Mechanism of Calcium Sensing Receptor Mediated Hematopoietic Stem Cell Lodgment In the Adult Bone Marrow Niche

Abstract Abstract 399 The ability of hematopoietic stem cells (HSCs) to maintain an undifferentiated state and undergo self-renewal is partly regulated by external signals originating from the stem cell niche. One receptor expressed on HSCs that is known to be involved in HSC niche biology is the calcium-sensing receptor (CaR). Our previous study using HSCs obtained from the fetal liver of mice deficient in CaR has shown the crucial role of CaR in HSC lodgment and engraftment in the bone marrow (BM), where CaR-/- HSCs lose their ability to lodge in the endosteal surface of the bone, leading to defective engraftment. To further investigate the mechanism of CaR-mediated lodgment of HSCs, we used a pharmacological approach to activate the receptor and assess the in vitro and in vivo effects. Cinacalcet treatment, which acts as a positive allosteric modulator of CaR to increase the sensitivity of the receptor to activation by extracellular Ca2+, leads to a 3-fold increase in primitive hematopoietic cell activity in vitro as assessed by the cobblestone forming cell assay. The increase in activity in vitro does not appear to be an effect of alterations in cell proliferation, cell survival or the expression of cell adhesion molecules such as VLA-4 or L-selectin. Rather, with CaR stimulation, long-term HSCs have an increased adhesion capability to collagen I, a major ECM molecule present predominantly in the BM endosteal region. We also observed that activation of the CaR following Cinacalcet treatment significantly enhances HSC homing to the BM, lodgment at the endosteal surface and in vivo engraftment capabilities as assessed by a competitive repopulation assay. This enhancement of in vivo activity correlates with increased CXCR4 signaling and migration towards an SDF-1alpha stimulus. Signaling through this receptor is known to be important in cell migration, proliferation, survival, and retention of HSCs in the BM following transplantation. Analysis of the CaR on human cells has demonstrated that there is a distinct population of CaR+ cells present in the CD34+ cell population. The frequency of this population varies between approximately 2% on mobilized peripheral blood CD34+ cells and 6% on BM CD34+ cells. Further analysis is being performed to define the identity of this subpopulation of CaR+CD34+ cells. These mechanisms by which the CaR dictates preferential localization of HSCs in the BM endosteal region may provide additional insights for the fundamental interrelationship between the stem cell niche and stem cell fate. These studies also have implications in the area of clinical stem cell transplantation, where ex vivo modulation of the CaR may be envisioned as a strategy to enhance HSC engraftment in the BM. Disclosures: No relevant conflicts of interest to declare.

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MESENCHYMAL TNFR1 EXPRESSION PRESERVES THE COLONIC EPITHELIAL STEM CELL NICHE

Inflammatory Bowel Diseases ◽

10.1093/ibd/izaa347.018 ◽

2021 ◽

Vol 27 (Supplement_1) ◽

pp. S7-S8

Author(s):

Safina Gadeock ◽

Cambrian Liu ◽

Brent Polk

Keyword(s):

Stem Cell ◽

Stem Cell Niche ◽

Mesenchymal Cells ◽

Epithelial Stem Cells ◽

Epithelial Stem Cell ◽

Long Term Treatment ◽

Lgr5 Expression ◽

Cell Niche

Abstract Tumor necrosis factor (TNF) is a highly expressed cytokine in inflammatory bowel disease (IBD). Although TNF can induce colonic epithelial dysfunction and apoptosis, recent studies suggest that TNF signalling promotes epithelial wound repair and stem cell function. Here we investigated the role of TNF receptor 1 (TNFR1) in mediating TNF’s effects on colonic epithelial stem cells, integral to mucosal healing in colitis. We demonstrate that Tnfr1-/- mice exhibit loss in Lgr5 expression (-52%, p<0.02; N=6) compared to wildtype (WT) controls. However, the opposite result was found in vitro, wherein murine Tnfr1-/- colonoids demonstrated a significant increase in Lgr5 expression (66%, p<0.007; N=6) compared to WT colonoids. Similarly, human colonoids treated with an anti-TNFR1 antibody also demonstrated an increase in Lgr5 expression, relative to IgG controls. To resolve the contradiction in the in vivo versus in vitro environment, we hypothesized that mesenchymal TNFR1 expression regulates the epithelial stem cell niche. To determine the relationships between these cell types, we co-cultured WT or Tnfr1-/- colonoids with WT or Tnfr1-/- colonic myofibroblasts (CMFs). We found that epithelial Lgr5 expression was significantly higher (by 52%, p<0.05; N=3) when co-cultured with WT compared to TNFR1-/- myofibroblasts. The loss of TNFR1 expression in vivo increases the number of αSMA+ mesenchymal cells by nearly 56% (N=6) but considerably reduces the pericryptal PDGFRα+ cells, suggesting modifications in mesenchymal populations that contribute to the epithelial stem cell niche. Functionally, primary Tnfr1-/--CMFs displayed PI3k (p<0.001; N=3) and MAPK (p<0.01; N=3)-dependent increases in migration, proliferation, and differentiation, but RNA profiling demonstrated by diminished levels of stem cell niche factors, Rspo3 (-80%, p<0.0001; N=6) and Wnt2b (-63%, p<0.008; N=6) compared to WT-CMFs. Supplementation with 50ng recombinant Rspo3 for 5 d to Lgr5-GFP organoids co-cultured with TNFR1-/--CMFs restored Lgr5 expression to wildtype levels. Therefore, TNFR1-mediated TNF signalling in mesenchymal cells promotes their ability to support an epithelial stem cell niche. These results should motivate future studies of the stem cell niche in the context of long-term treatment with anti-TNF therapies.

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Closer to Nature: The Role of MSCs in Recreating the Microenvironment of the Hematopoietic Stem Cell Niche in vitro

Transfusion Medicine and Hemotherapy ◽

10.1159/000520932 ◽

2022 ◽

pp. 1-10

Author(s):

Patrick Wuchter ◽

Anke Diehlmann ◽

Harald Klüter

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Stem Cell Niche ◽

Model Systems ◽

Bone Marrow Niche ◽

Hematopoietic Stem ◽

Hematopoietic Stem Cell Niche ◽

Cell Niche

Background: The stem cell niche in human bone marrow provides scaffolds, cellular frameworks and essential soluble cues to support the stemness of hematopoietic stem and progenitor cells (HSPCs). To decipher this complex structure and the corresponding cellular interactions, a number of in vitro model systems have been developed. The cellular microenvironment is of key importance, and mesenchymal stromal cells (MSCs) represent one of the major cellular determinants of the niche. Regulation of the self-renewal and differentiation of HSPCs requires not only direct cellular contact and adhesion molecules, but also various cytokines and chemokines. The C-X-C chemokine receptor type 4/stromal cell-derived factor 1 axis plays a pivotal role in stem cell mobilization and homing. As we have learned in recent years, to realistically simulate the physiological in vivo situation, advanced model systems should be based on niche cells arranged in a three-dimensional (3D) structure. By providing a dynamic rather than static setup, microbioreactor systems offer a number of advantages. In addition, the role of low oxygen tension in the niche microenvironment and its impact on hematopoietic stem cells need to be taken into account and are discussed in this review. Summary: This review focuses on the role of MSCs as a part of the bone marrow niche, the interplay between MSCs and HSPCs and the most important regulatory factors that need to be considered when engineering artificial hematopoietic stem cell niche systems. Conclusion: Advanced 3D model systems using MSCs as niche cells and applying microbioreactor-based technology are capable of simulating the natural properties of the bone marrow niche more closely than ever before.

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Isolation and characterization of human CD34−Lin− and CD34+Lin− hematopoietic stem cells using cell surface markers AC133 and CD7

Blood ◽

10.1182/blood.v95.9.2813.009k20_2813_2820 ◽

2000 ◽

Vol 95 (9) ◽

pp. 2813-2820 ◽

Cited By ~ 215

Author(s):

Lisa Gallacher ◽

Barbara Murdoch ◽

Dongmei M. Wu ◽

Francis N. Karanu ◽

Mike Keeney ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Surface Markers ◽

Cell Surface Markers ◽

Hematopoietic Stem ◽

Human Cd34 ◽

Cd34 Cells

Recent evidence indicates that human hematopoietic stem cell properties can be found among cells lacking CD34 and lineage commitment markers (CD34−Lin−). A major barrier in the further characterization of human CD34− stem cells is the inability to detect this population using in vitro assays because these cells only demonstrate hematopoietic activity in vivo. Using cell surface markers AC133 and CD7, subfractions were isolated within CD34−CD38−Lin− and CD34+CD38−Lin− cells derived from human cord blood. Although the majority of CD34−CD38−Lin− cells lack AC133 and express CD7, an extremely rare population of AC133+CD7− cells was identified at a frequency of 0.2%. Surprisingly, these AC133+CD7− cells were highly enriched for progenitor activity at a frequency equivalent to purified fractions of CD34+ stem cells, and they were the only subset among the CD34−CD38−Lin− population capable of giving rise to CD34+ cells in defined liquid cultures. Human cells were detected in the bone marrow of non-obese/severe combined immunodeficiency (NOD/SCID) mice 8 weeks after transplantation of ex vivo–cultured AC133+CD7− cells isolated from the CD34−CD38−Lin− population, whereas 400-fold greater numbers of the AC133−CD7− subset had no engraftment ability. These studies provide novel insights into the hierarchical relationship of the human stem cell compartment by identifying a rare population of primitive human CD34− cells that are detectable after transplantation in vivo, enriched for in vitro clonogenic capacity, and capable of differentiation into CD34+ cells.

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Enhancement of Megakaryocyte Interactions with the Osteoblastic Hematopoietic Stem Cell Niche Improves Engraftment Efficiency Following Hematopoietic Stem Cell Transplantation

Blood ◽

10.1182/blood.v120.21.211.211 ◽

2012 ◽

Vol 120 (21) ◽

pp. 211-211

Author(s):

Timothy S. Olson ◽

Satoru Otsuru ◽

Ted Hofmann ◽

Edwin M. Horwitz

Keyword(s):

Stem Cell ◽

Growth Factor ◽

Hematopoietic Stem Cell ◽

Stem Cell Niche ◽

Structural Changes ◽

Conflicts Of Interest ◽

Imatinib Treatment ◽

Hematopoietic Stem ◽

Cell Niche

Abstract Abstract 211 Bone marrow (BM) radioablation produces structural changes in the endosteal osteoblastic stem cell niche, a critical site of hematopoietic stem cell (HSC) engraftment following HSC transplantation (HSCT). We have previously shown that total body irradiation (TBI) in wildtype (WT) mice induces migration of recipient megakaryocytes to the niche and an expansion of niche osteoblasts that supports HSC engraftment following transplantation. We have also demonstrated that c-MPL-deficient (mpl−/−) recipients have decreases in total megakaryocytes (35% of WT), the percentage of megakaryocytes migrating to the endosteum (<20% of WT), and niche osteoblast expansion (<50% of WT) following TBI, leading to profound deficits in long-term (LT)-HSC engraftment following HSCT. We now present data examining mechanisms by which megakaryocytes facilitate both niche osteoblast expansion post-TBI and donor HSC engraftment following HSCT, and a therapeutic strategy utilizing these mechanisms to enhance donor HSC engraftment. The decrease in total megakaryocytes and absent thrombopoietin (TPO) signaling in mpl−/− mice resulted in a 90% reduction in post-TBI mpl−/− versus WT BM levels of platelet-derived growth factor beta (PDGFβ), a known osteoblast growth factor. In vitro, megakaryocytes cultured together or across a transwell membrane markedly enhanced osteoblast growth (> 2.5 fold, p < 0.001), but PDGFβ signaling inhibition completely abrogated megakaryocyte-driven osteoblast growth. In vivo, inhibition of PDGF receptor signaling in WT mice via imatinib treatment resulted in near complete blockade of TBI-induced osteoblast expansion, and imatinib treatment of primary recipients resulted in diminished LT-HSC engraftment in secondary transplant assays. Blockade of CD41 integrin-mediated adhesion of megakaryocytes in WT recipient BM blocked TBI-induced megakaryocyte migration to the endosteal niche and severely abrogated LT-HSC engraftment efficiency. However, in contrast to c-MPL deficiency, CD41 blockade did not decrease PDGFβ expression or niche osteoblast expansion, suggesting that in addition to PDGFβ-dependent effects on niche expansion, the megakaryocyte migration to the niche itself is also required to efficiently engraft HSC. Mice with decreased GATA-1 expression (Gata-1tm2sho/J), have a large increase in total BM megakaryocytes a >2-fold (p < 0.001) increase in PDGFβ levels, and greatly increased expansion of osteoblast and other mesenchymal elements 48 hours post-TBI compared to WT mice. However, Gata-1tm2sho/J megakaryocytes have known defective terminal differentiation and function including decreased platelet production, and Gata-1tm2sho/J primary recipients did not engraft LT-HSC more efficiently than WT primary recipients, demonstrating the need for fully functional megakaryocytes, and not only increased PDGFβ-induced mesenchymal proliferation, to foster HSC engraftment. Finally, we have examined whether TPO administration prior to radioablation and HSCT can enhance host megakaryocyte effects on the niche and HSC engraftment. TPO administration for 5 days prior to radioablation, resulted in a significant increase in BM megakaryocytes and a 50% increase in niche osteoblast expansion. Furthermore, competitive secondary transplantation assays demonstrated that TPO- versus sham-treatment of primary recipients prior to TBI and BM transplant, resulted in increased initial engraftment at 24 hours post-primary transplant (40% increase, p < 0.05) increased short-term HSC and progenitor engraftment 3–6 weeks following secondary transplant (4–20 fold increase, p < 0.02), and sustained LT-HSC engraftment at 28 weeks post-transplant in 47% versus 7% (p < 0.05) of secondary recipients of TPO- versus sham-treated primary recipient BM, respectively. Taken together, our results demonstrate that host megakaryocytes facilitate efficient HSC engraftment following TBI and HSCT through PDGFβ-dependent enhancement of niche osteoblast expansion and through direct interactions of megakaryocytes with the niche. TPO-treatment of transplant recipients prior to radioablation and stem cell infusion enhances these megakaryocyte-dependent pathways and subsequent donor HSC engraftment efficiency, providing a clinically applicable strategy to enhance niche function and stem cell engraftment following clinical transplantation. Disclosures: No relevant conflicts of interest to declare.

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Ectopic Human Mesenchymal Stem Cell-Coated Scaffolds Create An In Vivo Leukemia Niche in NOD/SCID Mice.

Blood ◽

10.1182/blood.v114.22.559.559 ◽

2009 ◽

Vol 114 (22) ◽

pp. 559-559

Author(s):

Sarah Rivkah Vaiselbuh ◽

Morris Edelman ◽

Jeffrey Michael Lipton ◽

Johnson M. Liu

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Stem Cell ◽

Stem Cell Niche ◽

Scid Mice ◽

Cxcr4 Antagonist ◽

Cell Niche

Abstract Abstract 559 Introduction: Different cellular components of the normal hematopoietic niche have been identified. However, the niche for malignant hematopoiesis remains to be elucidated. Recent work of other groups has suggested that hematopoietic stem cells (HSC) within the bone marrow anchor themselves in place by attaching to osteoblasts and/or vascular sinusoid endothelial cells. We have recently identified mesenchymal stem cells (MSC) as niche-maker cells and found a crucial role of the SDF-1/CXCR4 axis in this process. Stromal Derived Factor-1 (SDF-1/CXCL12) regulates stem cell trafficking and the cell cycle via its receptor CXCR4. Methods: Polyurethane scaffolds, coated in vitro with human bone marrow MSC, were implanted subcutaneously in non-irradiated NOD/SCID mice. CD34+ HSC or primary AML cells (from a leukapheresis product) were injected either in situ or retro-orbitally in the mice and analyzed for engraftment. The mice were treated twice per week with in situ injections of SDF-1, AMD3100 (a CXCR4 antagonist) or PBS (control). After 2 to 4 weeks, the scaffolds were processed and evaluated for cell survival in the mesenchymal niche by immunohistochemistry. Results: We created in vitro MSC-coated scaffolds that retained inoculated AML cells in the presence of SDF-1, while AML cells seeded on empty scaffolds were not retained. In vivo in NOD/SCID mice, the MSC-coated scaffolds, in the presence of SDF-1 enabled homing of both in situ injected normal CD34+ HSC and retroorbital- or in situ injected primary human AML cells. The scaffolds were vascularized and showed osteoclasts and adipocytes present, suggestive of an ectopic human bone marrow microenvironment in the murine host. Finally, the SDF-1-treated scaffolds showed proliferation of the MSC stromal layer with multiple adherent AML cells, while in the AMD3100-treated scaffolds the stromal lining was thin and disrupted at several points, leaving AML cells free floating in proximity. The PBS-treated control-scaffold showed a thin single cell MSC stromal layer without disruption, with few AML cells attached. Conclusion: The preliminary data of this functional ectopic human microenvironment in NOD/SCID mice suggest that AMD3100 (a CXCR4 antagonist) can disrupt the stem cell niche by modulation of the mesenchymal stromal. Further studies are needed to define the role of mesenchymal stem cells in maintaining the hematopoietic/leukemic stem cell niche in vivo. In Vivo Leukemia Stem Cell Niche: (A) Empty polyurethane scaffold. (B)Vascularization in SQ implanted MSC-coated scaffold (s) niche in NOD/SCID mice. (C) DAB Peroxidase (brown) human CD45 positive nests of AML cells (arrows) 1 week after direct in situ AML injection. (D) Human CD45 positive myeloid cells adhere to MSC in vivo (arrows). Disclosures: No relevant conflicts of interest to declare.

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Mir-886-3p Contributes to the Regulation of the hematopoietic microenvironment by down-Regulating SDF-1α (CXCL12).

Blood ◽

10.1182/blood.v114.22.561.561 ◽

2009 ◽

Vol 114 (22) ◽

pp. 561-561

Author(s):

Manoj M Pillai ◽

Xiaodong Yang ◽

Mineo Iwata ◽

Lynne Bemis ◽

Beverly Torok-Storb

Keyword(s):

Gene Expression ◽

Stem Cell ◽

Stromal Cells ◽

Stem Cell Niche ◽

Expression Profiles ◽

Hematopoietic Stem ◽

Cd34 Cells ◽

Proliferation And Differentiation ◽

Cell Niche ◽

Angiopoietin 1

Abstract Abstract 561 Two functionally distinct stromal cell lines were isolated from a primary long term culture (LTC) established from aspirated human marrow. Designated HS5 and HS27a, the lines were immortalized and extensively characterized including expression profiles for both messenger (mRNA) and micro-RNA (miRNA, a recently described class of small non-coding RNAs that regulate gene expression by binding to target mRNAs to prevent their translation). HS5 was found to secrete growth factors that stimulate proliferation and differentiation of hematopoietic progenitors (G-CSF, IL-6, IL-1α and IL1β), whereas HS27a expresses activities associated with the stem cell niche (SDF-1αa, angiopoietin-1 etc). In keeping with this HS5 conditioned media stimulated proliferation and differentiation of isolated CD34+ cells whereas HS27a supported CD34+ cells in an undifferentiated state. When cultured together to better mimic in vivo cell-cell interactions, the gene expression of HS27a and HS5 combined differed from the expected sum of the two parts, exemplified by the 5-fold down regulation of SDF-1α. Comparisons of miRNA expression profiles of HS5 and HS27a determined that mir-886-3p, (previously described by deep sequencing of small RNA libraries) was expressed > 40 fold in HS5 compared to HS27a, this was then confirmed by quantitative RT-PCR. Given the abundance of mir-886-3p and the possibility that it could be secreted by HS5 and taken up by cells in contact with HS5, we tested its effect on gene expression in HS27a. Transcript levels of genes associated with the stem cell niche (Jagged1, BMP4, Angiopoietin-1, SDF-1α, VCAM-1 and N-Cadherin) were determined by quantitative RT-PCR after direct transfection of mir-886-3p precursors into HS27a cells and compared to appropriate controls. Results show SDF-1α mRNA expression was down-regulated by as much as 8 fold 3 days after transfection. Levels of secreted SDF-1α in culture media, as determined by ELISA, were also decreased. Since SDF-1α is a chemokine known to be critical for the homing of hematopoietic stem and progenitor cells to their niche, the functional significance of the SDF-1α down-regulation by mir-886-3p was confirmed by decreased chemotaxis of T-lymphocytic cells (Jurkat) following miRNA transfection of stromal cells. To determine if mir-886-3p directly effects the SDF-1α transcript, the 1.5 kbp 3'untranslated region (UTR) of SDF-1α gene was cloned downstream of the luciferase gene, and co-transfected with mir-886-3p into HS27a cells. Results showed the luciferase activity was down-regulated greater than 50% in the presence of mir-886-3p, suggesting a direct effect on the SDF-1 α transcript. Given the concern over the relevance of immortalized cell lines we investigated Mir-886-3p expression in primary marrow stromal cells at early passage sorted on the basis of +/- expression of CD146. (CD146 or MCAM has been reported to define a population that supports the hematopoietic stem/ precursor cell niche and is expressed by HS27a and not HS5 cells). Results indicated that the CD146+ stromal cells had significantly lower expression of mir-886-3p when compared to CD146- cells. In summary, these data suggest a role for miRNA in modulating the expression of gene products that are associated with the hematopoietic stem cell niche. Disclosures: No relevant conflicts of interest to declare.

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Screen for Small Molecules Capable of Expanding Human Hematopoietic Stem Cell Ex Vivo

Blood ◽

10.1182/blood.v118.21.1919.1919 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1919-1919

Author(s):

Iman Hatem Fares ◽

Jalila Chagraoui ◽

Jana Krosl ◽

Denis-Claude Roy ◽

Sandra Cohen ◽

...

Keyword(s):

Stem Cell ◽

Hematopoietic Stem Cell ◽

Mononuclear Cells ◽

Conflicts Of Interest ◽

Ex Vivo ◽

Fold Increase ◽

Hematopoietic Stem ◽

Cd34 Cells

Abstract Abstract 1919 Hematopoietic stem cell (HSC) transplantation is a life saving procedure whose applicability is restricted by the lack of suitable donors, by poor responsiveness to mobilization regimens in preparation of autologous transplantations, by insufficient HSC numbers in individual cord blood units, and by the inability to sufficiently amplify HSCs ex vivo. Characterization of Stemregenin (SR1), an aryl hydrocarbon receptor (AHR) antagonist that promotes HSC expansion, provided a proof of principle that low molecular weight (LMW) compounds have the ability to promote HSC expansion. To identify novel putative agonists of HSC self-renewal, we initiated a high throughput screen (HTS) of a library comprising more than 5,000 LMW molecules using the in vitro maintenance of the CD34+CD45RA- phenotype as a model system. Our study was based on the fact that mobilized peripheral blood-derived CD34+CD45RA- cells cultured in media supplemented with: stem cell factor, thrombopoietin, FLT3 ligand and interleukin 6, would promote the expansion of mononuclear cells (MNC) concomitant with a decrease in CD34+CD45RA- population and HSC depletion. LMW compounds preventing this loss could therefore act as agonists of HSC expansion. In a 384-well plate, 2000 CD34+cells were initially cultured/well in 50μl medium comprising 1μM test compounds or 0.1% DMSO (vehicle). The proportions of CD34+CD45RA− cells were determined at the initiation of experiment and after a 7-day incubation. Six of 5,280 LMW compounds (0.11%) promoted CD34+CD45RA− cell expansion, and seventeen (0.32%) enhanced differentiation as determined by the increase in proportions of CD34−CD45RA+ cells compared to control (DMSO). The 6 LMW compounds promoting expansion of the CD34+CD45RA− cell population were re-analyzed in a secondary screen. Four out of these 6 molecules suppressed the transcriptional activity of AHR, suggesting that these compounds share the same molecular pathway as SR1 in stimulating HSC expansion, thus they were not further characterized. The remaining 2 compounds promoted, similar to SR1 or better, a 10-fold and 35-fold expansion of MNC during 7 and 12-day incubations, respectively. The expanded cell populations comprised 65–75% of CD34+ cells compared to 12–30% determined for DMSO controls. During 12-day incubation with these compounds, the numbers of CD34+ cells increased ∼25-fold over their input values, or ∼ 6-fold above the values determined for controls. This expansion of CD34+ cells was associated with a ∼5-fold increase in the numbers of multilineage CFC (granulocyte, erythroid, monocyte, and megakaryocyte, or CFU-GEMM) compared to that found in DMSO control cultures. The ability of the 2 newly identified compounds to expand functional HSCs is currently being evaluated in vivo usingimmunocompromised mice. In conclusion, results of our initial screen suggest that other mechanism, besides inhibition of AhR, are at play for expansion of human HSC. Disclosures: No relevant conflicts of interest to declare.

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Intestinal Stem Cell Niche Insights Gathered from Both In Vivo and Novel In Vitro Models

Stem Cells International ◽

10.1155/2017/8387297 ◽

2017 ◽

Vol 2017 ◽

pp. 1-10 ◽

Cited By ~ 10

Author(s):

Nikolce Gjorevski ◽

Paloma Ordóñez-Morán

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Stem Cell Niche ◽

Cell Function ◽

Complex Structure ◽

Intestinal Stem Cell ◽

Intestinal Damage ◽

Cell Niche

Intestinal stem cells are located at the base of the crypts and are surrounded by a complex structure called niche. This environment is composed mainly of epithelial cells and stroma which provides signals that govern cell maintenance, proliferation, and differentiation. Understanding how the niche regulates stem cell fate by controlling developmental signaling pathways will help us to define how stem cells choose between self-renewal and differentiation and how they maintain their undifferentiated state. Tractable in vitro assay systems, which reflect the complexity of the in vivo situation but provide higher level of control, would likely be crucial in identifying new players and mechanisms controlling stem cell function. Knowledge of the intestinal stem cell niche gathered from both in vivo and novel in vitro models may help us improve therapies for tumorigenesis and intestinal damage and make autologous intestinal transplants a feasible clinical practice.

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Characterization of substantia nigra neurogenesis in homeostasis and dopaminergic degeneration: beneficial effects of the microneurotrophin BNN-20

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02398-3 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Theodora Mourtzi ◽

Dimitrios Dimitrakopoulos ◽

Dimitrios Kakogiannis ◽

Charalampos Salodimitris ◽

Konstantinos Botsakis ◽

...

Keyword(s):

Stem Cell ◽

Substantia Nigra ◽

Mouse Model ◽

Dopaminergic Neurons ◽

Stem Cell Niche ◽

Substantia Nigra Pars Compacta ◽

Dopaminergic Cell ◽

Cell Niche

Abstract Background Loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) underlines much of the pathology of Parkinson’s disease (PD), but the existence of an endogenous neurogenic system that could be targeted as a therapeutic strategy has been controversial. BNN-20 is a synthetic, BDNF-mimicking, microneurotrophin that we previously showed to exhibit a pleiotropic neuroprotective effect on the dopaminergic neurons of the SNpc in the “weaver” mouse model of PD. Here, we assessed its potential effects on neurogenesis. Methods We quantified total numbers of dopaminergic neurons in the SNpc of wild-type and “weaver” mice, with or without administration of BNN-20, and we employed BrdU labelling and intracerebroventricular injections of DiI to evaluate the existence of dopaminergic neurogenesis in the SNpc and to assess the origin of newborn dopaminergic neurons. The in vivo experiments were complemented by in vitro proliferation/differentiation assays of adult neural stem cells (NSCs) isolated from the substantia nigra and the subependymal zone (SEZ) stem cell niche to further characterize the effects of BNN-20. Results Our analysis revealed the existence of a low-rate turnover of dopaminergic neurons in the normal SNpc and showed, using three independent lines of experiments (stereologic cell counts, BrdU and DiI tracing), that the administration of BNN-20 leads to increased neurogenesis in the SNpc and to partial reversal of dopaminergic cell loss. The newly born dopaminergic neurons, that are partially originated from the SEZ, follow the typical nigral maturation pathway, expressing the transcription factor FoxA2. Importantly, the pro-cytogenic effects of BNN-20 were very strong in the SNpc, but were absent in other brain areas such as the cortex or the stem cell niche of the hippocampus. Moreover, although the in vitro assays showed that BNN-20 enhances the differentiation of NSCs towards glia and neurons, its in vivo administration stimulated only neurogenesis. Conclusions Our results demonstrate the existence of a neurogenic system in the SNpc that can be manipulated in order to regenerate the depleted dopaminergic cell population in the “weaver” PD mouse model. Microneurotrophin BNN-20 emerges as an excellent candidate for future PD cell replacement therapies, due to its area-specific, pro-neurogenic effects.

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Matrix Glycoprotein Osteopontin Is a Stem Cell Niche Constituent That Constrains the Hematopoietic Stem Cell Pool Size.

Blood ◽

10.1182/blood.v104.11.664.664 ◽

2004 ◽

Vol 104 (11) ◽

pp. 664-664 ◽

Cited By ~ 2

Author(s):

Sebastian Stier ◽

Yon Ko ◽

Randolf Forkert ◽

Christoph Lutz ◽

Thomas Neuhaus ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cell ◽

Stem Cell Niche ◽

Wild Type ◽

Hematopoietic Stem ◽

Hematopoietic Stem Cell Niche ◽

Cell Niche ◽

Stem Cell Pool

Abstract Stem cells reside in a physical niche where a balance of signals controls their growth, differentiation and death. Niche components have generally been defined in terms of cells and positive effects on stem cell maintenance or expansion. Here we define a role for a matrix glycoprotein that provides a constraining function in the hematopoietic stem cell niche. Osteopontin (OPN) is an abundant glycoprotein in bone that can function as either cytokine or cell adhesion mediator. It is known to be produced by multiple cells types including osteoblasts, cells recently defined to be a regulatory component of the hematopoietic stem cell niche. Using studies combining OPN deficient mice and exogenous OPN, we demonstrate that OPN modifies primitive hematopoietic cell numbers and function. In OPN deficient mice, increased primitive cell numbers were observed in vivo associated with reduced progenitors and reduced primitive cell apoptotic fraction. To determine whether the effect of OPN deficiency was stroma dependent, we performed in vitro stem cell assays on OPN−/− stroma and observed greater LTC-IC supportive capacity compared with wild type stroma. Furthermore, OPN−/− recipients showed a significantly higher proportion of hematopoietic stem cells after transplantation of OPN+/+ bone marrow in comparison to wild-type recipients, indicating that the OPN null microenvironment was sufficient to increase stem cell number. A reduction in apoptotic fraction was seen in primitive cells in the OPN−/− recipient marrows. A role for OPN in apoptosis was confirmed by exogenous OPN in in-vitro studies. Hypothesizing that OPN may serve as a physiologic constraint on stem cell pool size, we compared OPN−/− with wild type animals following parathyroid hormone activation of the stem cell niche. The expansion of stem cells by PTH was superphysiologic in the absence of OPN. Therefore, OPN is a restricting element of the stem cell niche, limiting the number of stem cells produced by niche activation. Extracellular matrix components such as OPN may serve as modulable, regulatory participants in the stem cell niche.

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