scholarly journals Cell Tracking of Adult Neural Stem Cells From The Subependymal Zone In Vitro Reveals Cell Type-Specific Impacts On Cell Cycle Progression By The Extracellular Matrix Molecule Tenascin-C

2021 ◽  
Author(s):  
Elena Schaberg ◽  
Magdalena Götz ◽  
Andreas Faissner
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Stem Cell Niche ◽  
Cycle Length ◽  
Cell Tracking ◽  
Subependymal Zone ◽  
Cell Niche ◽  
Lineage Trees ◽  
Cell Cycle Length

Abstract Adult neurogenesis has been described in two canonical regions of the adult central nervous system (CNS) of rodents, the subgranular zone (SGZ) of the hippocampus and the subependymal zone (SEZ) of the lateral ventricles. The stem cell niche of the SEZ provides a privileged environment composed of a specialized extracellular matrix (ECM) that comprises the glycoproteins tenascin-C (Tnc) and laminin-1 (LN1). In the present study, we investigated the function of these ECM glycoproteins in the adult stem cell niche. Adult neural stem cells (aNSPCs) of the SEZ were prepared from wildtype (Tnc+/+) and Tnc knockout (Tnc-/-) mice and analyzed using molecular and cell biological approaches. A delayed maturation of aNSPCs in Tnc-/- tissue was reflected by a reduced capacity to form neurospheres in response to epidermal growth factor (EGF). In order to examine a potential influence of the ECM on cell proliferation, aNSPCs of both genotypes were studied by cell tracking using digital video microscopy. aNSPCs were cultivated on three different substrates, namely poly-D-Lysine (PDL) and PDL replenished with either LN1 or Tnc for up to six days in vitro. On each of the three substrates aNSPCs displayed lineage trees that could be investigated with regard to cell cycle length. The latter appeared reduced in Tnc-/- aNSPCs on PDL and LN1 substrates, less so on Tnc that seemed to compensate the absence of the ECM compound to some extent. Close inspection of the lineage trees revealed a subpopulation of late dividing aNSPCslate that engaged into cycling after a notable delay. aNSPCslate exhibited a clearly different morphology, with a larger cell body and conspicuous processes. aNSPCslate reiterated the reduction in cell cycle length on all substrates tested, which was not rescued on Tnc substrates. When the migratory activity of aNSPC-derived progeny was determined, Tnc-/- neuroblasts displayed significantly longer migration tracks. This was traced to an increased rate of migration episodes compared to the wildtype cells that rested for longer time periods. We conclude that Tnc intervenes in the proliferation of aNSPCs and modulates the motility of neuroblasts in the niche of the SEZ.

scholarly journals Live-cell time-lapse imaging and single-cell tracking of in vitro cultured neural stem cells – Tools for analyzing dynamics of cell cycle, migration, and lineage selection

Methods ◽  
2018 ◽  
Vol 133 ◽  
pp. 81-90 ◽  
Author(s):  
Katja M. Piltti ◽  
Brian J. Cummings ◽  
Krystal Carta ◽  
Ayla Manughian-Peter ◽  
Colleen L. Worne ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Neural Stem Cells ◽  
Single Cell ◽  
Cell Tracking ◽  
Time Lapse ◽  
Live Cell ◽  
Time Lapse Imaging

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

Blood ◽  
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.

scholarly journals Cell cycle length, cell size, and proliferation rate in hydra stem cells

1990 ◽  
Vol 142 (2) ◽  
pp. 392-400 ◽  
Author(s):  
Thomas W. Holstein ◽  
Charles N. David
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Cell Size ◽  
Cycle Length ◽  
Proliferation Rate ◽  
Cell Cycle Length

scholarly journals Renal capsule as a stem cell niche

2010 ◽  
Vol 298 (5) ◽  
pp. F1254-F1262 ◽  
Author(s):  
Hyeong-Cheon Park ◽  
Kaoru Yasuda ◽  
Mei-Chuan Kuo ◽  
Jie Ni ◽  
Brian Ratliff ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Stem Cell Niche ◽  
Cultured Cells ◽  
Renal Parenchyma ◽  
Renal Tubules ◽  
Stem Cell Markers ◽  
Renal Capsule ◽  
Cell Niche

Renal resident stem cells were previously reported within the renal tubules and papillary area. The aim of the present study was to determine whether renal capsules harbor stem cells and whether this pool can be recruited to the renal parenchyma after ischemic injury. We demonstrated the presence of label-retaining cells throughout the renal capsule, at a density of ∼10 cells/mm2, and their close apposition to the blood vessels. By flow cytometry, in vitro cultured cells derived from the renal capsule were positive for mesenchymal stem cell (MSC) markers (CD29+, vimentin+, Sca-1+, nestin+) but did not express hematopoietic and endothelial stem cell markers. Moreover, renal capsule-derived cells also exhibited self-renewal, clonogenicity, and multipotency in differentiation conditions, all favoring stem cell characteristics and identifying them with MSC. In situ labeling of renal capsules with CM-DiI CellTracker demonstrated in vivo a directed migration of CM-DiI-labeled cells to the ischemic renal parenchyma, with the rate of migration averaging 30 μm/h. Decapsulation of the kidneys during ischemia resulted in a modest, but statistically significant, deceleration of recovery of plasma creatinine compared with ischemic kidneys with intact renal capsule. Comparison of these conditions allows the conclusion that renal capsular cells may contribute ∼25–30% of the recovery from ischemia. In conclusion, the data suggest that the renal capsule may function as a novel stem cell niche harboring MSC capable of participating in the repair of renal injury.

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

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
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.

scholarly journals Phase register of Hes1 oscillations with mitoses underlies cell-cycle heterogeneity in ER+ breast cancer cells

2021 ◽  
Author(s):  
Nitin Sabherwal ◽  
Andrew Rowntree ◽  
Jochen Kursawe ◽  
Nancy Papalopulu
Keyword(s):  
Breast Cancer ◽  
Cell Cycle ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Cancer Cells ◽  
Cycle Length ◽  
Breast Cancer Cells ◽  
Transcriptional Repressor ◽  
Hes1 Expression ◽  
Cell Cycle Length

AbstractHere, we study the dynamical expression of endogenously labelled Hes1, a transcriptional repressor implicated in controlling cell proliferation, to understand how cell-cycle length heterogeneity is generated in ER+ breast cancer cells. We find that Hes1 shows oscillatory expression and during the cell-cycle has a variable peak in G1, a trough around G1-S transition and a less variable second peak in G2/M. Compared to other subpopulations, the cell-cycle in CD44HighCD24Low cancer stem cells is longest and most variable. Most cells divide around the peak of the Hes1 expression wave but mitoses in slow dividing CD44HighCD24Low cells appear phase-shifted, resulting in a late-onset Hes1 peak in G1. The position, duration and shape of this peak, rather than the Hes1 expression levels, are good predictors of cell-cycle length. Diminishing Hes1 oscillations impairs proliferation, indicating their functional importance for efficient cell-cycle progression. We propose that the position of mitosis in relation to the Hes1 wave underlies cell-cycle length heterogeneity in cancer cell subpopulations.Significance statementTumours exhibit heterogeneities that are not due to mutations, including Cancer Stem Cells with different potencies. We show that the cancer stem cell state predisposed to dormancy in vivo has a highly variable and long cell-cycle. Using single-cell live-imaging for the transcriptional repressor Hes1 (a key molecule in cancer), we show a new type of oscillatory expression of Hes1 in all cells in the population. The most potent cancer stem cells tend to divide around the trough of the Hes1 oscillatory wave, a feature predictive of a long cell-cycle. A novel concept proposed here is that the position that a cell is with respect to the Hes1 wave when it divides is predictive of its prospective cell-cycle length and characteristic of its cellular sub-state.Abstract in picture

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

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 664-664 ◽  
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.

Thrombopoietin-Mpl Signal Induces Hematopoietic Stem Cells into Quiescent State in the Bone Marrow Niche.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 858-858
Author(s):  
Hiroki Yoshihara ◽  
Fumio Arai ◽  
Kentaro Hosokawa ◽  
Takao Takahashi ◽  
Hiroshi Miyazaki ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Stem Cell ◽  
Stem Cell Niche ◽  
Osteoblastic Cells ◽  
Bone Surface ◽  
Hematopoietic Stem ◽  
Cycle Arrest ◽  
Cell Niche

Abstract Hematopoietic stem cells (HSCs) have the ability to self-renew and to differentiate to produce multi-lineage blood cells throughout the lifetime of individuals. Maintenance of these stem cell activities depends on the balance of intrinsic and extrinsic factors. The factors which regulate HSCs are provided by a microenvironment called stem cell niche, and interactions between HSCs and stem cell niche is critical for the maintenance of stem cell activities. Recently, it has been reported that HSCs exist frequently aside of the trabecular bone surface in bone marrow (BM). We have previously reported that side-population (SP) in HSC fraction is in the G0 phase and anti-apoptotic stem cells, and contacts osteoblasts. (Arai et al., Cell 2004). To further investigate the regulation of quiescence, cell adhesion, and survival of HSCs, we tried to clone the quiescent HSCs specific molecules by microarray analysis of c-Kit+Sca-1+Lin− (KSL)-SP vs. non-SP cells. We identified that Mpl, thrombopoietin (Tpo) receptor, was highly expressed in SP cells compared to non-SP cells. Tpo/Mpl signal is known as a physiological regulator of megakaryopoiesis, but the role of Tpo/Mpl signal in the maintenance of HSCs remains elusive. In this study, we investigated the role of Tpo/Mpl signal on the regulation of HSCs in the niche. The frequency of Mpl+ cells in KSL, KSL-SP, and KSL-non SP cells were 50.4 %, 88.9 %, and 44.7 %, respectively. In addition, Tpo was expressed in osteoblastic cells in BM. Immunohistochemical staining of BM showed that Mpl+ HSCs adhered to the bone surface and bone-lining osteoblastic cells produced Tpo. These data suggest that Tpo/Mpl signal contributes to the HSCs-niche interaction. BM transplantation (BMT) assay demonstrated that Mpl+KSL cells showed high long-term reconstitution (LTR)-activity, whereas Mpl−KSL cells did not, suggesting that LTR-HSCs were enriched in Mpl+ fraction. To investigate the function of Tpo/Mpl signal in HSCs, we performed CAFC assay and LTC-IC assay in the presence of anti-Mpl neutralizing antibody (AMM2). Inhibition of Tpo/Mpl signal reduced cobblestone formation and reduced LTC-IC formation. These data suggest that Tpo/Mpl signal maintained immature phenotypes of HSCs in vitro. It was reported that Mpl deficient mice showed the defect of stem cell function. For rapid and transient inhibition of Mpl signaling in vivo, we administrated AMM2 into the adult mice. AMM2 did not affect the frequency of non-SP fraction, but transiently decreased frequency of SP in KSL after 6 days of injection. Moreover, the combination of AMM2 and 5-FU capacitated BMT without irradiation. In contrast, injection of Tpo increased KSL-SP cells. To understand the mechanism of Tpo/Mpl signal, we cultured Mpl+KSL cells in the presence of SCF and/or Tpo, and analyzed the gene expression. We found that Tpo treatment up-regulate β1-integrin and p57, but not p21, p27 or p18. As the up-regulation of p57 is essential for TGF-β induced cell cycle arrest in hematopoiesis, Tpo/Mpl signal may also be related to cell cycle arrest. Altogether, these data suggest that Tpo/Mpl signal regulates HSCs-niche interaction and enhanced the quiescence of HSCs.

Evidence for a Metabolic Stem Cell Niche.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4046-4046 ◽  
Author(s):  
Michael Cross ◽  
Rudiger Alt ◽  
Lydia Schnapke-Hille ◽  
Thomas Riemer ◽  
Dietger Niederwieser
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Stem Cell ◽  
Stem Cell Niche ◽  
Culture Media ◽  
Self Renewal ◽  
Cell Culture Media ◽  
Hematopoietic Stem ◽  
Cell Niche

Abstract The hematopoietic stem cell niche presents a localised environment supporting the balanced maintenance, self-renewal and occasional expansion of the stem cell pool. These options are widely assumed to be regulated exclusively by signalling from specific combinations of stroma-bound or soluble ligands. However, a consideration of the rare conditions under which absolute numbers of stem cells increase in vivo as well as the selective pressures acting on regenerative systems during evolution has led us to propose a metabolic component to the stem cell niche which serves to limit cumulative damage, to avoid the selection of potentially oncogenic mutations and to tie symmetric division to slow proliferation. This would mean that traditional cell culture media based on “systemic” substrates such as glucose and glutamine may actively prevent the symmetric amplification of high quality stem cells, offering a possible explanation for the limited success in this area to date. To investigate this possibility, we have examined the effects of range of carbon and energy sources on the proliferation and maintenance of stem and progenitor cells. Our strategy is to screen a wide variety of culture conditions using murine FDCPmix cells, which are non-tumorigenic but have an innate tendency to amplify symmetrically in the presence of IL-3, and then to test key observations in human UCB CD133+ cells provided with SCF, TPO and FLT-3L. In both cell systems, we do indeed find an unusually low requirement for the systemic substrates glucose and glutamine normally included as major energy and carbon sources in cell culture media. Reducing glucose reduces the yield of committed cells from CD133+ cultures without affecting the accumulation of CD133+CD34+cKit+ progenitors. When provided with alternative substrates more likely to reflect a “niche” type environment, FDCPmix cells can be maintained for long periods in media containing only the trace levels of glucose or glutamine derived from dialysed serum, and show improved self-renewal under these conditions. We have also found that raising osmolarity reduces glucose dependence and simultaneously favours the maintenance both of self-renewing CFU (FDCPmix culture) and of CAFCweek13 (CD133+ culture). In parallel, the use of NMR and mass spectrometry techniques to profile intracellular metabolites in self-renewing and differentiating FDCPmix cells reveals a shift in the metabolite balance indicating reduced glycolysis in the early cells. Taken together, these results suggest that hematopoietic stem cells do indeed have remarkable metabolic characteristics consistent with adaptation to a metabolically limiting niche environment. It may therefore be necessary to identify niche substrates and to combine these with the relevant signalling environment in vitro in order to effectively increase stem cell numbers for research, stem cell transplantation and tissue engineering applications.

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