49 results found
MacLean AL, Lo Celso C, Stumpf MPH, 2017, Concise Review: Stem Cell Population Biology: Insights from Hematopoiesis, STEM CELLS, Vol: 35, Pages: 80-88, ISSN: 1066-5099
Khorshed RA, Lo Celso C, 2016, MACHINE LEARNING CLASSIFICATION OF COMPLEX VASCULATURE STRUCTURES FROM IN-VIVO BONE MARROW 3D DATA, IEEE 13th International Symposium on Biomedical Imaging (ISBI), Publisher: IEEE, Pages: 1217-1220, ISSN: 1945-7928
Khorshed RA, Lo Celso C, 2016, Automated identification and measurement of Hematopoietic Stem Cells in 3D Intravital Microscopy Data, Microscopy and Analysis, Editors: Stanciu, Publisher: InTech, ISBN: 978-953-51-2578-5
Image analysis and quantification of Haematopoietic stem cells (HSCs) position within their surrounding microenvironment in the bone marrow is a fast growing area of research, as it holds the key to understanding the dynamics of HSC-niche interactions and their multiple implications in normal tissue development and in response to various stress events. However, this area of research is very challenging due to the complex cellular structure of such images. Therefore, automated image analysis tools are required to simplify the biological interpretation of 3D HSC microenvironment images. In this chapter, we describe how 3D intravital microscopy data can be visualised and analysed using a computational method that allows the automated quantification of HSC position relative to surrounding niche components.
Silberstein L, Goncalves KA, Kharchenko PV, et al., 2016, Proximity-Based Differential Single-Cell Analysis of the Niche to Identify Stem/Progenitor Cell Regulators., Cell Stem Cell, Vol: 19, Pages: 530-543
Physiological stem cell function is regulated by secreted factors produced by niche cells. In this study, we describe an unbiased approach based on the differential single-cell gene expression analysis of mesenchymal osteolineage cells close to, and further removed from, hematopoietic stem/progenitor cells (HSPCs) to identify candidate niche factors. Mesenchymal cells displayed distinct molecular profiles based on their relative location. We functionally examined, among the genes that were preferentially expressed in proximal cells, three secreted or cell-surface molecules not previously connected to HSPC biology-the secreted RNase angiogenin, the cytokine IL18, and the adhesion molecule Embigin-and discovered that all of these factors are HSPC quiescence regulators. Therefore, our proximity-based differential single-cell approach reveals molecular heterogeneity within niche cells and can be used to identify novel extrinsic stem/progenitor cell regulators. Similar approaches could also be applied to other stem cell/niche pairs to advance the understanding of microenvironmental regulation of stem cell function.
Vainieri ML, Blagborough AM, MacLean AL, et al., 2016, Systematic tracking of altered haematopoiesis during sporozoite-mediated malaria development reveals multiple response points, OPEN BIOLOGY, Vol: 6, ISSN: 2046-2441
Khorshed RA, Hawkins ED, Duarte D, et al., 2015, Automated Identification and Localization of Hematopoietic Stem Cells in 3D Intravital Microscopy Data, STEM CELL REPORTS, Vol: 5, Pages: 139-153, ISSN: 2213-6711
Batista S, Maniati E, Reynolds LE, et al., 2014, Haematopoietic focal adhesion kinase deficiency alters haematopoietic homeostasis to drive tumour metastasis., Nat Commun, Vol: 5
Metastasis is the main cause of cancer-related death and thus understanding the molecular and cellular mechanisms underlying this process is critical. Here, our data demonstrate, contrary to established dogma, that loss of haematopoietic-derived focal adhesion kinase (FAK) is sufficient to enhance tumour metastasis. Using both experimental and spontaneous metastasis models, we show that genetic ablation of haematopoietic FAK does not affect primary tumour growth but enhances the incidence of metastasis significantly. At a molecular level, haematopoietic FAK deletion results in an increase in PU-1 levels and decrease in GATA-1 levels causing a shift of hematopoietic homeostasis towards a myeloid commitment. The subsequent increase in circulating granulocyte number, with an increase in serum CXCL12 and granulocyte CXCR4 levels, was required for augmented metastasis in mice lacking haematopoietic FAK. Overall our findings provide a mechanism by which haematopoietic FAK controls cancer metastasis.
Rashidi NM, Scott MK, Scherf N, et al., 2014, In vivo time-lapse imaging shows diverse niche engagement by quiescent and naturally activated hematopoietic stem cells, BLOOD, Vol: 124, Pages: 79-83, ISSN: 0006-4971
Scott MK, Akinduro O, Lo Celso C, 2014, In vivo 4-dimensional tracking of hematopoietic stem and progenitor cells in adult mouse calvarial bone marrow., J Vis Exp
Through a delicate balance between quiescence and proliferation, self renewal and production of differentiated progeny, hematopoietic stem cells (HSCs) maintain the turnover of all mature blood cell lineages. The coordination of the complex signals leading to specific HSC fates relies upon the interaction between HSCs and the intricate bone marrow microenvironment, which is still poorly understood([1-2]). We describe how by combining a newly developed specimen holder for stable animal positioning with multi-step confocal and two-photon in vivo imaging techniques, it is possible to obtain high-resolution 3D stacks containing HSPCs and their surrounding niches and to monitor them over time through multi-point time-lapse imaging. High definition imaging allows detecting ex vivo labeled hematopoietic stem and progenitor cells (HSPCs) residing within the bone marrow. Moreover, multi-point time-lapse 3D imaging, obtained with faster acquisition settings, provides accurate information about HSPC movement and the reciprocal interactions between HSPCs and stroma cells. Tracking of HSPCs in relation to GFP positive osteoblastic cells is shown as an exemplary application of this method. This technique can be utilized to track any appropriately labeled hematopoietic or stromal cell of interest within the mouse calvarium bone marrow space.
Carlson AL, Fujisaki J, Wu J, et al., 2013, Tracking single cells in live animals using a photoconvertible near-infrared cell membrane label., PLoS One, Vol: 8
We describe a novel photoconversion technique to track individual cells in vivo using a commercial lipophilic membrane dye, DiR. We show that DiR exhibits a permanent fluorescence emission shift (photoconversion) after light exposure and does not reacquire the original color over time. Ratiometric imaging can be used to distinguish photoconverted from non-converted cells with high sensitivity. Combining the use of this photoconvertible dye with intravital microscopy, we tracked the division of individual hematopoietic stem/progenitor cells within the calvarium bone marrow of live mice. We also studied the peripheral differentiation of individual T cells by tracking the gain or loss of FoxP3-GFP expression, a marker of the immune suppressive function of CD4(+) T cells. With the near-infrared photoconvertible membrane dye, the entire visible spectral range is available for simultaneous use with other fluorescent proteins to monitor gene expression or to trace cell lineage commitment in vivo with high spatial and temporal resolution.
Fink J, Kent D, Li J, et al., 2013, HOMOZYGOUS JAK2V617F DRIVES RAPID HEMATOPOIETIC STEM CELL PROLIFERATION AND DIFFERENTIATION AT THE EXPENSE OF SELF-RENEWAL, 42nd Annual Scientific Meeting of the International-Society-for-Experimental-Hematology-and-Stem-Cells (ISEH), Publisher: ELSEVIER SCIENCE INC, Pages: S15-S15, ISSN: 0301-472X
Hawkins ED, Lo Celso C, 2013, Subdivision of bone marrow microenvironments: purpose built homes for haematopoietic stem cells, EMBO JOURNAL, Vol: 32, Pages: 176-177, ISSN: 0261-4189
Joseph C, Quach JM, Walkley CR, et al., 2013, Deciphering hematopoietic stem cells in their niches: a critical appraisal of genetic models, lineage tracing, and imaging strategies., Cell Stem Cell, Vol: 13, Pages: 520-533
In recent years, technical developments in mouse genetics and imaging equipment have substantially advanced our understanding of hematopoietic stem cells (HSCs) and their niche. The availability of numerous Cre strains for targeting HSCs and microenvironmental cells provides extensive flexibility in experimental design, but it can also pose significant challenges due to strain-specific differences in cell specificity. Here we outline various genetic approaches for isolating, detecting, and ablating HSCs and niche components and provide a guide for advantages and caveats to consider. We also discuss opportunities and limitations presented by imaging technologies that allow investigation of HSC behavior in situ.
Lo Celso C, 2013, IN VIVO IMAGING OF QUIESCENT AND PHYSIOLOGICALLY ACTIVATED HAEMATOPOIETIC STEM CELLS, 42nd Annual Scientific Meeting of the International-Society-for-Experimental-Hematology-and-Stem-Cells (ISEH), Publisher: ELSEVIER SCIENCE INC, Pages: S4-S4, ISSN: 0301-472X
MacLean AL, Lo Celso C, Stumpf MPH, 2013, Population dynamics of normal and leukaemia stem cells in the haematopoietic stem cell niche show distinct regimes where leukaemia will be controlled, JOURNAL OF THE ROYAL SOCIETY INTERFACE, Vol: 10, ISSN: 1742-5689
Progatzky F, Dallman MJ, Lo Celso C, 2013, From seeing to believing: labelling strategies for in vivo cell-tracking experiments, INTERFACE FOCUS, Vol: 3, ISSN: 2042-8898
Roeder I, Krinner A, Scherf N, et al., 2013, QUANTIFICATION OF STEM CELL/NICHE INTERACTIONS BY COUPLING IN VIVO IMAGING AND IN SILICO SIMULATION, 42nd Annual Scientific Meeting of the International-Society-for-Experimental-Hematology-and-Stem-Cells (ISEH), Publisher: ELSEVIER SCIENCE INC, Pages: S31-S31, ISSN: 0301-472X
Barrett O, Sottocornola R, Lo Celso C, 2012, In vivo imaging of hematopoietic stem cells in the bone marrow niche., Methods Mol Biol, Vol: 916, Pages: 231-242
Even though hematopoietic stem cells (HSC) are amongst the first somatic stem cells exploited for therapeutic purposes, their application is still limited by the inability to expand them ex vivo without impairing their function. Moreover, it has recently emerged that several types of leukemia develop and relapse through complex interactions with bone marrow (BM) components and may directly affect the HSC and their niche. Increasing attention has therefore been dedicated to the BM microenvironment the HSC reside in, with the view that a better understanding of the molecular regulators of HSC-niche interaction in vivo will allow improving HSC mobilization, collection and transplantation and provide clues for the development of innovative leukemia treatments. This chapter focuses on a recently established technique for the visualization of transplanted hematopoietic stem and progenitor cells (HSPC) within the calvarium bone marrow of live mice (Lo Celso et al. Nature 457:92-96, 2007). Intravital microscopy is a rapidly developing field, driven by constant improvement in both detection technologies (i.e., spatial resolution, depth of penetration, spectral definition) and probe availability (i.e., increasingly sophisticated genetic and chemical reporter systems). We therefore discuss the current limitations and challenges related to intravital microscopy of the HSC niche and introduce a number of potential imaging approaches, which could be promising candidates for future development of this technique.
Sottocornola R, Lo Celso C, 2012, Dormancy in the stem cell niche, STEM CELL RESEARCH & THERAPY, Vol: 3, ISSN: 1757-6512
Cornejo MG, Mabialah V, Sykes SM, et al., 2011, Crosstalk between NOTCH and AKT signaling during murine megakaryocyte lineage specification., Blood, Vol: 118, Pages: 1264-1273
The NOTCH signaling pathway is implicated in a broad range of developmental processes, including cell fate decisions. However, the molecular basis for its role at the different steps of stem cell lineage commitment is unclear. We recently identified the NOTCH signaling pathway as a positive regulator of megakaryocyte lineage specification during hematopoiesis, but the developmental pathways that allow hematopoietic stem cell differentiation into the erythro-megakaryocytic lineages remain controversial. Here, we investigated the role of downstream mediators of NOTCH during megakaryopoiesis and report crosstalk between the NOTCH and PI3K/AKT pathways. We demonstrate the inhibitory role of phosphatase with tensin homolog and Forkhead Box class O factors on megakaryopoiesis in vivo. Finally, our data annotate developmental mechanisms in the hematopoietic system that enable a decision to be made either at the hematopoietic stem cell or the committed progenitor level to commit to the megakaryocyte lineage, supporting the existence of 2 distinct developmental pathways.
Fujisaki J, Wu J, Carlson AL, et al., 2011, In vivo imaging of Treg cells providing immune privilege to the haematopoietic stem-cell niche., Nature, Vol: 474, Pages: 216-219
Stem cells reside in a specialized regulatory microenvironment or niche, where they receive appropriate support for maintaining self-renewal and multi-lineage differentiation capacity. The niche may also protect stem cells from environmental insults including cytotoxic chemotherapy and perhaps pathogenic immunity. The testis, hair follicle and placenta are all sites of residence for stem cells and are immune-suppressive environments, called immune-privileged sites, where multiple mechanisms cooperate to prevent immune attack, even enabling prolonged survival of foreign allografts without immunosuppression. We sought to determine if somatic stem-cell niches more broadly are immune-privileged sites by examining the haematopoietic stem/progenitor cell (HSPC) niche in the bone marrow, a site where immune reactivity exists. We observed persistence of HSPCs from allogeneic donor mice (allo-HSPCs) in non-irradiated recipient mice for 30 days without immunosuppression with the same survival frequency compared to syngeneic HSPCs. These HSPCs were lost after the depletion of FoxP3 regulatory T (T(reg)) cells. High-resolution in vivo imaging over time demonstrated marked co-localization of HSPCs with T(reg) cells that accumulated on the endosteal surface in the calvarial and trabecular bone marrow. T(reg) cells seem to participate in creating a localized zone where HSPCs reside and where T(reg) cells are necessary for allo-HSPC persistence. In addition to processes supporting stem-cell function, the niche will provide a relative sanctuary from immune attack.
Lane SW, Wang YJ, Lo Celso C, et al., 2011, Differential niche and Wnt requirements during acute myeloid leukemia progression, BLOOD, Vol: 118, Pages: 2849-2856, ISSN: 0006-4971
Lo Celso C, Lin CP, Scadden DT, 2011, In vivo imaging of transplanted hematopoietic stem and progenitor cells in mouse calvarium bone marrow, NATURE PROTOCOLS, Vol: 6, Pages: 1-14, ISSN: 1754-2189
Lo Celso C, Scadden DT, 2011, The haematopoietic stem cell niche at a glance, JOURNAL OF CELL SCIENCE, Vol: 124, Pages: 3529-3535, ISSN: 0021-9533
Rodriguez S, Wang L, Mumaw C, et al., 2011, The SKP2 E3 ligase regulates basal homeostasis and stress-induced regeneration of HSCs, BLOOD, Vol: 117, Pages: 6509-6519, ISSN: 0006-4971
Sanchez-Aguilera A, Lee YJ, Lo Celso C, et al., 2011, Guanine nucleotide exchange factor Vav1 regulates perivascular homing and bone marrow retention of hematopoietic stem and progenitor cells., Proc Natl Acad Sci U S A, Vol: 108, Pages: 9607-9612
Engraftment and maintenance of hematopoietic stem and progenitor cells (HSPC) depend on their ability to respond to extracellular signals from the bone marrow microenvironment, but the critical intracellular pathways integrating these signals remain poorly understood. Furthermore, recent studies provide contradictory evidence of the roles of vascular versus osteoblastic niche components in HSPC function. To address these questions and to dissect the complex upstream regulation of Rac GTPase activity in HSPC, we investigated the role of the hematopoietic-specific guanine nucleotide exchange factor Vav1 in HSPC localization and engraftment. Using intravital microscopy assays, we demonstrated that transplanted Vav1(-/-) HSPC showed impaired early localization near nestin(+) perivascular mesenchymal stem cells; only 6.25% of Vav1(-/-) HSPC versus 45.8% of wild-type HSPC were located less than 30 μm from a nestin(+) cell. Abnormal perivascular localization correlated with decreased retention of Vav1(-/-) HSPC in the bone marrow (44-60% reduction at 48 h posttransplant, compared with wild-type) and a very significant defect in short- and long-term engraftment in competitive and noncompetitive repopulation assays (<1.5% chimerism of Vav1(-/-) cells vs. 53-63% for wild-type cells). The engraftment defect of Vav1(-/-) HSPC was not related to alterations in proliferation, survival, or integrin-mediated adhesion. However, Vav1(-/-) HSPC showed impaired responses to SDF1α, including reduced in vitro migration in time-lapse microscopy assays, decreased circadian and pharmacologically induced mobilization in vivo, and dysregulated Rac/Cdc42 activation. These data suggest that Vav1 activity is required specifically for SDF1α-dependent perivascular homing of HSPC and suggest a critical role for this localization in retention and subsequent engraftment.
Silberstein L, Osawa M, Lin C, et al., 2011, Real-Time RT-PCR Analysis of Individual Osteolineage Cells within the Hematopoietic Stem Cell Niche, 53rd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH)/Symposium on the Basic Science of Hemostasis and Thrombosis, Publisher: AMER SOC HEMATOLOGY, Pages: 1028-1029, ISSN: 0006-4971
Strydom N, Lo Celso C, Scott M, et al., 2011, Using intravital microscopy to investigate the molecules that regulate the homing of senescent neutrophils to the bone marrow and their phagocytosis by bone marrow macrophages, Annual Congress of the British-Society-for-Immunology, Publisher: WILEY-BLACKWELL, Pages: 127-127, ISSN: 0019-2805
Ferraro F, Celso CL, Scadden D, 2010, Adult stem cels and their niches., Adv Exp Med Biol, Vol: 695, Pages: 155-168, ISSN: 0065-2598
Stem cells participate in dynamic physiologic systems that dictate the outcome of developmental events and organismal stress, Since these cells are fundamental to tissue maintenance and repair, the signals they receive play a critical role in the integrity of the organism. Much work has focused on stem cell identification and the molecular pathways involved in their regulation. Yet, we understand little about how these pathways achieve physiologically responsive stem cell functions. This chapter will review the state of our understanding of stem cells in the context of their microenvironment regarding the relation between stem cell niche dysfunction, carcinogenesis and aging.
Lane SW, Sykes SM, Al-Shahrour F, et al., 2010, The Apc(min) mouse has altered hematopoietic stem cell function and provides a model for MPD/MDS., Blood, Vol: 115, Pages: 3489-3497
Apc, a negative regulator of the canonical Wnt signaling pathway, is a bona-fide tumor suppressor whose loss of function results in intestinal polyposis. APC is located in a commonly deleted region on human chromosome 5q, associated with myelodysplastic syndrome (MDS), suggesting that haploinsufficiency of APC contributes to the MDS phenotype. Analysis of the hematopoietic system of mice with the Apc(min) allele that results in a premature stop codon and loss of function showed no abnormality in steady state hematopoiesis. Bone marrow derived from Apc(min) mice showed enhanced repopulation potential, indicating a cell intrinsic gain of function in the long-term hematopoietic stem cell (HSC) population. However, Apc(min) bone marrow was unable to repopulate secondary recipients because of loss of the quiescent HSC population. Apc(min) mice developed a MDS/myeloproliferative phenotype. Our data indicate that Wnt activation through haploinsufficiency of Apc causes insidious loss of HSC function that is only evident in serial transplantation strategies. These data provide a cautionary note for HSC-expansion strategies through Wnt pathway activation, provide evidence that cell extrinsic factors can contribute to the development of myeloid disease, and indicate that loss of function of APC may contribute to the phenotype observed in patients with MDS and del(5q).
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