scholarly journals Oleic acid triggers hippocampal neurogenesis by binding to TLX/NR2E1

2020 ◽  
Author(s):  
Prasanna Kandel ◽  
Fatih Semerci ◽  
Aleksandar Bajic ◽  
Dodge Baluya ◽  
LiHua Ma ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Oleic Acid ◽  
Progenitor Cell ◽  
Monounsaturated Fatty Acids ◽  
Hippocampal Neurogenesis ◽  
Adult Hippocampal Neurogenesis ◽  
Orphan Nuclear Receptor ◽  
Endogenous Ligand ◽  
Self Renewal ◽  
Stem And Progenitor Cells

AbstractAdult hippocampal neurogenesis underpins learning, memory, and mood, but diminishes with age and illness. The orphan nuclear receptor TLX/NR2E1 is known to regulate neural stem and progenitor cell self-renewal and proliferation, but the precise mechanism by which it accomplishes this is unknown. We found that neural stem and progenitor cells require monounsaturated fatty acids to survive and proliferate. Specifically, oleic acid (18:1ω9) binds to TLX to convert it from a transcriptional repressor to a transcriptional activator of cell cycle and neurogenesis genes. We propose a model in which sufficient quantities of this endogenous ligand must bind to TLX to trigger the switch to proliferation. These findings pave the way for future therapeutic manipulations to counteract pathogenic impairments of neurogenesis.

Cell Cycle Progression and Self-Renewal Divisions of Cord Blood Derived CD34+ Cells Treated with the Polyamine Copper-Chelator Tetraethylenepentamine.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4237-4237
Author(s):  
Toni Peled ◽  
Noga R. Goudsmid ◽  
Frida Grynspan ◽  
Sophie Adi ◽  
Efrat Landau ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Progenitor Cells ◽  
Fluorescence Intensity ◽  
Cell Expansion ◽  
Progenitor Cell ◽  
Self Renewal ◽  
Cell Cycle Profile ◽  
Cd34 Cells ◽  
Copper Chelator

Abstract In vitro cell expansion is constrained by default pathways of commitment and differentiation resulting in limited expansion of hematopoietic stem-progenitor cells (HSPCs). Still, several ex vivo manipulations have been reported to achieve expansion of HSPCs by altering cell cycle kinetics and enhancing progression through the G1-S barrier. We have previously shown that addition of tetraethylenepentamine (TEPA), a polyamine copper chelator, to cytokine-supplemented CD34+ cell cultures modulates cytokine-driven hematopoietic cell fate in vitro, resulting in remarkable expansion of a cell population that displays phenotypic and functional characteristics of HSPCs (Exp Hematol.2004;32 (6):547–55). The objective of the present study was to evaluate the mechanism leading to expansion of early progenitor cells following short-term exposure to TEPA. To this end, cell cycle profile, tracking of proliferation history, as well as determination of actual numbers of progenitor subsets were studied. In order to follow the extent of proliferation by tracking the number of cellular divisions, freshly isolated CD34+ cells were labeled with PKH2, a membrane dye that is sequentially diluted during every cell division. Fluorescence intensities of CD34+ and that of a more immature CD34+CD38− cell subset were determined immediately after staining. The cells were then cultured in serum-containing medium and a cocktail of cytokines (SCF, TPO, IL-6, Flt3-ligand, at 50 ng/ml each and IL-3 at 20 ng/ml), with and without TEPA. Total nucleated cells (TNC), purified CD34+ cells and CD34+CD38− cells were analyzed for PKH2 fluorescence intensity during the first two weeks of culture. Cell cycle profile was detected with the DNA intercalating agent propidium iodide, which determines cellular DNA content. FACS analysis of the cultured cells as well as progenitor cell quantification by immuno-affinity purification revealed comparable expansion levels of TNC and CD34+ cells in both TEPA-treated and control cultures during the first two weeks, as previously published. Although similar CD34+ cell numbers were observed, the mean frequency of CD34+CD38− and CD34+CD38-Lin- cells within the CD34+ cell population was significantly higher in TEPA-treated cultures over the control (0.2 vs. 0.04 and 0.07 vs. 0.01, respectively; n=6, p<0.05). Median PKH2 fluorescence intensity of CD34+CD38− subset was two fold higher in TEPA than in control cultures, demonstrating that early progenitor cells derived from TEPA-treated cultures consistently accomplished less proliferation cycles as compared to early progenitor cells derived from control cultures. This effect was not mirrored by a significant alteration of the cell cycle profile (Control (%): G1=26±14, S=2.6±0.1, G2=0.7±0.4; TEPA(%): G1=29±12, S=1.7±0.9, G2=0.4±0.2). Taken together, the data suggest that during cycling, the CD34+CD38− phenotype is preserved more successfully in TEPA-treated than in control cultures, suggesting retention of self-renewing potential of early progenitor cells under these culture conditions. This mechanism also supports a role for TEPA in inhibition of early progenitor cell differentiation. Ongoing work is aimed at further defining whether phenotype reversion or self-renewal (or both) lie at the foundation of TEPA-mediated progenitor cell expansion.

Malignant Myeloma Cells Impair Phenotype and Function of stem and Progenitor Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1799-1799
Author(s):  
Ingmar Bruns ◽  
Sebastian Büst ◽  
Akos G. Czibere ◽  
Ron-Patrick Cadeddu ◽  
Ines Brückmann ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Progenitor Cells ◽  
Plasma Cells ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Healthy Donors ◽  
Stem And Progenitor Cells

Abstract Abstract 1799 Poster Board I-825 Multiple myeloma (MM) patients often present with anemia at the time of initial diagnosis. This has so far only attributed to a physically marrow suppression by the invading malignant plasma cells and the overexpression of Fas-L and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) by malignant plasma cells triggering the death of immature erythroblasts. Still the impact of MM on hematopoietic stem cells and their niches is scarcely established. In this study we analyzed highly purified CD34+ hematopoietic stem and progenitor cell subsets from the bone marrow of newly diagnosed MM patients in comparison to normal donors. Quantitative flowcytometric analyses revealed a significant reduction of the megakaryocyte-erythrocyte progenitor (MEP) proportion in MM patients, whereas the percentage of granulocyte-macrophage progenitors (GMP) was significantly increased. Proportions of hematopoietic stem cells (HSC) and myeloid progenitors (CMP) were not significantly altered. We then asked if this is also reflected by clonogenic assays and found a significantly decreased percentage of erythroid precursors (BFU-E and CFU-E). Using Affymetrix HU133 2.0 gene arrays, we compared the gene expression signatures of stem cells and progenitor subsets in MM patients and healthy donors. The most striking findings so far reflect reduced adhesive and migratory potential, impaired self-renewal capacity and disturbed B-cell development in HSC whereas the MEP expression profile reflects decreased in cell cycle activity and enhanced apoptosis. In line we found a decreased expression of the adhesion molecule CD44 and a reduced actin polymerization in MM HSC by immunofluorescence analysis. Accordingly, in vitro adhesion and transwell migration assays showed reduced adhesive and migratory capacities. The impaired self-renewal capacity of MM HSC was functionally corroborated by a significantly decreased long-term culture initiating cell (LTC-IC) frequency in long term culture assays. Cell cycle analyses revealed a significantly larger proportion of MM MEP in G0-phase of the cell cycle. Furthermore, the proportion of apoptotic cells in MM MEP determined by the content of cleaved caspase 3 was increased as compared to MEP from healthy donors. Taken together, our findings indicate an impact of MM on the molecular phenotype and functional properties of stem and progenitor cells. Anemia in MM seems at least partially to originate already at the stem and progenitor level. Disclosures Off Label Use: AML with multikinase inhibitor sorafenib, which is approved by EMEA + FDA for renal cell carcinoma.

scholarly journals Reactive, Adult Neurogenesis From Increased Neural Progenitor Cell Proliferation Following Alcohol Dependence in Female Rats

2021 ◽  
Vol 15 ◽  
Author(s):  
Natalie N. Nawarawong ◽  
K. Ryan Thompson ◽  
Steven P. Guerin ◽  
Chinchusha Anasooya Shaji ◽  
Hui Peng ◽  
...  
Keyword(s):  
Alcohol Dependence ◽  
Dentate Gyrus ◽  
Adult Neurogenesis ◽  
Progenitor Cell ◽  
Neural Progenitor Cell ◽  
Neural Progenitor ◽  
Hippocampal Neurogenesis ◽  
Adult Hippocampal Neurogenesis ◽  
Female Rats ◽  
Immature Neurons

Hippocampal neurodegeneration is a consequence of excessive alcohol drinking in alcohol use disorders (AUDs), however, recent studies suggest that females may be more susceptible to alcohol-induced brain damage. Adult hippocampal neurogenesis is now well accepted to contribute to hippocampal integrity and is known to be affected by alcohol in humans as well as in animal models of AUDs. In male rats, a reactive increase in adult hippocampal neurogenesis has been observed during abstinence from alcohol dependence, a phenomenon that may underlie recovery of hippocampal structure and function. It is unknown whether reactive neurogenesis occurs in females. Therefore, adult female rats were exposed to a 4-day binge model of alcohol dependence followed by 7 or 14 days of abstinence. Immunohistochemistry (IHC) was used to assess neural progenitor cell (NPC) proliferation (BrdU and Ki67), the percentage of increased NPC activation (Sox2+/Ki67+), the number of immature neurons (NeuroD1), and ectopic dentate gyrus granule cells (Prox1). On day seven of abstinence, ethanol-treated females showed a significant increase in BrdU+ and Ki67+ cells in the subgranular zone of the dentate gyrus (SGZ), as well as greater activation of NPCs (Sox2+/Ki67+) into active cycling. At day 14 of abstinence, there was a significant increase in the number of immature neurons (NeuroD1+) though no evidence of ectopic neurogenesis according to either NeuroD1 or Prox1 immunoreactivity. Altogether, these data suggest that alcohol dependence produces similar reactive increases in NPC proliferation and adult neurogenesis. Thus, reactive, adult neurogenesis may be a means of recovery for the hippocampus after alcohol dependence in females.

Self-Renewal and Differentiation in Hematopoietic Stem and Progenitor Cells Is Controlled By the APC/C Coactivator Cdh1

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2370-2370
Author(s):  
Daniel Ewerth ◽  
Stefanie Kreutmair ◽  
Birgit Kügelgen ◽  
Dagmar Wider ◽  
Julia Felthaus ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Research Funding ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Nsg Mice ◽  
Human Cd34 ◽  
Cd34 Cells ◽  
Stem And Progenitor Cells

Abstract Introduction: Hematopoietic stem and progenitor cells (HSPCs) represent the lifelong source of all blood cells and continuously renew the hematopoietic system by differentiation into mature blood cells. The process of differentiation is predominantly initiated in G1 phase of the cell cycle when stem cells leave their quiescent state. During G1 the anaphase-promoting complex or cyclosome (APC/C) associated with the coactivator Cdh1 is highly active and marks proteins for proteasomal degradation to regulate proliferation. In addition, Cdh1 has been shown to control terminal differentiation in neurons, muscle cells or osteoblasts. Here we show that Cdh1 is also a critical regulator of human HSPC differentiation and self-renewal. Methods: Human CD34+ cells were collected from peripheral blood (PB) of G-CSF mobilized donors and cultured in the presence of different cytokine combinations. To analyze cell division and self-renewal versus differentiation, CFSE staining was used in combination with flow cytometric detection of CD34 expression. The knockdown and overexpression of Cdh1 was achieved by lentiviral delivery of suitable vectors into target cells. After cell sorting transduced (GFP+) CD34+ cells were used for in vitro differentiation in liquid culture or CFU assay. For in vivo experiments purified cells were transplanted into NSG mice. Results: G-CSF mobilized CD34+ cells showed effective differentiation into granulocytes (SCF, G-CSF), erythrocytes (SCF, EPO) or extended self-renewal (SCF, TPO, Flt3-L) when stimulated in vitro. The differentiation was characterized by a fast downregulation of Cdh1 on protein level, while Cdh1 remained expressed under self-renewal conditions. A detailed analysis of different subsets, both in vitro and in vivo, showed high Cdh1 level in CD34+ cells and low expression in myeloid cells. Analysis of proliferation revealed lowest division rates during self-renewal, accompanied by higher frequency of CD34+ cells. The fastest proliferation was found after induction of erythropoiesis. These experiments also showed a more rapid decrease of HSPCs' colony-forming ability and of CD34+ cells during granulopoiesis after 2-3 cell divisions in contrast to a moderate decline under self-renewal conditions. The depletion of Cdh1 (Cdh1-kd) had no effect on total cell numbers or proliferation detected by CFSE during differentiation and self-renewal, but showed an increase in S phase cells. These results were confirmed at the single cell level by measuring the cell cycle length of individual cells. Independent of cell cycle regulation, Cdh1-kd cells showed a significant maintenance of CD34+ cells under self-renewal conditions and during erythropoiesis with lower frequency of Glycophorin A+ cells. In CFU assays, the Cdh1-kd resulted in less primary colony formation, notably CFU-GM and BFU-E, but significantly more secondary colonies compared to control cells. These results suggest that the majority of cells reside in a more undifferentiated state due to Cdh1-kd. The overexpression of Cdh1 showed reversed results with less S phase cells and tendency to increased differentiation in liquid culture and CFU assays. To further validate our results in vivo, we have established a NSG xenotransplant mouse model. Human CD34+ cells depleted of Cdh1 engrafted to a much higher degree in the murine BM 8 and 12 weeks after injection as shown by higher frequencies of human CD45+ cells. Moreover, we also found an increased frequency of human CD19+ B cells after transplantation of CD34+ Cdh1-kd cells. These results suggest an enhanced in vivo repopulation capacity of human CD34+ HSCs in NSG mice when Cdh1 is depleted. Preliminary data in murine hematopoiesis support our hypothesis showing enhanced PB chimerism upon Cdh1-kd. Looking for a mediator of these effects, we found the Cdh1 target protein TRRAP, a cofactor of many HAT complexes, increased upon Cdh1-kd under self-renewal conditions. We use currently RT-qPCR to determine, if this is caused by a transcriptional or post-translational mechanism. Conclusions: Loss of the APC/C coactivator Cdh1 supports self-renewal of CD34+ cells, represses erythropoiesis in vitro and facilitates engraftment capacity and B cell development of human HSPCs in vivo. This work was supported by Josè Carreras Leukemia Foundation grant DCJLS R10/14 (to ME+RW) Disclosures Ewerth: Josè Carreras Leukemia Foundation: Research Funding. Wäsch:German Cancer Aid: Research Funding; Comprehensiv Cancer Center Freiburg: Research Funding; Janssen-Cilag: Research Funding; MSD: Research Funding.

scholarly journals Single Cell-Resolution Analysis of HSC Dysfunction in Mir-146a knockout Mice

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 714-714
Author(s):  
Jennifer Grants ◽  
Joanna Wegrzyn ◽  
David Knapp ◽  
Tony Hui ◽  
Kieran O'Neill ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Transcriptome Profiling ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Proliferation And Differentiation ◽  
Stem And Progenitor Cells ◽  
Resolution Analysis ◽  
Signaling Activation

Abstract MicroRNA miR-146a is frequently depleted in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Loss of miR-146a may be an initiating event in tumorigenesis, as miR-146a loss in mouse models is sufficient to cause features of MDS and eventual progression to AML. To define how miR-146a loss initiates tumorigenesis, we analyzed hematopoietic stem cell (HSC) function from miR-146a knockout (KO) mice prior to onset of an overt malignant phenotype. Tracking cell division kinetics, proliferation, and differentiation of single long-term HSC (LT-HSC; EPCR+CD45+CD48-CD150+) in culture, we found evidence that miR-146a KOreduces HSC quiescence and promotes differentiating cell divisions. Our data show that miR-146a KO HSC dysfunction may stem from loss of a CD150-bright EPCR-bright sub-population, which has previously been associated with robust HSC activity. In line with this, single cell DNA methylation profiling revealed a reduction in a primitive sub-population of LT-HSCs in miR-146a KO animals. In addition, single cell LT-HSC transplants revealed a myeloid repopulation bias. As reduced HSC cell cycle quiescence has been linked to impaired HSC self-renewal upon hematopoietic stress, such as serial transplantation, we assessed the frequency of serially transplantable HSCs by performing secondary transplants with limiting dilution. Serially transplantable HSC frequency was reduced in miR-146a KO compared to wild type, suggesting impaired HSC self-renewal. Transcriptome profiling of miR-146a KO hematopoietic stem and progenitor cells identified tumor necrosis factor (TNF) signaling activation as a potential driver of HSC dysfunction. LT-HSC cell cycle quiescence and the CD150-bright EPCR-bright LT-HSC sub-population were restored in miR-146a/TNF double KO mice, suggesting that aberrant TNF signaling activation drives HSC dysfunction upon loss of miR-146a. Gene expression levels in the TNF signaling network are inversely correlated with miR-146a levels in human AML, implying that TNF signaling may similarly disrupt HSC function in miR-146a- depleted myeloid malignancies. Overall, our findings suggest that miR-146a promotes HSC cell cycle quiescence and inhibits differentiation by antagonizing TNF signaling, in order to maintain a primitive sub-population of long-term self-renewing HSCs. Disclosures Eaves: Experimental Hematology: Other: Editor of journal; StemCell Technologies Inc: Other: Wife of owner.

Models of Pluripotent and Somatic Stem Cells to Study Tissue-Specific Sensitivities in Fanconi Anemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 168-168 ◽  
Author(s):  
Timothy Michael Chlon ◽  
Susanne I Wells ◽  
Sonya Ruiz-Torres ◽  
Matthew Kuhar ◽  
James M Wells
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Progenitor Cells ◽  
Fanconi Anemia ◽  
Squamous Epithelium ◽  
Somatic Stem Cells ◽  
Self Renewal ◽  
Stem And Progenitor Cells ◽  
Pathway Function

Abstract The Fanconi Anemia (FA) DNA Repair pathway functions through homologous recombination for error-free repair of DNA interstrand crosslinks. Loss of function of this pathway causes a complex genetic disease that is characterized by congenital abnormalities, bone marrow failure (BMF), and extreme incidence of squamous cell carcinomas. BMF is caused by exhaustion of hematopoietic stem and progenitor cells (HSPCs) and is nearly 100% penetrant by age 40 in FA patients, indicating a profound sensitivity of HSPCs to FA pathway deficiency. In contrast, stem cells in other rapidly regenerating tissues, such as the skin and intestine, are not similarly exhausted. Interestingly, squamous epithelium is highly prone to transformation while intestinal epithelium is not. In order to explore the developmental origins of such striking tissue-specific phenotypes in FA patients, we have generated induced pluripotent stem cell (iPSC) lines conditional for FA pathway function (cFA-iPSCs) and used them to derive FA-proficient and deficient in vitro models of diverse tissues. FA patient cells are refractory to reprogramming. To circumvent this defect and prevent the selection of FA-resistant iPSC clones, fibroblasts from 2 FANCA patients were inducibly complemented with a FANCA transgene under the control of a tetracycline-inducible promoter and then were reprogrammed to iPSC. In this way, the FA pathway was functional throughout reprogramming and could then be turned on or off in established iPSC lines by the addition or withdrawal of doxycycline (DOX) to the media. Here, we describe the effect of FA pathway loss on iPSCs, and present preliminary data on iPSC-derived equivalents of three lineages: hematopoietic, squamous, and intestinal. First, functional consequences of FA pathway loss on iPSC pluripotency and self-renewal were examined. Upon withdrawal of DOX from the culture media, the complementing FA transgene was effectively silenced, resulting in loss of FA pathway function within 7 days. FA-deficient iPSCs maintained normal expression of OCT-3/4 and NANOG and formed teratomas in NSG mice, indicating that pluripotency was maintained. However, profound cell cycle arrest and apoptosis were observed under normal in vitro culture conditions within 7 days of DOX-withdrawal, and the iPSCs failed to expand by 2-3 passages. Thus, we concluded that iPSCs require an intact FA pathway for self-renewal in vitro. Mechanistic studies of FA pathway-deficient iPSCs revealed a 10-fold increase in gH2AX foci in the G2-M phase of the cell cycle. This correlated with activated DNA damage response signaling through ATR and CHK1. Inhibition of CHK1 completely restored the growth of FA-deficient iPSCs to that of their FA-proficient counterparts through a remarkable rapid bypass of the G2-M checkpoint. Unexpectedly, cells maintained in CHK1 inhibitor for over 40days accrued few karypotypic abnormalities (<5% of cells), of which most were trisomies, and only 1 cell out of 40 contained translocations. The rarity of deletions and translocations in CHK1 inhibited iPSC suggests that error-free repair still occurs by an unknown mechanism. We next differentiated the cFA-iPSCs into 3D squamous epithelium and intestine with timed-withdrawal of the FA pathway to determine the effect of FA pathway loss on tissue development and homeostasis. Squamous epithelial rafts and intestinal organoids were generated in the presence and absence of DOX using established protocols. These demonstrated that FA pathway loss does not cause gross abnormalities in epithelial tissues, in line with patient phenotypes. We will present the latest results on proliferation, survival, and differentiation of stem and progenitor cells within each tissue. These will include an assessment of epithelial hyperplasia, which has been observed previously in immortalized FA patient keratinocyte-derived organotypic squamous epithelial rafts and in the oral epithelia of FANCD2 knockout mice. Finally, we will present our latest data on the effects of FA deficiency on hematopoietic progenitor cells, which are currently being generated from the cFA-iPSCs. Collectively, these experiments will quantify cell intrinsic sensitivity of pluripotent versus somatic stem cells that reside in diverse tissue types to loss of the FA pathway. The results may inform the development of novel therapeutics to treat FA BMF without increasing disease risk in other tissues. Disclosures No relevant conflicts of interest to declare.

Expansion of functionally defined mouse hematopoietic stem and progenitor cells by a short isoform of RUNX1/AML1

Blood ◽  
2012 ◽  
Vol 119 (3) ◽  
pp. 727-735 ◽  
Author(s):  
Shinobu Tsuzuki ◽  
Masao Seto
Keyword(s):  
Progenitor Cell ◽  
Gene Expression Analysis ◽  
Ex Vivo ◽  
Transcriptional Regulators ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Limited Success ◽  
Stem And Progenitor Cells

Abstract Self-renewal activity is essential for the maintenance and regeneration of the hematopoietic system. The search for molecules capable of promoting self-renewal and expanding hematopoietic stem cells (HSCs) has met with limited success. Here, we show that a short isoform (AML1a) of RUNX1/AML1 has such activities. Enforced AML1a expression expanded functionally defined HSCs, with an efficiency that was at least 20 times greater than that of the control in vivo and by 18-fold within 7 days ex vivo. The ex vivo–expanded HSCs could repopulate hosts after secondary transplantations. Moreover, AML1a expression resulted in vigorous and long-term (> 106-fold at 4 weeks) ex vivo expansion of progenitor cell populations capable of differentiating into multilineages. Gene expression analysis revealed that AML1a expression was associated with up-regulation of genes, including Hoxa9, Meis1, Stat1, and Ski. shRNA-mediated silencing of these genes attenuated AML1a-mediated activities. Overall, these findings establish AML1a as an isoform-specific molecule that can influence several transcriptional regulators associated with HSCs, leading to enhanced self-renewal activity and hematopoietic stem/progenitor cell expansion ex vivo and in vivo. Therefore, the abilities of AML1a may have implications for HSC transplantation and transfusion medicine, given that the effects also can be obtained by cell-penetrating AML1a protein.

Intravital Imaging of Young and Aged Stem Cells in the Marrow of Long Bones: Visualizing Mammalian Stem Cell Behavior in Real-Time in Vivo

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2418-2418
Author(s):  
Anja Köhler ◽  
Vince Schmithorst ◽  
Marie-Dominique Filippi ◽  
Marnie A. Ryan ◽  
Deidre Daria ◽  
...  
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Progenitor Cell ◽  
Intravital Imaging ◽  
Long Bones ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Close Vicinity ◽  
Stem And Progenitor Cells

Abstract Hematopoiesis, the process in which blood cells are generated from hematopoietic stem and progenitor cells (HSPCs) is primarily confined to the bone cavities. The interactions of hematopoietic cells with stroma cells forming niches inside the bone cavities are central to hematopoiesis, as these regulate cell proliferation, self-renewal and differentiation. Hematopoietic cell/stroma interactions have thus been, in analogy to the immunological synapse, named stem/progenitor cell synapses. So far, visualization of the behavior of somatic stem and progenitor cells in an undisturbed in vivo environment has not been reported for the mammalian system and consequently, the cellular dynamics of stem, progenitor and differentiated cells in vivo are only poorly defined. We developed and performed intravital time-lapse 2-photon microscopy in the marrow of the long bones (tibia) of mice to study the behavior and dynamics of differentiated hematopoietic cells as well as HPCs and HSCs in close vicinity to the endosteum in vivo over time. We demonstrate that HPCs as well as HSCs reside in close vicinity to the endosteum, further supporting the notion of an endosteal stem cell niche, and that they are, in contrast to differentiated macrophages and dendritic cells, solitary and immobile. Both HPCs and HSCs occupy distinct positions relative to the endosteum and show cell protrusion movement consistent with an active stem/progenitor cell synapse. Lastly, we report that aged HSCs show increased protrusion movement and localize more distantly to the endosteum compared to young HSCs. In addition, aged HSCs present with reduced adhesion to stroma as well as reduced polarity upon adhesion in vitro, implying a connection between altered stem cell dynamics in vivo and stem cell aging. The intravital imaging technology developed might establish a basis for further delineating additional important questions in stem cell biology like cellular mechanisms of hematopoietic stem cell self-renewal and differentiation in the context of the stroma/niche in vivo.

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