Characterization of Intercellular Junctional Complexes between Human Hematopoietic and Mesenchymal Stem Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1396-1396
Author(s):  
Patrick Wuchter ◽  
Rainer Saffrich ◽  
Beate Straub ◽  
Judit Boda-Heggemann ◽  
Katrin Miesala ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adherens Junctions ◽  
Cellular Interaction ◽  
Bovine Bone ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Junctional Complexes ◽  
Intercellular Connections

Abstract The interaction between hematopoietic stem cells and their niche is essential for the balance between self-renewal and differentiation. We previously demonstrated that intercellular connections in mesenchymal stem cells (MSC) are realized by occasional gap junctions and frequent adherens junctions, comprising specific cadherin-catenin-complexes. Using MSC-feeder-layer as a surrogate model for the hematopoietic stem cell (HSC) niche, we have analyzed the intercellular junctional complexes between HSC and MSC. MSC were obtained from bone marrow aspirates from healthy voluntary donors. HSC were isolated from umbilical cord blood. Using advanced confocal laser scanning in combination with deconvolution and volume rendering software, we were able to produce 3D-images of intercellular junctions between HSC and MSC. We used a panel of antibodies specific for various components of tight, gap and adherens junctions. Additionally, we compared the data to human and bovine bone marrow tissue in situ. We could show that intercellular connections between HSC and MSC are mainly realized by podia formation of the HSC linking to the adjacent MSC. These podia vary greatly in length and shape (uropodia, filopodia). Along these podia and especially at the contact zone to the MSC, we have identified the cytoplasmic plaque proteins alpha- and beta-catenin and protein p120ctn, as well as the transmembrane glycoprotein N-cadherin. This study provided solid evidence for the direct and intimate cellular interaction of HSCs with their niche. Direct cell contact represents a key factor for the regulation of self-renewal versus differentiation. The examination of the specific function of catenins, p120ctn and N-cadherin in this process is concurrently underway.

Mesenchymal Stem Cells Self-Regulate Their Differentiation To Maintain the Bone Marrow Stromal System.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2335-2335
Author(s):  
Iekuni Oh ◽  
Akira Miyazato ◽  
Hiroyuki Mano ◽  
Tadashi Nagai ◽  
Kazuo Muroi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cell Lines ◽  
Molecular Mechanisms ◽  
Adipocyte Differentiation ◽  
Expression Profiles ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Steady Level

Abstract Mesenchymal stem cells (MSCs) account for a very small population in bone marrow stroma as a non-hematopoietic component with multipotency of differentiation into adipocytes, osteocytes and chondrocytes. MSC-derived cells are known to have hematopoiesis-supporting and immunomodulatory abilities. Although clinical applications of MSCs have already been conducted for the suppression of graft versus host disease in allogeneic stem cell transplantation and for tissue regeneration, underlying mechanisms of the biological events are still obscure. Previously, we established a differentiation model of MSCs using a mouse embryo fibroblast cell line, C3H10T1/2 (10T1/2) (Nishikawa M et al: Blood81:1184–1192, 1993). Preadipocyte (A54) and myoblast (M1601) cell lines were cloned by treatment with 5-azacytidine. A54 cells and M1601 cells can terminally differentiate into adipocytes and myotubes, respectively, under appropriate conditions, while parent 10T1/2 cells remain undifferentiated. Moreover, A54 cells show a higher ability to support hematopoiesis compared with the other cell lines. In this study, we analyzed gene expression profiles of the three cell lines by using DNA microarray and real-time PCR to investigate molecular mechanisms for maintaining immaturity of parent 10T1/2 cells. In A54 cells, 202 genes were up-regulated, including those encoding critical factors for hematopoiesis such as SCF, Angiopoietin-1, and SDF-1 as well as genes known to be involved in adipocyte differentiation such as C/EBPα, C/EBPδ and PPAR-γ genes. These data are consistent with the hematopoiesis-supporting ability of A54 cells. During adipocyte differentiation, SCF and SDF-1 expression levels decreased in A54 cells while C/EBPα expression showed a steady level. Recently, osteoblasts have been reported to play crucial roles in “niche” for self-renewal of hematopoietic stem cells. Our results also implicate that precursor cells of non-hematopoietic components may have important roles for hematopoiesis in bone marrow. Meanwhile, in parent 10T1/2 cells, 105 genes were up-regulated, including CD90, Dlk, Wnt5α and many functionally unknown genes. Although C/EBPα expression was induced in 10T1/2 cells without differentiation under the adipocyte differentiation conditions, CD90 expression decreased, Dlk showed a steady level and Wnt5α was up-regulated. Assuming that some regulatory mechanisms are needed to keep an immature state of parent 10T1/2 cells even under the differentiation-inducible conditions, we performed following experiments. First, enforced Dlk expression in A54 cells did not inhibit terminal differentiation to adipocytes under the differentiation conditions. Second, when we cultured A54 cells in the conditioned media of parent 10T1/2 cells under the differentiation-inducible conditions, adipocyte differentiation was inhibited, suggesting that 10T1/2 cells produce some soluble molecules that can inhibit adipocyte differentiation. Since Wnt family is known to be involved in the regulation of self-renewal of several stem cells, Wnt5α may be one candidate for maintenance of “stemness” of MSCs. Taken together, the data of 10T1/2 cells suggest that MSCs can self-regulate their differentiation in the bone marrow stromal system. This concept may be important to investigate the fatty change of bone marrow in aging and in aplastic anemia.

Molecular Characterization of Unique Junctional Complexes as Communication Pathways among Mesenchymal Stem Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1399-1399 ◽  
Author(s):  
Patrick Wuchter ◽  
Judit Boda-Heggemann ◽  
Beate Straub ◽  
Ulf Krause ◽  
Anja Seckinger ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Light And Electron Microscopy ◽  
Beta Catenin ◽  
Unique Type ◽  
Hematopoietic Stem ◽  
Junctional Complexes ◽  
Adhering Junctions

Abstract The interaction between stem cells and their niche is essential for the balance between self-renewal and differentiation. Mesenchymal stem cells (MSC) from bone marrow are able to differentiate into various tissues, such as bone, cartilage, muscle and fat. We have characterized the intercellular contacts among MSC and previously demonstrated the occurrence of a novel kind of adhering junction, consisting of villiform-to-vermiform cell projections (processus adhaerentes). The molecular compositions of these junctions have now been elucidated. Using a panel of antibodies specific for various components of tight, gap and adhering junctions (the latter comprising desmosomes and adhering junctions), we systematically analyzed the junctional complexes of MSC obtained from bone marrow aspirates from healthy voluntary donors and compared the data to bone marrow tissue in situ. Light and electron microscopy was used and several biochemical analyses were applied, including immunoprecipitation and RT-PCR. We could show that intercellular connections in MSC are realized in vitro by occasional gap junctions and frequent adhering junctions. Intercellular projections are noted, often deeply intruding into the neighbouring cell. These findings have been confirmed in situ in bone marrow biopsy material. Furthermore, we analyzed the molecular composition of these junctions and cell-projections. In most of them, we found the transmembrane glycoproteins cadherin 11 and N-cadherin, together with the cytoplasmic plaque proteins alpha- and beta-catenin and protein p120ctn. Interestingly, similar junctions had been already noted in primary mesenchymal stem cells of day 7 to 8.5 mouse embryos. Our hypothesis is that this junction type might be a prerequisite for early mesenchymal cells. Our findings indicate that intercellular communication in MSC in human marrow is realized through a unique type of junctional complexes not described before. These structures are crucial for the long-term fate and differentiation of the cells and might have significant impact on the interaction with hematopoietic stem cells.

scholarly journals Pleiotrophin Regulates the Retention and Self-Renewal of Hematopoietic Stem Cells in the Bone Marrow Vascular Niche

Cell Reports ◽  
2012 ◽  
Vol 2 (4) ◽  
pp. 964-975 ◽  
Author(s):  
Heather A. Himburg ◽  
Jeffrey R. Harris ◽  
Takahiro Ito ◽  
Pamela Daher ◽  
J. Lauren Russell ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Vascular Niche ◽  
Bone Marrow Vascular Niche

182 In Vitro Derivation of Male Germ Cells from Bovine Bone Marrow-Derived Mesenchymal Stem Cells

2018 ◽  
Vol 30 (1) ◽  
pp. 231
Author(s):  
J. Cortez ◽  
J. Bahamonde ◽  
J. Palomino ◽  
M. De los Reyes ◽  
C. Torres ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Germ Cell ◽  
Mrna Expression ◽  
Male Germ Cell ◽  
Bovine Bone ◽  
Mrna Levels ◽  
Germ Cell Differentiation

During the last few years, the in vitro derivation of germ cell lineages from stem cells has emerged as an exciting new strategy for obtaining mature gametes. In vitro gamete derivation technology has potential applications as an alternative method for dissemination of elite animal genetics, production of transgenic animals, and conservation of endangered species. Germ cell differentiation and gametogenesis is a complex process and potential of different stem cell donors (i.e. SSC, ESC, iPSC) for in vitro male germ cell derivation has been inconsistent. Mesenchymal stem cells (MSC) may be suitable candidates for in vitro gamete derivation considering their (1) plasticity that is not limited to mesodermal derivatives, (2) availability of abundant tissues sources for isolation, (3) high proliferative potential, (4) simple and inexpensive isolation, and (5) high potential for cell therapy, including autologous or allogenic transplantation. The present study aimed to induce differentiation of MSC isolated from bone marrow derived from bovine male fetuses (bfMSC) into the germ cell lineage using an in vitro approach based on the exogenous effect of retinoic acid (RA) and bone morphogenetic protein 4 (BMP4). Differentiation media consisted in control media (DMEM with high glucose plus 10% fetal bovine serum, 100 IU mL−1 penicillin, 100 μg mL−1 streptomycin, and 0.25 μg mL−1amphotericin B) supplemented with RA (0.01, 0.1, or 1 µM) or BMP4 (10, 50, or 100 ng mL−1). Cell samples were obtained from differentiating and control bfMSC cultures and analysed for expression of housekeeping genes β-ACTIN and GAPDH, pluripotent genes OCT4 and NANOG, germ cell genes FRAGILLIS, STELLA, and VASA, male germ cell genes DAZL, PIWIl2, and STRA8, and meiotic biomarker SCP3 by quantitative-PCR (Q-PCR). OCT4, NANOG, and DAZL were immunodetected in undifferentiated and differentiated bfMSC using flow-cytometry analysis. The mRNA expression of DAZL was activated by RA or BMP4 supplementation, although no differences (P > 0.05) were detected among different concentrations. DAZL and NANOG mRNA levels increased (P < 0.05) from Day 7 to Day 21 during supplementation of RA (0.1 μM). In comparison, DAZL mRNA levels increased (P < 0.05) at Day 14 during supplementation of BMP4 (100 ng). OCT4 and SCP3 mRNA levels were not affected by RA or BMP4 treatments. Transcripts of FRAGILLIS, STELLA, VASA, PIWIl2, and STRA8 were not detected in control or differentiated bfMSC. Higher (P < 0.05) percentages of undifferentiated bfMSC were positive for NANOG (80.6%) and OCT4 (83.4%). DAZL- and NANOG-positive cells were 2.1% and 2.9%, and 95.9% and 97.8% at Days 0 and 21 of RA treatment, respectively. Data indicated that expression of germ cell biomarker DAZL in bfMSC is activated and increased after in vitro supplementation of RA and BMP4. Moreover, NANOG mRNA levels were regulated by RA treatment. Similar levels of SCP3 mRNA expression suggest that differentiated bfMSC were not induced into meiosis. Thus, exposure of bfMSC to RA or BMP4 under in vitro conditions might induce an early stage of premeiotic germinal differentiation.

scholarly journals Molecular Modulation of Fetal Liver Hematopoietic Stem Cell Mobilization into Fetal Bone Marrow in Mice

2020 ◽  
Vol 2020 ◽  
pp. 1-12
Author(s):  
Huihong Zeng ◽  
Jiaoqi Cheng ◽  
Ying Fan ◽  
Yingying Luan ◽  
Juan Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Vascular Endothelial Cells ◽  
Fetal Liver ◽  
Bone Marrow Niche ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Fetal Bone

Development of hematopoietic stem cells is a complex process, which has been extensively investigated. Hematopoietic stem cells (HSCs) in mouse fetal liver are highly expanded to prepare for mobilization of HSCs into the fetal bone marrow. It is not completely known how the fetal liver niche regulates HSC expansion without loss of self-renewal ability. We reviewed current progress about the effects of fetal liver niche, chemokine, cytokine, and signaling pathways on HSC self-renewal, proliferation, and expansion. We discussed the molecular regulations of fetal HSC expansion in mouse and zebrafish. It is also unknown how HSCs from the fetal liver mobilize, circulate, and reside into the fetal bone marrow niche. We reviewed how extrinsic and intrinsic factors regulate mobilization of fetal liver HSCs into the fetal bone marrow, which provides tools to improve HSC engraftment efficiency during HSC transplantation. Understanding the regulation of fetal liver HSC mobilization into the fetal bone marrow will help us to design proper clinical therapeutic protocol for disease treatment like leukemia during pregnancy. We prospect that fetal cells, including hepatocytes and endothelial and hematopoietic cells, might regulate fetal liver HSC expansion. Components from vascular endothelial cells and bones might also modulate the lodging of fetal liver HSCs into the bone marrow. The current review holds great potential to deeply understand the molecular regulations of HSCs in the fetal liver and bone marrow in mammals, which will be helpful to efficiently expand HSCs in vitro.

scholarly journals Conditioned Medium from Human Tonsil-Derived Mesenchymal Stem Cells Enhances Bone Marrow Engraftment via Endothelial Cell Restoration by Pleiotrophin

Cells ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 221
Author(s):  
Yu-Hee Kim ◽  
Kyung-Ah Cho ◽  
Hyun-Ji Lee ◽  
Minhwa Park ◽  
Sang-Jin Shin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Conditioned Medium ◽  
Histological Analysis ◽  
Conditioning Regimen ◽  
Mouse Bone Marrow ◽  
Human Tonsil ◽  
Hematopoietic Stem ◽  
Bone Marrow Engraftment

Cotransplantation of mesenchymal stem cells (MSCs) with hematopoietic stem cells (HSCs) has been widely reported to promote HSC engraftment and enhance marrow stromal regeneration. The present study aimed to define whether MSC conditioned medium could recapitulate the effects of MSC cotransplantation. Mouse bone marrow (BM) was partially ablated by the administration of a busulfan and cyclophosphamide (Bu–Cy)-conditioning regimen in BALB/c recipient mice. BM cells (BMCs) isolated from C57BL/6 mice were transplanted via tail vein with or without tonsil-derived MSC conditioned medium (T-MSC CM). Histological analysis of femurs showed increased BM cellularity when T-MSC CM or recombinant human pleiotrophin (rhPTN), a cytokine readily secreted from T-MSCs with a function in hematopoiesis, was injected with BMCs. Microstructural impairment in mesenteric and BM arteriole endothelial cells (ECs) were observed after treatment with Bu–Cy-conditioning regimen; however, T-MSC CM or rhPTN treatment restored the defects. These effects by T-MSC CM were disrupted in the presence of an anti-PTN antibody, indicating that PTN is a key mediator of EC restoration and enhanced BM engraftment. In conclusion, T-MSC CM administration enhances BM engraftment, in part by restoring vasculature via PTN production. These findings highlight the potential therapeutic relevance of T-MSC CM for increasing HSC transplantation efficacy.

Characterization of a novel decellularized bone marrow scaffold as an inductive environment for hematopoietic stem cells

2019 ◽  
Vol 7 (4) ◽  
pp. 1516-1528 ◽  
Author(s):  
Juares E. Romero Bianco ◽  
Renata Giardini Rosa ◽  
Ada Congrains-Castillo ◽  
Paulo P. Joazeiro ◽  
Stephen D. Waldman ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Bovine Bone ◽  
Hematopoietic Stem ◽  
Bone Marrow Study ◽  
Increasing Demand

Due to the increasing demand for a bone marrow study model, we developed a natural scaffold from decellularized bovine bone marrow (DeBM).

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