units to regulate hematopoiesis.

The bone marrow niche is recognized as a central player in maintaining and regulating the behavior of hematopoietic stem and progenitor cells. Specific gain-of and loss-of function experiments perturbing a range of osteolineage cells or their secreted proteins had been shown to affect stem cell maintenance (Calvi et al, 2003 [1]; Stier et al., 2005 [2]; Zhang et al., 2003 [3]; Nilsson et al., 2005 [4]; Greenbaum et al., 2013 [5]) and engraftment (Adam et al., 2006, 2009 [6,7]). We used specific in vivo cell deletion approaches to dissect the niche cell-parenchymal cell dependency in a complex bone marrow microenvironment. Endogenous deletion of osteocalcin-expressing (Ocn+) cells led to a loss of T immune cells (Yu et al., 2015 [8]. Ocn+ cells express the Notch ligand DLL4 to communicate with T-competent progenitors, and thereby ensuring T precursor production and expression of chemotactic molecules on their cell surface for subsequent thymic seeding. In contrast, depletion of osterix-expressing (Osx+) osteoprogenitors led to reduced B immune cells. These distinct hematopoietic phenotypes suggest specific pairing of mesenchymal niche cells and parenchymal hematopoietic cells in the bone marrow to create unique functional units to support hematopoiesis. Here, we present the global gene expression profiles of these osteolineage subtypes utilizing a triple fluorescent transgenic mouse model (OsxCre+;Rosa-mCh+;Ocn:Topaz+) that labels Osx+ cells red, Ocn+ cells green, and Osx+Ocn+ cells yellow. This system allows isolation of distinct osteolineage subsets within the same animal by flow cytometry. Array data that have been described in our study [8] are also publically available from NCBI Gene Expression Omnibus (GEO) with the accession number GSE66042. Differences in gene expression may correlate with functional difference in supporting hematopoiesis.

Background Residual chronic myeloid leukemia disease following imatinib treatment has been attributed to the presence of quiescent leukemic stem cells intrinsically resistant to imatinib. Mesenchymal stromal cells in the bone marrow may favor the persistence and progression of leukemia by preserving the proliferation and self-renewal capacities of the malignant progenitor cells.Design and Methods BV173 or primary chronic myeloid leukemia cells were co-cultured with human mesenchymal stromal cells and imatinib-induced cell death was then measured. The roles of pro-and anti-apoptotic proteins and chemokine CXCL12 in this context were evaluated. We also studied the ability of BV173 cells to repopulate NOD/SCID mice following in vitro exposure to imatinib and mesenchymal stromal cells.Results Whilst imatinib induced dose-dependent apoptosis of BV173 cells and primary chronic myeloid leukemia cells, co-culture with mesenchymal stromal cells protected both types of chronic myeloid leukemia cells. Molecular analysis indicated that mesenchymal stromal cells reduced caspase-3 activation and modulated the expression of the anti-apoptotic protein Bcl-XL. Furthermore, chronic myeloid leukemia cells exposed to imatinib in the presence of mesenchymal stromal cells retained the ability to engraft into NOD/SCID mice. We observed that chronic myeloid leukemia cells and mesenchymal stromal cells express functional levels of CXCR4 and CXCL12, respectively. Finally, the CXCR4 antagonist, AMD3100 restored apoptosis by imatinib and the susceptibility of the SCID leukemia repopulating cells to the tyrosine kinase inhibitor.Conclusions Human mesenchymal stromal cells mediate protection of chronic myeloid leukemia cells from imatinib-induced apoptosis. Disruption of the CXCL12/CXCR4 axis restores, at least in part, the leukemic cells’ sensitivity to imatinib. The combination of anti-CXCR4 antagonists with tyrosine kinase inhibitors may represent a powerful approach to the treatment of chronic myeloid leukemia.

ABSTRACT Objective: The aim of the present study was to evaluate and compare the cytotoxic effects of Biodentine and MTA on dental pulp stem cells (DPSCs) and to assess cell viability and adherence after material exposure to an acidic environment. Material and Methods: DPSCs were cultured either alone or in contact with either: Biodentine; MTA set for 1 hour; or MTA set for 24 hours. After 4 and 7 days, cell viability was measured using the MTT assay. Biodentine and MTA were also prepared and packed into standardized bovine dentin disks and divided into three groups according to the storage media (n=6/group): freshly mixed materials without storage medium (Group A); materials stored in saline (Group B); materials stored in citric acid buffered at pH 5.4 (Group C). After 24 hours, DPSCs were introduced in the wells and cell adherence, viability, and cellular morphology were observed via confocal microscopy after three days of culture. Cell viability was analyzed using repeated-measures analysis of variance test with Tukey's post hoc tests (α=0.05). Results: Biodentine expressed significantly higher cell viability compared with all other groups after 4 days, with no differences after 7 days. Notably, cell viability was significantly greater in 24-hour set MTA compared with 1-hour set MTA and control groups after 7 days. Material exposure to an acidic environment showed an increase in cell adherence and viability in both groups. Conclusions: Biodentine induced a significantly accelerated cell proliferation compared with MTA. Setting of these materials in the presence of citric acid enhanced DPSC viability and adherence.