scholarly journals Comparative characterization and osteogenic / adipogenic differentiation of mesenchymal stem cells derived from male rat hair follicles and bone marrow

2020 ◽  
Vol 9 (1) ◽  
Author(s):  
Abdel Kader A. Zaki ◽  
Tariq I. Almundarij ◽  
Faten A. M. Abo-Aziza
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Adipogenic Differentiation ◽  
Adult Stem Cell ◽  
Cell Counting ◽  
Population Doubling ◽  
Male Rat ◽  
Cell Source

AbstractClinical applications of cell therapy and tissue regeneration under different conditions need a multiplicity of adult stem cell sources. Up to date, little is available on the comparative isolation, characterization, proliferation, rapid amplification, and osteogenic/adipogenic differentiation of rat mesenchymal stem cells (MSCs) isolated from living bulge cells of the hair follicle (HF) and bone marrow (BM) from the same animal. This work hopes to use HF-MSCs as an additional adult stem cell source for research and application. After reaching 80% confluence, the cell counting, viability %, and yields of HF-MSCs and BM-MSCs were nearly similar. The viability % was 91.41 ± 2.98 and 93.11 ± 3.06 while the cells yield of initial seeding was 33.15 ± 2.76 and 34.22 ± 3.99 and of second passage was 28.76 ± 1.01 and 29.56 ± 3.11 for HF-MSCs and BM-MSCs respectively. Clusters of differentiation (CDs) analysis revealed that HF-MSCs were positively expressed CD34, CD73 and CD200 and negatively expressed CD45. BM-MSCs were positively expressed CD73 and CD200 and negatively expressed of CD34 and CD45. The proliferation of HF-MSCs and BM-MSCs was determined by means of incorporation of Brd-U, population doubling time (PDT) assays and the quantity of formazan release. The percentage of Brd-U positive cells and PDT were relatively similar in both types of cells. The proliferation, as expressed by the quantity of formazan assay in confluent cells, revealed that the quantity of release by BM-MSCs was slightly higher than HF-MSCs. Adipogenic differentiated BM-MSCs showed moderate accumulation of oil red-O stained lipid droplets when compared to that of HF-MSCs which exhibited high stain. The total lipid concentration was significantly higher in adipogenic differentiated HF-MSCs than BM-MSCs (P < 0.05). It was found that activity of bone alkaline phosphatase and calcium concentration were significantly higher (P < 0.01 and P < 0.05 respectively) in osteogenic differentiated BM-MSCs than that of HF-MSCs. The present findings demonstrate that the HF-MSCs are very similar in most tested characteristics to BM-MSCs with the exception of differentiation. Additionally; no issues have been reported during the collection of HF-MSCs. Therefore, the HF may represent a suitable and accessible source for adult stem cells and can be considered an ideal cell source for adipogenesis research.

390 ISOLATION AND ADIPOGENESIS OF GRIZZLY BEAR MESENCHYMAL STEM CELLS

2010 ◽  
Vol 22 (1) ◽  
pp. 351
Author(s):  
A. J. Maki ◽  
I. Omelogu ◽  
E. Monaco ◽  
M. E. McGee-Lawrence ◽  
R. M. Bradford ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Stem Cell ◽  
In Vitro Model ◽  
Adipogenic Differentiation ◽  
Grizzly Bear ◽  
Vitro Model

During winter hibernation, grizzly bears (Ursus arctos horribilis) do not eat but instead rely on internal fat stores as a primary source of metabolic energy. The resulting seasonal fluctuations in appetite and body mass make the grizzly bear a naturally occurring animal model for human conditions such as obesity and anorexia. An in vitro model of hibernating bear stem cells might enhance our understanding of processes such as stem cell proliferation and differentiation. Mesenchymal stem cells, derived from bone marrow and adipose tissue among others, differentiate into adipocytes and might play important roles in energy metabolism. In the current study, we examined the in vitro viability and morphology of mesenchymal stem cells isolated from grizzly bear adipose tissue (ADSC) and bone marrow (BMSC); these ADSC and BMSCs underwent adipogenic differentiation for 0, 7, 14, 21, and 28 days. Bone marrow stem cells and ADSC were isolated using mechanical disaggregation, collagenase digestion, centrifugation, and plating onto tissue culture polystyrene. Cell viability and proliferation was quantified using the colony forming unit assay and a hemocytometer. Both stem cell types were differentiated into adipocytes using 10 μM insulin, 1 μM dexamethasone, and 0.5 mM isobutylmethylxanthine (all Sigma- Aldrich, St. Louis, MO, USA) with the addition of 10% fetal bovine (FBS) or bear serum from the active feeding period. Adipogenic differentiation was confirmed using Oil Red O and quantified using ImageJ. Statistical analysis was performed using an unpaired t-test between treatments of the same time point. All cells were isolated within 28 h of tissue harvest. Adipose-derived stem cells formed an average of 11 colonies (0.011%), whereas BMSC formed 1.5 colonies (0.0015%) per 100 000 cells. Doubling time forADSC was approximately 54 h in 10% FBS. BothADSC and BMSC had an initial spindle-shaped morphology, which gradually became more rounded during adipogenic differentiation. For bear serum at Day 28, ADSC had a significantly (P < 0.01) greater stained area per cell than did BMSC. In summary, both types of mesenchymal stem cells successfully differentiated into adipocytes and maintained viability. In conclusion, grizzly bear mesenchymal stem cells canbesuccessfully isolated, expanded, and differentiated in culture. These results allow for future studies using the bear as an in vitro model for fat metabolism during hibernation and active periods. This work was partially supported by the Carle Foundation Hospital, the Intel Scholar’s Research Program, USDA Multi-State Research Project W1171, and the Illinois Regenerative Medicine Institute (IDPH # 63080017). In addition, the authors would like to thank Agatha Luszpak for support with the analysis.

Following Autologous Stem Cell Transplantation (1 Yr) Mesenchymal Stem Cells Are Functionally Impaired with Reduced Hematopoietic Support

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3888-3888
Author(s):  
Catharina Hazenberg ◽  
Fiona A.J. van den Heuvel ◽  
Edo Vellenga ◽  
Annet Z. Brouwers-Vos ◽  
Gerbrig Berger ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Autologous Stem Cell Transplantation ◽  
Population Doubling ◽  
Bone Marrow Biopsies

Abstract Autologous stem cell transplantation (ASCT) is frequently applied in patients with multiple myeloma and malignant lymphoma. Although adequate steady state hematopoiesis with normal peripheral blood counts is attained after ASCT, marked cytopenias may occur in times of stress such as sepsis or re-exposure to chemotherapy. Our group has previously shown impairment of the hematopoietic stem cell (HSC) compartment 1 year post ASCT (pASCT), reflected by reduced HSC frequency and quiescence, and increased ROS production (Haematologica 2013;98:1264). Considering the essential role for mesenchymal stem cells (MSCs) in supporting hematopoiesis, we studied the MSC compartment 1 year post ASCT. Bone marrow biopsies from pASCT patients (n=17) were studied and compared to normal bone marrow from healthy donors (NBM, n=20) by performing immunohistochemistry staining of endothelial cells by CD34 (indicating microvessel density, MVD) and MSCs by nestin, CD146 (Melanoma Cell Adhesion Molecule, MCAM) and CD271 (Nerve Growth Factor Receptor, NGFR). A significant increase in CD271+ MSCs was observed in pASCT bone marrow biopsies compared to NBM (p<0.0001), while the expression of additional markers did not differ between pASCT vs. NBM. MSCs were cultured from the CD34- fraction of bone marrow mononuclear cells, obtained from pASCT patients (n=17) and MSCs derived from NBM (n=20). MSCs were selected by their plastic-adherency and replated to generate MSCs. Although pASCT MSCs and NBM MSCs had similar population doubling times (1.92±0.22 and 3.52±1.02 in passage 4 (P4), pASCT MSCs cultured in vitro demonstrated a change in morphology from the onset of P4. We also observed premature exhaustion of growth in 45% of the studied patients at P5 (vs. 18% in NBM) and increased senescence shown by B-galactosidase staining in P5/P6 (p=0.04). Differentiation assays did not show impairment in differentiation towards osteoblasts or adipocytes of pASCT MSCs. Gene expression analysis on early passage MSCs showed upregulation of pro-inflammatory and cell cycle genes, such as IL6 and p21, in pASCT MSCs compared to NBM MSCs. Co-culture studies with cord blood-derived CD34+ cells on pASCT MSCs showed a significant reduction in output in CFC assays and significant reduction in number of cobblestone-area forming cells in pASCT co-cultures versus NBM (p < 0.05). Given the higher incidence of MDS and AML after ASCT, we questioned whether the observed phenotype of pASCT MSCs resembles MSCs from patients with MDS and AML. Therefore the endothelial and mesenchymal compartments of MDS (n=20) and AML (n=23) patients were studied. An increase in MVD was detected in MDS/AML bone marrow biopsies in contrast to NBM and pASCT (p < 0.05), while the expression of CD146, CD271 and nestin in MDS/AML patients was not significantly increased. 25% of AML MSC cultures showed no growth in the first passage. When MSC growth did occur, the remaining cultures did not show a difference in population doubling time or expansion. However, a change in morphology of MDS/AML MSCs similar to pASCT MSCs was observed. Studies of early passages of MDS/AML MSCs demonstrated a significantly increased gene expression of IL-6 and p21 comparable to pASCT MSCs. In addition PITX2 and Foxc1 expression was increased but no difference was observed in pASCT vs. MDS/AML MSCs. PITX2 has been linked to increased senescence of MDS MSCs while Foxc1 is linked to adipo-osteoprogenitor cell differentiation thereby affecting the HSC compartment. Since none of the pASCT patients did develop MDS, immunohistochemical stainings were also performed on bone marrow biopsies of patients that developed therapy related (t-)MDS/AML following ASCT for lymphoma and myeloma (n=7), after a mean of 117 (MDS) and 50 months (AML). An increase in MVD was observed shortly before or during MDS/AML development, which is probably related to the emergence of malignant cells. No major changes in the phenotype of the MSC compartment were observed before or during the emergence of t-MDS/AML, indicating that t-MDS/AML is preceded by an increase in MVD without distinct changes in the MSC compartment. In summary our results demonstrate that MSCs are affected after ASCT, as shown by expression pattern and functionality. These changes result in a pro-inflammatory phenotype with premature senescence and impaired support of hematopoietic cells, which may account for the reduced bone marrow reserve observed in pASCT patients. Disclosures No relevant conflicts of interest to declare.

scholarly journals Cytotoxic Effect of Hypoxic Environment in Mesenchymal Stem Cell

Proceedings ◽  
2018 ◽  
Vol 2 (25) ◽  
pp. 1592
Author(s):  
Sevil Özer ◽  
H. Seda Vatansever ◽  
Feyzan Özdal-Kurt
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Mesenchymal Stem Cell ◽  
Human Mesenchymal Stem Cells ◽  
Hypoxic Condition ◽  
Immunoperoxidase Technique ◽  
Cellular Functions ◽  
Marrow Mesenchymal Stem Cells

Bone marrow mesenchymal stem cells (BM-MSCs) are used to repair hypoxic or ischemic tissue. After hypoxic the level of ATP is decreases, cellular functions do not continue and apoptosis or necrosis occur. Apoptosis is a progress of programmed cell death that occurs in normal or pathological conditions. In this study, we were investigated the hypoxic effect on apoptosis in mesenchymal stem cell. Bone marrow-derived stem cells were cultured in hypoxic (1% or 3%) or normoxic conditions 24, 96 well plates for 36 h. Cell viability was shown by MTT assay on 36 h. After fixation of cells with 4% paraformaldehyde, distributions of caspase-3, Bcl-2 and Bax with indirect immunoperoxidase technique, apoptotic cells with TUNEL assay were investigated. All staining results were evaluated using H-score analyses method with ANOVA, statistically. As a result, hypoxic condition was toxic for human mesenchymal stem cells and the number of death cell was higher in that than normoxic condition.

scholarly journals Population doubling level-dependent change of secreted glycosaminoglycan in equine bone marrow-derived mesenchymal stem cells

2015 ◽  
Vol 26 (3) ◽  
pp. 73-80 ◽  
Author(s):  
Takafumi SASAO ◽  
Yuki FUKUDA ◽  
Sayako YOSHIDA ◽  
Shihori MIYABARA ◽  
Yoshinori KASASHIMA ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Population Doubling ◽  
Population Doubling Level ◽  
Dependent Change ◽  
Equine Bone

Effect of Sirtuin1 (Sirt1) on Bone Marrow Mesenchymal Stem Cells (BMSCs) Osteogenesis/Adipogenesis via β-Catenin/The Transcription Factor T Cell Factor 1 (TCF1)/Runt-Related Transcription Factor 2 (RUNX2)

2021 ◽  
Vol 11 (10) ◽  
pp. 2070-2075
Author(s):  
Wenji Shi ◽  
Mingxing Zhao ◽  
Guangxia Shi
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipogenic Differentiation ◽  
Sham Operation ◽  
Runx2 Expression ◽  
Sham Operation Group ◽  
Marrow Mesenchymal Stem Cells ◽  
Operation Group ◽  
Sirna Group

Bone marrow mesenchymal stem cells (BMSCs) have self-renewal potential. Sirt1 regulates cell differentiation and apoptosis. However, Sirt1’s effect on BMSCs osteogenic/adipogenic differentiation has not been fully elucidated. SD rats were randomly divided into Osteoporosis (OP) group and sham operation group. OP rat BMSCs were isolated and assigned into control group, NC group and Sirt1 siRNA group followed by analysis of Sirt1 level by Real-time PCR, cell proliferation by MTT assay, expression of OC, OPN and FABP4 level by real time PCR, and β-Catenin/TCF1/Runx2 protein expression by Western blot. In OP group, Sirt1 expression was significantly increased and BMSCs proliferation was decreased along with reduced OC and OPN mRNA expression, increased FABP4 expression and reduced β-Catenin/TCF1/Runx2 expression compared with sham operation group (P < 0.05). In Sirt1 siRNA group, Sirt1 expression was significantly reduced, BMSCs proliferation was increased, OC and OPN mRNA expression was increased, FABP4 expression was decreased, and β-Catenin/TCF1/Runx2 expression was increased compared to OP group (P < 0.05). Sirt1 is increased in osteoporosis. Down-regulating Sirt1 in osteoporotic BMSCs can regulate β-Catenin/TCF1/Runx2 signaling and promote BMSCs osteogenic differentiation and inhibit adipogenic differentiation.

Neuroprotective effect of bone marrow derived mesenchymal stem cell secretome in 6-OHDA-induced Parkinson's disease

2021 ◽  
Author(s):  
Dhruv Mahendru ◽  
Ashish Jain ◽  
Seema Bansal ◽  
Deepti Malik ◽  
Neha Dhir ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Stem Cells ◽  
Bone Marrow ◽  
Parkinson's Disease ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Neuroprotective Effect ◽  
Behavioral Parameters ◽  
Old Rats ◽  
6 Hydroxydopamine

Aim: The aim of the study was to evaluate the neuroprotective effect of bone marrow stem cell secretome in the 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. Materials & methods: Secretome prepared from mesenchymal stem cells of 3-month-old rats was injected daily for 7 days between days 7 and 14 after 6-OHDA administration. After 14 days, various neurobehavioral parameters were conducted. These behavioral parameters were further correlated with biochemical and molecular findings. Results & conclusion: Impaired neurobehavioral parameters and increased inflammatory, oxidative stress and apoptotic markers in the 6-OHDA group were significantly modulated by secretome-treated rats. In conclusion, mesenchymal stem cells-derived secretome could be further explored for the management of Parkinson's disease.

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.

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