scholarly journals Improved Isolation of Mesenchymal Stem Cells Based on Interactions between N-Acetylglucosamine-Bearing Polymers and Cell-Surface Vimentin

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
Author(s):  
Hirohiko Ise ◽  
Kumiko Matsunaga ◽  
Marie Shinohara ◽  
Yasuyuki Sakai
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Tissue Culture ◽  
Mesenchymal Stem Cells ◽  
Bone Marrow Cells ◽  
Cell Types ◽  
Binding Activity ◽  
Isolation System ◽  
Multiple Cell

Mesenchymal stem cells (MSCs) in bone marrow and adipose tissues are expected to be effective tools for regenerative medicine to treat various diseases. To obtain MSCs that possess both high differentiation and tissue regenerative potential, it is necessary to establish an isolation system that does not require long-term culture. It has previously been reported that the cytoskeletal protein vimentin, expressed on the surfaces of multiple cell types, possesses N-acetylglucosamine- (GlcNAc-) binding activity. Therefore, we tried to exploit this interaction to efficiently isolate MSCs from rat bone marrow cells using GlcNAc-bearing polymer-coated dishes. Cells isolated by this method were identified as MSCs because they were CD34-, CD45-, and CD11b/c-negative and CD90-, CD29-, CD44-, CD54-, CD73-, and CD105-positive. Osteoblast, adipocyte, and chondrocyte differentiation was observed in these cells. In total, yields of rat MSCs were threefold to fourfold higher using GlcNAc-bearing polymer-coated dishes than yields using conventional tissue-culture dishes. Interestingly, MSCs isolated with GlcNAc-bearing polymer-coated dishes strongly expressed CD106, whereas those isolated with conventional tissue-culture dishes had low CD106 expression. Moreover, senescence-associated β-galactosidase activity in MSCs from GlcNAc-bearing polymer-coated dishes was lower than that in MSCs from tissue-culture dishes. These results establish an improved isolation method for high-quality MSCs.

130 Burn Injury Reduces Bone Marrow Mesenchymal Stem Cells and Sensitizes Their Adrenergic Receptor Subgroups in a Murine Model

2021 ◽  
Vol 42 (Supplement_1) ◽  
pp. S87-S88
Author(s):  
Kuzhali Muthumalaiappan ◽  
Maria Camargo Johnson ◽  
Julia Walczak ◽  
Vimal Subramaniam ◽  
Anthony J Baldea ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adrenergic Receptor ◽  
Burn Injury ◽  
Bone Marrow Cells ◽  
Ex Vivo ◽  
Traumatic Injury ◽  
Hematopoietic Cell ◽  
The Right

Abstract Introduction Previous burn and traumatic injury studies have established that adrenergic signaling is increased after burn injury and may lead to an impairment of hematopoietic cell development in the bone marrow (BM). Nonetheless, mesenchymal stem cells (MSCs), which have gained momentum in regenerative medicine also play a predominant role in the BM niche. Understanding the propensity of the adrenergic receptor (AR) response by MSCs can be utilized for devising targeted therapies. However, the traditional plastic adherence procedure using ex vivo culture of BM cells for several weeks may skew the actual characteristics of MSCs. Our current study focused on isolating MSCs from freshly obtained BM in a murine scald burn model with a goal to characterize the expression pattern of native AR subgroups present on BM MSCs as compared to sham mice. Methods Eight, two-month-old adult female mice were subjected to a 15% total body 3rd degree burn or sham burn. The mice were sacrificed 7 days later. Femurs were removed and total bone marrow cells were flushed out. Multi parametric flow cytometry was used to gate for cells negative for hematopoietic cell markers (CD45, CD11B) and positive for MSC markers (CD105, CD106, SSEA, Ly6A) and AR subgroups (α1, α2, β1, β2, β3). We measured the number of BM MSCs, quantified the subtypes of ARs present on MSCs, and compared the ratio of AR antibody binding per total MSC population. Results Overall the frequency of MSCs per million total BM cells decreased by 48% post-burn injury with165,300 ± 194 in sham versus 110,000 ± 30 in burn displayed as bar graph in Panel A. Over 90% of MSCs consistently express β2 AR and only 10% express α2 AR subgroup in both scald and sham burn. Presence of other subgroups ranged from 50% to 80% of MSCs as seen in histograms to the right of dotted line in Panel B. Our AR propensity score based on AR mean fluorescence intensity adjusted to total number of MSCs present was increased by 2.8-fold for α1, 2.5-fold for β1, 1.6-fold for β3, and 1.3-fold for β2 AR subgroups (Panel C). These findings indicate burn injury not only decreases the frequency of BM MSCs but also increases the affinity of certain AR subgroups present on MSCs. Since BM MSCs are the major source of cytokines, chemokines and growth factors; detailed studies on AR mediated signaling in BM MSCs is warranted. Conclusions Polarization of AR signaling in BM MSCs by burn-induced catecholamines may have broader implications for comorbidities such as bone resorption and muscle wasting observed in human patients post burn trauma.

Evaluation of Canine Bone Marrow-derived Mesenchymal Stem Cells After Long-term Cryopreservation

2013 ◽  
Vol 30 (12) ◽  
pp. 1032-1037 ◽  
Author(s):  
Xiaofeng Zhu ◽  
Fang Yuan ◽  
Houxuan Li ◽  
Yuqian Zheng ◽  
Yin Xiao ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells

scholarly journals Comparison of the Proliferation and Differentiation Potential of Human Urine-, Placenta Decidua Basalis-, and Bone Marrow-Derived Stem Cells

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Chengguang Wu ◽  
Long Chen ◽  
Yi-zhou Huang ◽  
Yongcan Huang ◽  
Ornella Parolini ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Stem Cell Differentiation ◽  
Cell Types ◽  
Stem Cell Marker ◽  
Differentiation Potential ◽  
Multipotent Stem Cells ◽  
Decidua Basalis

Human multipotent stem cell-based therapies have shown remarkable potential in regenerative medicine and tissue engineering applications due to their abilities of self-renewal and differentiation into multiple adult cell types under appropriate conditions. Presently, human multipotent stem cells can be isolated from different sources, but variation among their basic biology can result in suboptimal selection of seed cells in preclinical and clinical research. Thus, the goal of this study was to compare the biological characteristics of multipotent stem cells isolated from human bone marrow, placental decidua basalis, and urine, respectively. First, we found that urine-derived stem cells (USCs) displayed different morphologies compared with other stem cell types. USCs and placenta decidua basalis-derived mesenchymal stem cells (PDB-MSCs) had superior proliferation ability in contrast to bone marrow-derived mesenchymal stem cells (BMSCs); these cells grew to have the highest colony-forming unit (CFU) counts. In phenotypic analysis using flow cytometry, similarity among all stem cell marker expression was found, excluding CD29 and CD105. Regarding stem cell differentiation capability, USCs were observed to have better adipogenic and endothelial abilities as well as vascularization potential compared to BMSCs and PDB-MSCs. As for osteogenic and chondrogenic induction, BMSCs were superior to all three stem cell types. Future therapeutic indications and clinical applications of BMSCs, PDB-MSCs, and USCs should be based on their characteristics, such as growth kinetics and differentiation capabilities.

scholarly journals Label-Free Rapid Separation and Enrichment of Bone Marrow-Derived Mesenchymal Stem Cells from a Heterogeneous Cell Mixture Using a Dielectrophoresis Device

Sensors ◽  
2018 ◽  
Vol 18 (9) ◽  
pp. 3007 ◽  
Author(s):  
Junya Yoshioka ◽  
Yu Ohsugi ◽  
Toru Yoshitomi ◽  
Tomoyuki Yasukawa ◽  
Naoki Sasaki ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Gradient Method ◽  
Label Free ◽  
Rapid Separation ◽  
Isolation System ◽  
Tissue Samples ◽  
Cell Based Therapy

Bone marrow-derived mesenchymal stem cells (BMSCs) are an important cell resource for stem cell-based therapy, which are generally isolated and enriched by the density-gradient method based on cell size and density after collection of tissue samples. Since this method has limitations with regards to purity and repeatability, development of alternative label-free methods for BMSC separation is desired. In the present study, rapid label-free separation and enrichment of BMSCs from a heterogeneous cell mixture with bone marrow-derived promyelocytes was successfully achieved using a dielectrophoresis (DEP) device comprising saw-shaped electrodes. Upon application of an electric field, HL-60 cells as models of promyelocytes aggregated and floated between the saw-shaped electrodes, while UE7T-13 cells as models of BMSCs were effectively captured on the tips of the saw-shaped electrodes. After washing out the HL-60 cells from the device selectively, the purity of the UE7T-13 cells was increased from 33% to 83.5% within 5 min. Although further experiments and optimization are required, these results show the potential of the DEP device as a label-free rapid cell isolation system yielding high purity for rare and precious cells such as BMSCs.

scholarly journals Differentiation of Adipose-Derived Mesenchymal Stem Cells into Neuron-Like Cells induced by using β-mercaptoethanol

2020 ◽  
Vol 17 (1(Suppl.)) ◽  
pp. 0235
Author(s):  
Maeda Mohammad ◽  
Ahmed Majeed Al-Shammari ◽  
Rafal H Abdulla ◽  
Aesar Ahmed ◽  
Aseel Khalid
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Light Chain ◽  
Bone Marrow Cells ◽  
Neurofilament Light Chain ◽  
Neurofilament Light ◽  
Study Objective ◽  
Marrow Cells

Background: Adipose derived-mesenchymal stem cells have been used as an alternative to bone marrow cells in this study. Objective: We investigated the in vitro isolation, identification, and differentiation of stem cells into neuron cells, in order to produce neuron cells via cell culture, which would be useful in nerve injury treatment. Method: Mouse adipose mesenchymal stem cells were dissected from the abdominal subcutaneous region. Neural differentiation was induced using β-mercaptoethanol. This study included two different neural stage markers, i.e. nestin and neurofilament light-chain, to detect immature and mature neurons, respectively. Results: The immunocytochemistry results showed that the use of β-mercaptoethanol resulted in the successful production of neuron cells. This was attributable to the increase and significant overexpression of the nestin protein during the early exposure period, which resulted in the expression of the highest levels of nestin. In comparison, the expression level of neurofilament light-chain protein also increased with time but less than nestin. Non-treated mesenchymal stem cells, considered as control showed very low expression for both markers. Conclusion: The results of this study indicate that adipose mesenchymal cells represent a good, easily obtainable source of bone marrow cells used to developing the differentiation process.

Effects Of Hypoxia in Long-Term In Vitro Expansion of Human Bone Marrow Derived Mesenchymal Stem Cells

2017 ◽  
Vol 118 (10) ◽  
pp. 3072-3079 ◽  
Author(s):  
Annelise Pezzi ◽  
Bruna Amorin ◽  
Álvaro Laureano ◽  
Vanessa Valim ◽  
Alice Dahmer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Human Bone ◽  
Human Bone Marrow ◽  
In Vitro Expansion

scholarly journals Comparison of the Biological Characteristics of Mesenchymal Stem Cells Derived from Bone Marrow and Skin

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Ruifeng Liu ◽  
Wenjuan Chang ◽  
Hong Wei ◽  
Kaiming Zhang
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cell Types ◽  
Culture Conditions ◽  
Biological Characteristics ◽  
Cytokine Secretion ◽  
Surface Markers ◽  
Tissue Repair And Regeneration

Mesenchymal stem cells (MSCs) exhibit high proliferation and self-renewal capabilities and are critical for tissue repair and regeneration during ontogenesis. They also play a role in immunomodulation. MSCs can be isolated from a variety of tissues and have many potential applications in the clinical setting. However, MSCs of different origins may possess different biological characteristics. In this study, we performed a comprehensive comparison of MSCs isolated from bone marrow and skin (BMMSCs and SMSCs, resp.), including analysis of the skin sampling area, separation method, culture conditions, primary and passage culture times, cell surface markers, multipotency, cytokine secretion, gene expression, and fibroblast-like features. The results showed that the MSCs from both sources had similar cell morphologies, surface markers, and differentiation capacities. However, the two cell types exhibited major differences in growth characteristics; the primary culture time of BMMSCs was significantly shorter than that of SMSCs, whereas the growth rate of BMMSCs was lower than that of SMSCs after passaging. Moreover, differences in gene expression and cytokine secretion profiles were observed. For example, secretion of proliferative cytokines was significantly higher for SMSCs than for BMMSCs. Our findings provide insights into the different biological functions of both cell types.

scholarly journals Myelosuppressive conditioning is required to achieve engraftment of pluripotent stem cells contained in moderate doses of syngeneic bone marrow

Blood ◽  
1994 ◽  
Vol 83 (4) ◽  
pp. 939-948 ◽  
Author(s):  
Y Tomita ◽  
DH Sachs ◽  
M Sykes
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Whole Body ◽  
Mixed Chimerism ◽  
Hematopoietic Stem ◽  
Donor Cells ◽  
Donor Type ◽  
Low Levels

Abstract We have investigated the requirement for whole body irradiation (WBI) to achieve engraftment of syngeneic pluripotent hematopoietic stem cells (HSCs). Recipient B6 (H-2b; Ly-5.2) mice received various doses of WBI (0 to 3.0 Gy) and were reconstituted with 1.5 x 10(7) T-cell-depleted (TCD) bone marrow cells (BMCs) from congenic Ly-5.1 donors. Using anti-Ly-5.1 and anti-Ly-5.2 monoclonal antibodies and flow cytometry, the origins of lymphoid and myeloid cells reconstituting the animals were observed over time. Chimerism was at least initially detectable in all groups. However, between 1.5 and 3 Gy WBI was the minimum irradiation dose required to permit induction of long-term (at least 30 weeks), multilineage mixed chimerism in 100% of recipient mice. In these mice, stable reconstitution with approximately 70% to 90% donor-type lymphocytes, granulocytes, and monocytes was observed, suggesting that pluripotent HSC engraftment was achieved. About 50% of animals conditioned with 1.5 Gy WBI showed evidence for donor pluripotent HSC engraftment. Although low levels of chimerism were detected in untreated and 0.5-Gy-irradiated recipients in the early post-BM transplantation (BMT) period, donor cells disappeared completely by 12 to 20 weeks post-BMT. BM colony assays and adoptive transfers into secondary lethally irradiated recipients confirmed the absence of donor progenitors and HSCs, respectively, in the marrow of animals originally conditioned with only 0.5 Gy WBI. These results suggest that syngeneic pluripotent HSCs cannot readily engraft unless host HSCs sustain a significant level of injury, as is induced by 1.5 to 3.0 Gy WBI. We also attempted to determine the duration of the permissive period for syngeneic marrow engraftment in animals conditioned with 3 Gy WBI. Stable multilineage chimerism was uniformly established in 3-Gy-irradiated Ly-5.2 mice only when Ly-5.1 BMC were injected within 7 days of irradiation, suggesting that repair of damaged host stem cells or loss of factors stimulating engraftment may prevent syngeneic marrow engraftment after day 7.

scholarly journals Efficient retroviral gene transfer to purified long-term repopulating hematopoietic stem cells

Blood ◽  
1994 ◽  
Vol 84 (1) ◽  
pp. 74-83 ◽  
Author(s):  
SJ Szilvassy ◽  
S Cory
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Cell Biology ◽  
Bone Marrow Cells ◽  
High Titer ◽  
Marker Gene ◽  
Mouse Bone Marrow ◽  
Hematopoietic Stem

Abstract Efficient gene delivery to multipotential hematopoietic stem cells would greatly facilitate the development of effective gene therapy for certain hematopoietic disorders. We have recently described a rapid multiparameter sorting procedure for significantly enriching stem cells with competitive long-term lymphomyeloid repopulating ability (CRU) from 5-fluorouracil (5-FU)-treated mouse bone marrow. The sorted cells have now been tested as targets for retrovirus-mediated delivery of a marker gene, NeoR. They were cocultured for 4 days with fibroblasts producing a high titer of retrovirus in medium containing combinations of the hematopoietic growth factors interleukin-3 (IL-3), IL-6, c-kit ligand (KL), and leukemia inhibitory factor (LIF) and then injected into lethally irradiated recipients, together with sufficient “compromised” bone marrow cells to provide short-term support. Over 80% of the transplanted mice displayed high levels (> or = 20%) of donor- derived leukocytes when analyzed 4 to 6 months later. Proviral DNA was detected in 87% of these animals and, in half of them, the majority of the hematopoietic cells were marked. Thus, infection of the stem cells was most effective. The tissue and cellular distribution of greater than 100 unique clones in 55 mice showed that most sorted stem cells had lymphoid as well as myeloid repopulating potential. Secondary transplantation provided strong evidence for infection of very primitive stem cells because, in several instances, different secondary recipients displayed in their marrow, spleen, thymus and day 14 spleen colony-forming cells the same proviral integration pattern as the primary recipient. Neither primary engraftment nor marking efficiency varied for stem cells cultured in IL-3 + IL-6, IL-3 + IL-6 + KL, IL-3 + IL-6 + LIF, or all four factors, but those cultured in IL-3 + IL-6 + LIF appeared to have lower secondary engraftment potential. Provirus expression was detected in 72% of the strongly marked mice, albeit often at low levels. Highly efficient retroviral marking of purified lymphomyeloid repopulating stem cells should enhance studies of stem cell biology and facilitate analysis of genes controlling hematopoietic differentiation and transformation.

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