scholarly journals Cell culture dimensionality influences mesenchymal stem cell fate through cadherin-2 and cadherin-11

2019 ◽  
Author(s):  
Fiona R Passanha ◽  
Thomas Geuens ◽  
Simon Konig ◽  
Clemens A van Blitterswijk ◽  
Vanessa LS LaPointe
Keyword(s):  
Cell Culture ◽  
Cell Fate ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Positive Influence ◽  
Specific Cell ◽  
Aggregate Culture ◽  
Low Levels ◽  
The Difference ◽  
Cells Cultured

ABSTRACTThe acquisition of a specific cell fate is one of the core aims of tissue engineering and regenerative medicine. Significant evidence shows that aggregate cultures have a positive influence on fate decisions, presumably through cell-cell interactions, but little is known about the specific mechanisms. To investigate the difference between cells cultured as a monolayer and as aggregates, we started by looking at cadherin expression, an important protein involved in cell adhesion, during the differentiation of bone marrow-derived human mesenchymal stem cells (hMSCs) in aggregate and monolayer cultures. We observed that proliferating hMSCs in monolayer culture express cadherin-2 and undergo a switch to cadherin-11 over time, which was not evident in the aggregate cultures. By knocking down cadherin-2 and cadherin-11, we found that both cadherins were required for adipogenic differentiation in a monolayer as well as aggregate culture. However, during osteogenic differentiation, low levels of cadherin-2 were found to be favorable for cells cultured as a monolayer and as aggregates, whereas cadherin-11 was dispensable for cells cultured as aggregates. Together, these results provide compelling evidence for the important role that cadherins play in regulating the differentiation of hMSCs and how this is affected by the dimensionality of cell culture.

scholarly journals Ca2+-activated mitochondrial biogenesis and functions improve stem cell fate in Rg3-treated human mesenchymal stem cells

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Taeui Hong ◽  
Moon Young Kim ◽  
Dat Da Ly ◽  
Su Jung Park ◽  
Young Woo Eom ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Fate ◽  
Mitochondrial Biogenesis ◽  
Biological Activities ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Stem Cell Fate ◽  
Mitochondrial Functions

Abstract Although mitochondrial functions are essential for cell survival, their critical roles in stem cell fate, including proliferation, differentiation, and senescence, remain elusive. Ginsenoside Rg3 exhibits various biological activities and reportedly increases mitochondrial biogenesis and respiration. Herein, we observed that Rg3 increased proliferation and suppressed senescence of human bone marrow-derived mesenchymal stem cells. Osteogenic, but not adipogenic, differentiation was facilitated by Rg3 treatment. Rg3 suppressed reactive oxygen species production and upregulated mitochondrial biogenesis and antioxidant enzymes, including superoxide dismutase. Consistently, Rg3 strongly augmented basal and ATP synthesis-linked respiration with high spare respiratory capacity. Rg3 treatment elevated cytosolic Ca2+ concentration contributing to mitochondrial activation. Reduction of intracellular or extracellular Ca2+ levels strongly inhibited Rg3-induced activation of mitochondrial respiration and biogenesis. Taken together, Rg3 enhances capabilities of mitochondrial and antioxidant functions mainly through a Ca2+-dependent pathway, which improves the proliferation and differentiation potentials and prevents the senescence of human mesenchymal stem cells.

scholarly journals Ca2+-activated mitochondrial biogenesis and functions improve stem cell fate in Rg3-treated human mesenchymal stem cells

2020 ◽  
Author(s):  
Taeui Hong ◽  
Moon Young Kim ◽  
Dat Da Ly ◽  
Su Jung Park ◽  
Young Woo Eom ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Fate ◽  
Mitochondrial Biogenesis ◽  
Biological Activities ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Stem Cell Fate ◽  
Mitochondrial Functions

Abstract Although mitochondrial functions are essential for cell survival, their critical roles in stem cell fate, including proliferation, differentiation, and senescence, remain elusive. Ginsenoside Rg3 exhibits various biological activities and reportedly increases mitochondrial biogenesis and respiration. Herein, we observed that Rg3 increased proliferation and suppressed senescence of human bone marrow-derived mesenchymal stem cells. Osteogenic, but not adipogenic, differentiation was facilitated by Rg3 treatment. Rg3 suppressed reactive oxygen species production and upregulated mitochondrial biogenesis and antioxidant enzymes, including superoxide dismutase. Consistently, Rg3 strongly augmented basal and ATP synthesis-linked respiration with high spare respiratory capacity. Rg3 treatment elevated cytosolic Ca2+ concentration contributing to mitochondrial activation. Reduction of intracellular or extracellular Ca2+ levels strongly inhibited Rg3-induced activation of mitochondrial respiration and biogenesis. Taken together, Rg3 enhances capabilities of mitochondrial and antioxidant functions mainly through a Ca2+-dependent pathway, which improves the proliferation and differentiation potentials and prevents the senescence of human mesenchymal stem cells.

scholarly journals Connexin43 is Dispensable for Early Stage Human Mesenchymal Stem Cell Adipogenic Differentiation But is Protective against Cell Senescence

Biomolecules ◽  
2019 ◽  
Vol 9 (9) ◽  
pp. 474 ◽  
Author(s):  
Qing Shao ◽  
Jessica L. Esseltine ◽  
Tao Huang ◽  
Nicole Novielli-Kuntz ◽  
Jamie E. Ching ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Fate ◽  
Early Stage ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Human Mesenchymal Stem Cell ◽  
Gap Junctional Intercellular Communication ◽  
Cx43 Expression ◽  
Junction Protein

In the last couple of decades, there has been a growing optimism surrounding the potential transformative use of human mesenchymal stem cells (MSCs) and human-induced pluripotent stem cells (iPSCs) for regenerative medicine and disease treatment. In order for this to occur, it is first essential to understand the mechanisms underpinning their cell-fate specification, which includes cell signaling via gap junctional intercellular communication. Here, we investigated the role of the prototypical gap junction protein, connexin43 (Cx43), in governing the differentiation of iPSCs into MSCs and MSC differentiation along the adipogenic lineage. We found that control iPSCs, as well as iPSCs derived from oculodentodigital dysplasia patient fibroblasts harboring a GJA1 (Cx43) gene mutation, successfully and efficiently differentiated into LipidTox and perilipin-positive cells, indicating cell differentiation along the adipogenic lineage. Furthermore, the complete CRISPR-Cas9 ablation of Cx43 from iPSCs did not prevent their differentiation into bona fide MSCs or pre-adipocytes, strongly suggesting that even though Cx43 expression is upregulated during adipogenesis, it is expendable. Interestingly, late passage Cx43-ablated MSCs senesced more quickly than control cells, resulting in failure to properly differentiate in vitro. We conclude that despite being upregulated during adipogenesis, Cx43 plays no detectable role in the early stages of human iPSC-derived MSC adipogenic differentiation. However, Cx43 may play a more impactful role in protecting MSCs from premature senescence.

MicroRNA expression profile during adipogenic differentiation in mouse embryonic stem cells

2011 ◽  
Vol 43 (10) ◽  
pp. 611-620 ◽  
Author(s):  
Julia M. Knelangen ◽  
Mark B. van der Hoek ◽  
Wee-Ching Kong ◽  
Julie A. Owens ◽  
Bernd Fischer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Fate ◽  
Adipogenic Differentiation ◽  
Embryonic Stem Cell Line ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cell ◽  
Specific Cell ◽  
Cell Fate Determination ◽  
Fate Determination

Pluripotent embryonic stem cells (ESC) have the potential to differentiate into any cell type of the three germ layers. Differentiation processes depend on genetic and epigenetic factors. The guidance of cell fate determination by microRNAs (miRs) seems important for embryonic development and cell lineage decisions. MiRs are short, single-stranded, noncoding RNA molecules that regulate through posttranscriptional modulation, a subset of target genes involved in cell differentiation and specific cell function. We have used microarray profiling of miRs in the mouse embryonic stem cell line CGR8. Comparison of the miR profiles of undifferentiated stem cells with mesodermal progenitors cells ( day 5), preadipocytes ( day 10), and adipocytes ( day 21) showed that the expression level of 129 miRs changed (twofold) during adipogenic differentiation. We identified 10 clusters of differentially expressed miRs, which contain putative markers and regulators of mesodermal differentiation and cell fate determination into adipocytes. Notably, the adipocyte-specific miRs 143 and 103 were upregulated from day 10 onward. We have therefore demonstrated and characterized the dynamic profile of miR expression during murine adipogenic differentiation in vitro, including the initial differentiation from ESC via mesenchymal progenitors up to adipocytes. Our findings and experimental approach provide a suitable system to directly interrogate the role of miRs during adipogenic differentiation of embryonic stem cells.

scholarly journals Ca2+-Activated Mitochondrial Biogenesis and Functions Improve Stem Cell Fate in Rg3-Treated Human Mesenchymal Stem Cells

2020 ◽  
Author(s):  
Taeui Hong ◽  
Moon Young Kim ◽  
Dat Da Ly ◽  
Su Jung Park ◽  
Young Woo Eom ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Fate ◽  
Mitochondrial Biogenesis ◽  
Biological Activities ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Stem Cell Fate ◽  
Mitochondrial Functions

Abstract Although mitochondrial functions are essential for cell survival, their critical roles in stem cell fate, including proliferation, differentiation, and senescence, remain elusive. Ginsenoside Rg3 exhibits various biological activities and reportedly increases mitochondrial biogenesis and respiration. Herein, we observed that Rg3 increased proliferation and suppressed senescence of human bone marrow-derived mesenchymal stem cells. Osteogenic, but not adipogenic, differentiation was facilitated by Rg3 treatment. Rg3 suppressed reactive oxygen species production and upregulated mitochondrial biogenesis and antioxidant enzymes, including superoxide dismutase. Consistently, Rg3 strongly augmented basal and ATP synthesis-linked respiration with high spare respiratory capacity. Rg3 treatment elevated cytosolic Ca2+ concentration contributing to mitochondrial activation. Reduction of intracellular or extracellular Ca2+ levels strongly inhibited Rg3-induced activation of mitochondrial respiration and biogenesis. Taken together, Rg3 enhances capabilities of mitochondrial and antioxidant functions mainly through a Ca2+-dependent pathway, which improves the proliferation and differentiation potentials and prevents the senescence of human mesenchymal stem cells.

scholarly journals The Effects of Andrographolide on the Enhancement of Chondrogenesis and Osteogenesis in Human Suprapatellar Fat Pad Derived Mesenchymal Stem Cells

Molecules ◽  
2021 ◽  
Vol 26 (7) ◽  
pp. 1831
Author(s):  
Thitianan Kulsirirat ◽  
Sittisak Honsawek ◽  
Mariko Takeda-Morishita ◽  
Nuttanan Sinchaipanid ◽  
Wanvisa Udomsinprasert ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipogenic Differentiation ◽  
Human Mesenchymal Stem Cells ◽  
Fat Pad ◽  
Alizarin Red ◽  
Lineage Differentiation ◽  
Expression Of Genes ◽  
Dose Dependent ◽  
Oil Red O Staining

Andrographolide is a labdane diterpenoid herb, which is isolated from the leaves of Andrographis paniculata, and widely used for its potential medical properties. However, there are no reports on the effects of andrographolide on the human suprapatellar fat pad of osteoarthritis patients. In the present study, our goal was to evaluate the innovative effects of andrographolide on viability and Tri-lineage differentiation of human mesenchymal stem cells from suprapatellar fat pad tissues. The results revealed that andrographolide had no cytotoxic effects when the concentration was less than 12.5 µM. Interestingly, andrographolide had significantly enhanced, dose dependent, osteogenesis and chondrogenesis as evidenced by a significantly intensified stain for Alizarin Red S, Toluidine Blue and Alcian Blue. Moreover, andrographolide can upregulate the expression of genes related to osteogenic and chondrogenic differentiation, including Runx2, OPN, Sox9, and Aggrecan in mesenchymal stem cells from human suprapatellar fat pad tissues. In contrast, andrographolide suppressed adipogenic differentiation as evidenced by significantly diminished Oil Red O staining and expression levels for adipogenic-specific genes for PPAR-γ2 and LPL. These findings confirm that andrographolide can specifically enhance osteogenesis and chondrogenesis of mesenchymal stem cells from human suprapatellar fat pad tissues. It has potential as a therapeutic agent derived from natural sources for regenerative medicine.

scholarly journals Identification of Novel FNIN2 and FNIN3 Fibronectin-Derived Peptides That Promote Cell Adhesion, Proliferation and Differentiation in Primary Cells and Stem Cells

2021 ◽  
Vol 22 (6) ◽  
pp. 3042
Author(s):  
Eun Ju Lee ◽  
Khurshid Ahmad ◽  
Shiva Pathak ◽  
SunJu Lee ◽  
Mohammad Hassan Baig ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Cell Adhesion ◽  
3D Culture ◽  
Human Mesenchymal Stem Cells ◽  
Cell Types ◽  
Primary Cells ◽  
Cell Culture Techniques ◽  
Culture Techniques ◽  
Promote Cell Adhesion

In recent years, a major rise in the demand for biotherapeutic drugs has centered on enhancing the quality and efficacy of cell culture and developing new cell culture techniques. Here, we report fibronectin (FN) derived, novel peptides fibronectin-based intergrin binding peptide (FNIN)2 (18-mer) and FNIN3 (20-mer) which promote cell adhesion proliferation, and the differentiation of primary cells and stem cells. FNIN2 and 3 were designed based on the in silico interaction studies between FN and its receptors (integrin α5β1, αvβ3, and αIIbβ3). Analysis of the proliferation of seventeen-cell types showed that the effects of FNINs depend on their concentration and the existence of expressed integrins. Significant rhodamine-labeled FNIN2 fluorescence on the membranes of HeLa, HepG2, A498, and Du145 cells confirmed physical binding. Double coating with FNIN2 or 3 after polymerized dopamine (pDa) or polymerized tannic acid (pTA) precoating increased HBEpIC cell proliferation by 30–40 percent, suggesting FNINs potently affect primary cells. Furthermore, the proliferation of C2C12 myoblasts and human mesenchymal stem cells (MSCs) treated with FNINs was significantly increased in 2D/3D culture. FNINs also promoted MSC differentiation into osteoblasts. The results of this study offer a new approach to the production of core materials (e.g., cell culture medium components, scaffolds) for cell culture.

scholarly journals Factors to consider when interrogating 3D culture models with plate readers or automated microscopes

2021 ◽  
Author(s):  
Terry Riss ◽  
O. Joseph Trask
Keyword(s):  
Cell Culture ◽  
Three Dimensional ◽  
3D Culture ◽  
Fluorescent Imaging ◽  
Culture Models ◽  
Assay Methods ◽  
Diffusion Rates ◽  
Cell Culture Models ◽  
Dimensional Cell ◽  
Cells Cultured

AbstractAlong with the increased use of more physiologically relevant three-dimensional cell culture models comes the responsibility of researchers to validate new assay methods that measure events in structures that are physically larger and more complex compared to monolayers of cells. It should not be assumed that assays designed using monolayers of cells will work for cells cultured as larger three-dimensional masses. The size and barriers for penetration of molecules through the layers of cells result in a different microenvironment for the cells in the outer layer compared to the center of three-dimensional structures. Diffusion rates for nutrients and oxygen may limit metabolic activity which is often measured as a marker for cell viability. For assays that lyse cells, the penetration of reagents to achieve uniform cell lysis must be considered. For live cell fluorescent imaging assays, the diffusion of fluorescent probes and penetration of photons of light for probe excitation and fluorescent emission must be considered. This review will provide an overview of factors to consider when implementing assays to interrogate three dimensional cell culture models.

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