scholarly journals Potential Marker Genes for Predicting Adipogenic Differentiation of Mesenchymal Stromal Cells

2019 ◽  
Vol 9 (14) ◽  
pp. 2942 ◽  
Author(s):  
Masami Kanawa ◽  
Akira Igarashi ◽  
Katsumi Fujimoto ◽  
Veronica Sainik Ronald ◽  
Yukihito Higashi ◽  
...  
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Expression Profiles ◽  
Adipogenic Differentiation ◽  
Expression Patterns ◽  
Marker Genes ◽  
Mrna Levels ◽  
Differentiation Potential ◽  
Potential Marker ◽  
Promising Source

Mesenchymal stromal cells (MSCs) are a promising source for tissue engineering of soft connective tissues. However, the differentiation capacity of MSCs varies among individual cell lines. Here, we show marker genes to predict the adipogenic potential of MSCs. To clarify the correlation between gene expression patterns before adipogenic induction and the differentiation level of MSCs after differentiation, we compared mRNA levels of 95 genes and glycerol-3-phosphate dehydrogenase (GPDH) activities in 15 MSC lines (five jaw and 10 ilium MSCs) from 15 donors. Expression profiles of 22 genes before differentiation significantly correlated with GPDH activities after differentiation. Expression levels of 11 out of the 22 genes in highly potent ilium MSCs were at least three times higher compared with jaw MSCs, which have limited differentiation potential. Furthermore, three-dimensional scatter plot for mRNA expression of ITGA5, CDKN2D, and CD74 could completely distinguish highly potent MSCs from poorly potent MSCs for adipogenesis. The treatment of MSC cultures with the anti-ITGA5 antibody reduced adipogenic differentiation of MSCs. Collectively, these results suggest that the three genes play a role in adipogenesis before induction and can serve as predictors to select potent MSCs for adipogenic differentiation.

scholarly journals The Identification of Marker Genes for Predicting the Osteogenic Differentiation Potential of Mesenchymal Stromal Cells

2021 ◽  
Vol 43 (3) ◽  
pp. 2157-2166
Author(s):  
Masami Kanawa ◽  
Akira Igarashi ◽  
Katsumi Fujimoto ◽  
Tania Saskianti ◽  
Ayumu Nakashima ◽  
...  
Keyword(s):  
Regenerative Medicine ◽  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Marker Genes ◽  
Mrna Levels ◽  
Differentiation Potential ◽  
Alp Activity ◽  
Osteogenic Induction ◽  
Promising Source

Mesenchymal stromal cells (MSCs) have the potential to differentiate into a variety of mature cell types and are a promising source of regenerative medicine. The success of regenerative medicine using MSCs strongly depends on their differentiation potential. In this study, we sought to identify marker genes for predicting the osteogenic differentiation potential by comparing ilium MSC and fibroblast samples. We measured the mRNA levels of 95 candidate genes in nine ilium MSC and four fibroblast samples before osteogenic induction, and compared them with alkaline phosphatase (ALP) activity as a marker of osteogenic differentiation after induction. We identified 17 genes whose mRNA expression levels positively correlated with ALP activity. The chondrogenic and adipogenic differentiation potentials of jaw MSCs are much lower than those of ilium MSCs, although the osteogenic differentiation potential of jaw MSCs is comparable with that of ilium MSCs. To select markers suitable for predicting the osteogenic differentiation potential, we compared the mRNA levels of the 17 genes in ilium MSCs with those in jaw MSCs. The levels of 7 out of the 17 genes were not substantially different between the jaw and ilium MSCs, while the remaining 10 genes were expressed at significantly lower levels in jaw MSCs than in ilium MSCs. The mRNA levels of the seven similarly expressed genes were also compared with those in fibroblasts, which have little or no osteogenic differentiation potential. Among the seven genes, the mRNA levels of IGF1 and SRGN in all MSCs examined were higher than those in any of the fibroblasts. These results suggest that measuring the mRNA levels of IGF1 and SRGN before osteogenic induction will provide useful information for selecting competent MSCs for regenerative medicine, although the effectiveness of the markers is needed to be confirmed using a large number of MSCs, which have various levels of osteogenic differentiation potential.

scholarly journals Genetic Markers Can Predict Chondrogenic Differentiation Potential in Bone Marrow-Derived Mesenchymal Stromal Cells

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Masami Kanawa ◽  
Akira Igarashi ◽  
Katsumi Fujimoto ◽  
Yukihito Higashi ◽  
Hidemi Kurihara ◽  
...  
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Chondrogenic Differentiation ◽  
Expression Profiles ◽  
Three Dimensional ◽  
Marker Genes ◽  
Differentiation Potential ◽  
Expression Levels ◽  
Group 2 ◽  
Msc Differentiation

The precise predictions of the differentiation direction and potential of mesenchymal stromal cells (MSCs) are an important key to the success of regenerative medicine. The expression levels of fate-determining genes may provide tools for predicting differentiation potential. The expression levels of 95 candidate marker genes and glycosaminoglycan (GAG) contents after chondrogenic induction in 10 undifferentiated ilium and 5 jaw MSC cultures were determined, and their correlations were analyzed. The expression levels of eight genes before the induction of chondrogenic MSC differentiation were significantly correlated with the GAG levels after induction. Based on correlation patterns, the eight genes were classified into two groups: group 1 genes (AURKB, E2F1, CDKN2D, LIF, and ACLY), related to cell cycle regulation, and group 2 genes (CD74, EFEMP1, and TGM2), involved in chondrogenesis. The expression levels of the group 2 genes were significantly correlated with the ages of the cell donors. The expression levels of CDKN2D, CD74, and TGM2 were >10-fold higher in highly potent MSCs (ilium MSCs) than in MSCs with limited potential (jaw MSCs). Three-dimensional (3D) scatter plot analyses of the expression levels of these genes showed reduced variability between donors and confirmed predictive potential. These data suggest that group 2 genes are involved in age-dependent decreases in the chondrogenic differentiation potential of MSCs, and combined 3D analyses of the expression profiles of three genes, including two group 2 genes, were predictive of MSC differentiation potential.

scholarly journals Uniaxial Cyclic Tensile Stretching at 8% Strain Exclusively Promotes Tenogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stromal Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-16 ◽  
Author(s):  
Hui Yin Nam ◽  
Belinda Pingguan-Murphy ◽  
Azlina Amir Abbas ◽  
Azhar Mahmood Merican ◽  
Tunku Kamarul
Keyword(s):  
Gene Expression ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Mechanical Strain ◽  
Marker Genes ◽  
Actin Stress Fiber ◽  
Differentiation Potential ◽  
Cyclic Stretching ◽  
Tenogenic Differentiation ◽  
Collagen Iii

The present study was conducted to establish the amount of mechanical strain (uniaxial cyclic stretching) required to provide optimal tenogenic differentiation expression in human mesenchymal stromal cells (hMSCs) in vitro, in view of its potential application for tendon maintenance and regeneration. Methods. In the present study, hMSCs were subjected to 1 Hz uniaxial cyclic stretching for 6, 24, 48, and 72 hours; and were compared to unstretched cells. Changes in cell morphology were observed under light and atomic force microscopy. The tenogenic, osteogenic, adipogenic, and chondrogenic differentiation potential of hMSCs were evaluated using biochemical assays, extracellular matrix expressions, and selected mesenchyme gene expression markers; and were compared to primary tenocytes. Results. Cells subjected to loading displayed cytoskeletal coarsening, longer actin stress fiber, and higher cell stiffness as early as 6 hours. At 8% and 12% strains, an increase in collagen I, collagen III, fibronectin, and N-cadherin production was observed. Tenogenic gene expressions were highly expressed (p<0.05) at 8% (highest) and 12%, both comparable to tenocytes. In contrast, the osteoblastic, chondrogenic, and adipogenic marker genes appeared to be downregulated. Conclusion. Our study suggests that mechanical loading at 8% strain and 1 Hz provides exclusive tenogenic differentiation; and produced comparable protein and gene expression to primary tenocytes.

scholarly journals CD14 dictates differential activation of mesenchymal stromal cells through AKT, NF-κB and P38 signals

2019 ◽  
Vol 39 (7) ◽  
Author(s):  
Menghui Jiang ◽  
Tianlin Gao ◽  
Yuansheng Liu ◽  
Xue Cao ◽  
Yanting Li ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Expression Patterns ◽  
Mrna Level ◽  
Human Mscs ◽  
Differentiation Potential ◽  
Tlr4 Signaling ◽  
Tlr4 Signaling Pathway ◽  
Different Sources

Abstract Mesenchymal stromal cells (MSCs) widely exist in many tissues and have multiple differentiation potential and immunomodulatory capacities. Recently, MSCs have become promising tools for the treatment of various degenerative disorders and autoimmune diseases. The properties of MSCs could be modified in different microenvironments. Thus, it is important to explore the factors controlling MSC function. The presence of Toll-like receptors (TLRs) in MSCs was demonstrated according to previous studies. Consistently, we also illustrated the expression of TLRs in both murine and human MSCs, and displayed that the expression patterns of TLRs in MSCs from different sources. Furthermore, we explored the role of TLR and TLR signaling pathway in MSCs. Interestingly, activation of TLR4-induced expression of cytokines and some specific genes in MSCs. However, MSCs retained much lower mRNA level compared with macrophages. We explored the expression of CD14 in MSCs from different sources, which played a vital role in TLR4 signaling pathway, and found that MSCs are almost negative for CD14. Moreover, only partial activation of TLR4 signaling pathway was observed in MSCs, with no activation of AKT, NF-κB and P38. Here, in the study we defined TLR expression, function and activation in MSCs, which is critical for designing MSC-based therapies.

scholarly journals Laminin-5 and type I collagen promote adhesion and osteogenic differentiation of animal serum-free expanded human mesenchymal stromal cells

2012 ◽  
Vol 4 (4) ◽  
pp. 36 ◽  
Author(s):  
Falk Mittag ◽  
Eva-Maria Falkenberg ◽  
Alexandra Janczyk ◽  
Marco Götze ◽  
Tino Felka ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Type I Collagen ◽  
Cell Attachment ◽  
Expression Patterns ◽  
Marker Genes ◽  
Type I ◽  
Laminin 5 ◽  
Laminin 1

Mesenchymal stromal cells (MSC) are differentiation competent cells and may generate, among others, mature osteoblasts or chondrocytes<em> in vitro</em> and <em>in vivo</em>. Laminin-5 and type I collagen are important components of the extracellular matrix. They are involved in a variety of cellular and extracellular activities including cell attachment and osteogenic differentiation of MSC. MSC were isolated and expanded using media conforming good medical practice (GMP)-regulations for medical products. Cells were characterized according to the defined minimal criteria for multipotent MSC. MTT- and BrdU-assays were performed to evaluate protein-dependent (laminin-5, laminin-1, type I collagen) metabolic activity and proliferation of MSC. MSC-attachment assays were performed using protein-coated culture plates. Osteogenic differentiation of MSC was measured by protein-dependant mineralization and expression of osteogenic marker genes (osteopontin, alkaline phophatase, Runx2) after three, seven and 28 days of differentiation. Marker genes were identified using quantitative reverse-transcription polymerase chain reaction. Expansion of MSC in GMP-conforming media yielded vital cells meeting all minimal criteria for MSC. Attachment assay revealed a favorable binding of MSC to laminin-5 and type I collagen at a protein concentration of 1-5 fmol/mL. Compared to plastic, osteogenic differentiation was significantly increased by laminin-5 after 28 days of culture (P&lt;0.04). No significant differences in gene expression patterns were observed. We conclude that laminin-5 and type I collagen promote attachment, but laminin-1 and laminin-5 promote osteogenic differentiation of MSC. This may influence future clinical applications.

scholarly journals The Phytoestrogen Genistein Enhances Osteogenesis and Represses Adipogenic Differentiation of Human Primary Bone Marrow Stromal Cells

Endocrinology ◽  
2004 ◽  
Vol 145 (2) ◽  
pp. 848-859 ◽  
Author(s):  
M. Heim ◽  
O. Frank ◽  
G. Kampmann ◽  
N. Sochocky ◽  
T. Pennimpede ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Adipogenic Differentiation ◽  
Marrow Stromal Cells ◽  
Marker Genes ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Genistein Treatment ◽  
17Β Estradiol

Abstract In the present study, we investigated the role of the phytoestrogen genistein and 17β-estradiol in human bone marrow stromal cells, undergoing induced osteogenic or adipogenic differentiation. Profiling of estrogen receptors (ERs)-α, -β1, -β2, -β3, -β4, -β5, and aromatase mRNAs revealed lineage-dependent expression patterns. During osteogenic differentiation, the osteoblast-determining core binding factor-α1 showed a progressive increase, whereas the adipogenic regulator peroxisome proliferator-activated receptor γ (PPARγ) was sequentially decreased. This temporal regulation of lineage-determining marker genes was strongly enhanced by genistein during the early osteogenic phase. Moreover, genistein increased alkaline phosphatase mRNA levels and activity, the osteoprotegerin:receptor activator of nuclear factor-κB ligand gene expression ratio, and the expression of TGFβ1. During adipogenic differentiation, down-regulation in the mRNA levels of PPARγ and CCAAT/enhancer-binding protein-α at d 3 and decreased lipoprotein lipase and adipsin mRNA levels at d 21 were observed after genistein treatment. This led to a lower number of adipocytes and a reduction in the size of their lipid droplets. At d 3 of adipogenesis, TGFβ1 was strongly up-regulated by genistein in an ER-dependent manner. Blocking the TGFβ1 pathway abolished the effects of genistein on PPARγ protein levels and led to a reduction in the proliferation rate of precursor cells. Overall, genistein enhanced the commitment and differentiation of bone marrow stromal cells to the osteoblast lineage but did not influence the late osteogenic maturation markers. Adipogenic differentiation and maturation, on the other hand, were reduced by genistein (and 17β-estradiol) via an ER-dependent mechanism involving autocrine or paracrine TGFβ1 signaling.

Analysis of Key Distinct Biological Characteristics of Human Placenta-Derived Mesenchymal Stromal Cells and Individual Heterogeneity Attributing to Donors

2021 ◽  
pp. 1-13
Author(s):  
Chenxu Tai ◽  
Liudi Wang ◽  
Yuanyuan Xie ◽  
Tianyun Gao ◽  
Feifei Huang ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Human Placenta ◽  
Immune Regulation ◽  
Stromal Cells ◽  
Mononuclear Cells ◽  
Adipogenic Differentiation ◽  
Individual Heterogeneity ◽  
Differentiation Potential ◽  
Different Origin

For potential clinical applications in the future, we investigated the distinct biological features of mesenchymal stromal cells (MSCs) derived from different origin areas of human placenta and individual heterogeneity among different donors. Chorionic plate MSCs (CP-MSCs), amniotic membrane MSCs (AM-MSCs), and decidual plate MSCs (DP-MSCs) were isolated from 5 human placentae and were analyzed in terms of main features of MSCs including surface marker profile, growth, differentiation potential, immune regulation capability, and tubulin acetylation (Ac-tubulin). The expression profile of surface markers in the 3 types of MSCs derived from the 5 donors was relatively stable. Heterogeneity was found in growth, differentiation potential, and immune regulation among MSCs according to the different areas of isolation and different donors. CP-MSCs and AM-MSCs derived from the placentae of donors 1–3 had a higher osteogenic differentiation potential than the corresponding DP-MSCs, but those derived from the placentae of donors 4 and 5 had a markedly lower osteogenic differentiation potential than DP-MSCs. All CP-MSCs derived from donors 1–3 had the highest adipogenic differentiation potential, but CP-MSCs derived from donors 4 and 5 did not show strong capability of adipogenic differentiation. CP-MSCs markedly inhibited the proliferation of peripheral blood mononuclear cells (PBMCs) induced by phytohemagglutinin, whereas AM- and DP-MSCs did not. All MSCs decreased the proportion of CD3+/CD8–/IFN-γ+ Th1 and CD3+/CD8–/IL17+ Th17 cells, but increased the proportion of Treg cells in PBMCs, with individual differences among the 5 donors. DP-MSCs from donors 1 and 2 had higher levels of Ac-tubulin compared with CP- and AM-MSCs. However, the levels of Ac-tubulin in AM-MSCs from donors 3 and 5 were higher than those of the other 2 types of MSCs. Our results revealed that there was tissue-specific heterogeneity among the 3 types of MSCs from different origin tissues of placenta and individual heterogeneity among donors. In future, the pre-selected placenta-derived MSCs with specific biological advantages may improve the curative effect of cell therapy in different situations.

scholarly journals Multi-pronged approach to human mesenchymal stromal cells senescence quantification with a focus on label-free methods

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Weichao Zhai ◽  
Jerome Tan ◽  
Tobias Russell ◽  
Sixun Chen ◽  
Dennis McGonagle ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Preclinical Model ◽  
Label Free ◽  
Current Standard ◽  
Differentiation Potential ◽  
Human Mesenchymal Stromal Cells ◽  
Endogenous Fluorophores

AbstractHuman mesenchymal stromal cells (hMSCs) have demonstrated, in various preclinical settings, consistent ability in promoting tissue healing and improving outcomes in animal disease models. However, translation from the preclinical model into clinical practice has proven to be considerably more difficult. One key challenge being the inability to perform in situ assessment of the hMSCs in continuous culture, where the accumulation of the senescent cells impairs the culture’s viability, differentiation potential and ultimately leads to reduced therapeutic efficacies. Histochemical $$\upbeta $$ β -galactosidase staining is the current standard for measuring hMSC senescence, but this method is destructive and not label-free. In this study, we have investigated alternatives in quantification of hMSCs senescence, which included flow cytometry methods that are based on a combination of cell size measurements and fluorescence detection of SA-$$\upbeta $$ β -galactosidase activity using the fluorogenic substrate, C$${_{12}}$$ 12 FDG; and autofluorescence methods that measure fluorescence output from endogenous fluorophores including lipopigments. For identification of senescent cells in the hMSC batches produced, the non-destructive and label-free methods could be a better way forward as they involve minimum manipulations of the cells of interest, increasing the final output of the therapeutic-grade hMSC cultures. In this work, we have grown hMSC cultures over a period of 7 months and compared early and senescent hMSC passages using the advanced flow cytometry and autofluorescence methods, which were benchmarked with the current standard in $$\upbeta $$ β -galactosidase staining. Both the advanced methods demonstrated statistically significant values, (r = 0.76, p $$\le $$ ≤ 0.001 for the fluorogenic C$${_{12}}$$ 12 FDG method, and r = 0.72, p $$\le $$ ≤ 0.05 for the forward scatter method), and good fold difference ranges (1.120–4.436 for total autofluorescence mean and 1.082–6.362 for lipopigment autofluorescence mean) between early and senescent passage hMSCs. Our autofluroescence imaging and spectra decomposition platform offers additional benefit in label-free characterisation of senescent hMSC cells and could be further developed for adoption for future in situ cellular senescence evaluation by the cell manufacturers.

scholarly journals Bone marrow stromal cells from MDS and AML patients show increased adipogenic potential with reduced Delta-like-1 expression

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marie-Theresa Weickert ◽  
Judith S. Hecker ◽  
Michèle C. Buck ◽  
Christina Schreck ◽  
Jennifer Rivière ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Fate ◽  
Stromal Cells ◽  
Adipogenic Differentiation ◽  
Cell Types ◽  
Bone Marrow Niche ◽  
Differentiation Potential ◽  
Hematopoietic Stem ◽  
Bm Niche

AbstractMyelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are clonal hematopoietic stem cell disorders with a poor prognosis, especially for elderly patients. Increasing evidence suggests that alterations in the non-hematopoietic microenvironment (bone marrow niche) can contribute to or initiate malignant transformation and promote disease progression. One of the key components of the bone marrow (BM) niche are BM stromal cells (BMSC) that give rise to osteoblasts and adipocytes. It has been shown that the balance between these two cell types plays an important role in the regulation of hematopoiesis. However, data on the number of BMSC and the regulation of their differentiation balance in the context of hematopoietic malignancies is scarce. We established a stringent flow cytometric protocol for the prospective isolation of a CD73+ CD105+ CD271+ BMSC subpopulation from uncultivated cryopreserved BM of MDS and AML patients as well as age-matched healthy donors. BMSC from MDS and AML patients showed a strongly reduced frequency of CFU-F (colony forming unit-fibroblast). Moreover, we found an altered phenotype and reduced replating efficiency upon passaging of BMSC from MDS and AML samples. Expression analysis of genes involved in adipo- and osteogenic differentiation as well as Wnt- and Notch-signalling pathways showed significantly reduced levels of DLK1, an early adipogenic cell fate inhibitor in MDS and AML BMSC. Matching this observation, functional analysis showed significantly increased in vitro adipogenic differentiation potential in BMSC from MDS and AML patients. Overall, our data show BMSC with a reduced CFU-F capacity, and an altered molecular and functional profile from MDS and AML patients in culture, indicating an increased adipogenic lineage potential that is likely to provide a disease-promoting microenvironment.

scholarly journals Comprehensive Comparison of Amnion Stromal Cells and Chorion Stromal Cells by RNA-Seq

2021 ◽  
Vol 22 (4) ◽  
pp. 1901
Author(s):  
Brielle Jones ◽  
Chaoyang Li ◽  
Min Sung Park ◽  
Anne Lerch ◽  
Vimal Jacob ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stromal Cells ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Principal Component ◽  
Differentially Expressed ◽  
Inflammatory Factor ◽  
Next Generation Sequencing Technology ◽  
Angiogenic Potential ◽  
Anti Inflammatory

Mesenchymal stromal cells derived from the fetal placenta, composed of an amnion membrane, chorion membrane, and umbilical cord, have emerged as promising sources for regenerative medicine. Here, we used next-generation sequencing technology to comprehensively compare amniotic stromal cells (ASCs) with chorionic stromal cells (CSCs) at the molecular and signaling levels. Principal component analysis showed a clear dichotomy of gene expression profiles between ASCs and CSCs. Unsupervised hierarchical clustering confirmed that the biological repeats of ASCs and CSCs were able to respectively group together. Supervised analysis identified differentially expressed genes, such as LMO3, HOXA11, and HOXA13, and differentially expressed isoforms, such as CXCL6 and HGF. Gene Ontology (GO) analysis showed that the GO terms of the extracellular matrix, angiogenesis, and cell adhesion were significantly enriched in CSCs. We further explored the factors associated with inflammation and angiogenesis using a multiplex assay. In comparison with ASCs, CSCs secreted higher levels of angiogenic factors, including angiogenin, VEGFA, HGF, and bFGF. The results of a tube formation assay proved that CSCs exhibited a strong angiogenic function. However, ASCs secreted two-fold more of an anti-inflammatory factor, TSG-6, than CSCs. In conclusion, our study demonstrated the differential gene expression patterns between ASCs and CSCs. CSCs have superior angiogenic potential, whereas ASCs exhibit increased anti-inflammatory properties.

Sign in / Sign up

Export Citation Format

Share Document