AFM Assessment of the Mechanical Properties of Stem Cells During Differentiation

MRS Advances ◽  
2019 ◽  
Vol 5 (12-13) ◽  
pp. 601-607
Author(s):  
Jie Zou ◽  
Weiwei Wang ◽  
Xianlei Sun ◽  
Wingtai Tung ◽  
Nan Ma ◽  
...  
Keyword(s):  
Mechanical Properties ◽  
Stem Cells ◽  
Flow Cytometry ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Cell Fate ◽  
Stem Cell Differentiation ◽  
Cell Nuclei ◽  
Gene And Protein Expression ◽  
Marrow Mesenchymal Stem Cells

ABSTRACTThe dynamic mechanical force transmitted through microenvironments during tissue formation and regeneration continuously impacts the mechanics of cells and thereby regulates gene and protein expression. The mechanical properties are altered during the process of stem cells differentiating into different lineages. At different stages of differentiation, stem cells display different mechanical properties in response to surrounding microenvironments, which depend on the subcellular structures, especially the cytoskeleton and nucleus. The mechanical properties of the cell nucleus affect protein folding and transport as well as the condensation of chromatin, through which the cell fate is regulated. These findings raise the question as to how cell mechanics change during differentiation. In this study, the mechanical properties of human bone marrow mesenchymal stem cells (hBMSCs) were determined during adipogenic and osteogenic differentiation by atomic force microscopy (AFM). The cytoskeletal structure and the modification of histone were investigated using laser confocal microscope and flow cytometry. The mechanical properties of cell nuclei at different stages of cell differentiation were compared. The stiffness of nuclei increased with time as osteogenesis was induced in hBMSCs. The H3K27me3 level increased during osteogenesis and adipogenesis according to flow cytometry analysis. Our results show conclusively that AFM is a facile and effective method to monitor stem cell differentiation. The measurement of cell mechanical properties by AFM improves our understanding on the connection between mechanics and stem cell fate.

Identification of mesenchymal stem cell differentiation state using dual-micropore microfluidic impedance flow cytometry

2016 ◽  
Vol 8 (41) ◽  
pp. 7437-7444 ◽  
Author(s):  
Hongjun Song ◽  
Jenna M. Rosano ◽  
Yi Wang ◽  
Charles J. Garson ◽  
Balabhaskar Prabhakarpandian ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Mesenchymal Stem Cell ◽  
Electrical Impedance ◽  
Stem Cell Differentiation ◽  
Non Invasive ◽  
Mesenchymal Stem Cell Differentiation

A dual-micropore-based microfluidic electrical impedance flow cytometer for non-invasive identification of the differentiation state of mesenchymal stem cells.

scholarly journals ID: 1037 Effect of fluorescent nanodiamonds on umbilical cord mesenchymal stem cell differentiation into hepatocyte-like cell

2017 ◽  
Vol 4 (S) ◽  
pp. 115
Author(s):  
Trung Kien Do ◽  
Nguyen Thi Thanh Nga ◽  
Nguyen Quynh Anh ◽  
Dinh Minh Pham ◽  
Chu Hoang Ha
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Umbilical Cord ◽  
Stem Cell Differentiation ◽  
Hepatic Differentiation ◽  
Mesenchymal Stem Cell Differentiation ◽  
Or Gene

Fluorescent nanodiamond (FND) indicated that it has excellent biocompatibility and photostability,so it well suited for long-term labeling and tracking of stem cells. There are many reports concerning the factors controlling stem cell differentiation. However, still little knowledge about the biomaterials properties influence stem cell alive, growth and differentiation processing. In this study, we evaluate the effect of fluorescent nanodiamond in in vitro culture and differentiation of ucMSC into hepatocyte-like cell. Mesenchymal stem cells (MSCs) were isolated from the umbilical cord (UC) and CD markers were analyzed by flow cytometry and genes expression. For hepatic differentiation of UC-MSCs, cells were induced with HGF and DMSO treated. FND was supply in the experimental group which 10 g/ml in 4 hours. The FND uptake was detected of fluorescence intensity of FND in cells by flow cytometry and laser scan microscopy. The effect of FND into UCMSCs was not only evaluated by the cell alive and growth assay but also effective differentiation throughout morphology charging or gene expression levels of AFP, ALB, and HNF4 were determined by RT-PCR and real-time PCR. The result showed that the FND was well uptake in UCMSCs. It was no affected into ability of the cell alive and growth. The existence of FNDs does not disturb the functions of UC-MSCs differentiation into hepatocyte-like cell. FND can be utilized for the labeling and tracking of UC-MSCs and hepatocyte-like cell in homing research.

Guiding Stem Cell Differentiation Into Neural Lineages With Tunable Collagen Biomaterials

2009 ◽  
Author(s):  
Gary A. Monteiro ◽  
David I. Shreiber
Keyword(s):  
Mechanical Properties ◽  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Type I Collagen ◽  
Striated Muscle ◽  
Stem Cell Differentiation ◽  
Type I ◽  
Collagen Gels ◽  
Biomaterial Scaffolds

The long-term objective of this research is to develop tunable collagen-based biomaterial scaffolds for directed stem cell differentiation into neural lineages to aid in CNS diseases and trauma. Type I collagen is a ubiquitous protein that provides mechanostructural and ligand-induced biochemical cues to cells that attach to the protein via integrin receptors. Previous studies have demonstrated that the mechanical properties of a substrate or tissue can be an important regulator of stem cell differentiation. For example, the mechanical properties polyacrylamide gels can be tuned to induce neural differentiation from stem cells [1, 2]. Mesenchymal stem cells (MSCs) cultured on ployacrylamide gels with low elastic modulus (0.1–1 kPa) resulted in a neural like population. MSCs on 10-fold stiffer matrices that mimic striated muscle elasticity (Emuscle ∼8–17 kPa) lead to spindle-shaped cells similar in shape to myoblasts. Still stiffer gels (25–40 kPa) resulted in osetoblast differentiation. Based on these observations, collagen gels may provide an ideal material for differentiation into neural lineages because of their low compliance.

scholarly journals PASsing on Signals: PAS Kinase (PASK)-mTOR Signaling Conveys Nutrient Sufficiency Signals to Epigenetic (COMPASS) Complexes to Activate Stem Cell Differentiation Program (P15-009-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Chintan Kikani ◽  
Michael Xiao ◽  
Xiaoying Wu ◽  
Jared Rutter
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Cell Fate ◽  
Early Stage ◽  
Stem Cell Differentiation ◽  
Mtor Signaling ◽  
Kinase Assay ◽  
Cell Functions ◽  
Nutrient Signaling

Abstract Objectives To determine how nutrient signaling impacts stem cell functions Methods PASK phosphorylation: We measured in situ phosphorylation of PASK by metabolic 32P labeling of stem cells expressing WT or mutant versions of PASK. PASK Activation: PASK activation was measured using in vitro kinase assay using radio-labeled ATP. Myogenesis: Myogenesis was measured by immunohistological, and immunofluorescent analysis of differentiating muscle stem cells. Antibodies used were: Myogenin (F5D-Developmental Hybridoma), MF20 (Myosin heavy chain), Pax7 and MyoD. Results Stem cell fate in the tissue niche is intimately connected with intracellular metabolic state and the extracellular hormonal stimulations. We have identified PAS domain containing Kinase (PASK) as a stem cell enriched protein kinase that is required for establishment of the differentiation program in many stem cell paradigms. For this function, PASK phosphorylates Wdr5, a member of the COMPASS family of histone methyltransferases, to activate the epigenetic processes required for the stem cell differentiation (eLife, 2016). Here we show that a master nutrient sensor, mTOR complex 1 (mTORC1) activates PASK via multi-site phosphorylation during stem cell differentiation. This phosphorylation of PASK by mTORC1 is required for epigenetic activation of the Myogenin transcription, exit from the self-renewal and induction of the myogenesis program. Our data suggest that mTORC1-PASK signaling generates MyoG + committed myoblasts (epigenetically - an early stage of myogenesis), whereas mTORC1-S6K1 signaling is required for myoblast fusion (translationally - later stage of myogenesis). Conclusions Our discoveries show that nutrient signaling can partition stem cell fates during different stages of the myogenesis program downstream of mTOR signaling via activation of two distinct protein kinases. Funding Sources NIH R01 (Chintan Kikani), HHMI (Jared Rutter) Supporting Tables, Images and/or Graphs

scholarly journals The Mechanical Interplay Between Differentiating Mesenchymal Stem Cells and Gelatin-Based Substrates Measured by Atomic Force Microscopy

2021 ◽  
Vol 9 ◽  
Author(s):  
Hongxu Meng ◽  
Tina T. Chowdhury ◽  
Núria Gavara
Keyword(s):  
Mechanical Properties ◽  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Stem Cell Differentiation ◽  
Multiple Time ◽  
Time Points ◽  
Surrounding Matrix ◽  
Multiple Time Points ◽  
Highly Correlated

Traditional methods to assess hMSCs differentiation typically require long-term culture until cells show marked expression of histological markers such as lipid accumulation inside the cytoplasm or mineral deposition onto the surrounding matrix. In parallel, stem cell differentiation has been shown to involve the reorganization of the cell’s cytoskeleton shortly after differentiation induced by soluble factors. Given the cytoskeleton’s role in determining the mechanical properties of adherent cells, the mechanical characterization of stem cells could thus be a potential tool to assess cellular commitment at much earlier time points. In this study, we measured the mechanical properties of hMSCs cultured on soft gelatin-based hydrogels at multiple time points after differentiation induction toward adipogenic or osteogenic lineages. Our results show that the mechanical properties of cells (stiffness and viscosity) and the organization of the actin cytoskeleton are highly correlated with lineage commitment. Most importantly, we also found that the mechanical properties and the topography of the gelatin substrate in the vicinity of the cells are also altered as differentiation progresses toward the osteogenic lineage, but not on the adipogenic case. Together, these results confirm the biophysical changes associated with stem cell differentiation and suggest a mechanical interplay between the differentiating stem cells and their surrounding extracellular matrix.

scholarly journals Using single-cell entropy to describe the dynamics of reprogramming and differentiation of induced pluripotent stem cells

2020 ◽  
Vol 34 (30) ◽  
pp. 2050288
Author(s):  
Y. Ye ◽  
Z. Yang ◽  
M. Zhu ◽  
J. Lei
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Single Cell ◽  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Stem Cell Differentiation ◽  
Cell Level ◽  
Macroscopic Variable ◽  
Induced Pluripotent

Induced pluripotent stem cells (iPSCs) provide a great model to study the process of stem cell reprogramming and differentiation. Single-cell RNA sequencing (scRNA-seq) enables us to investigate the reprogramming process at single-cell level. Here, we introduce single-cell entropy (scEntropy) as a macroscopic variable to quantify the cellular transcriptome from scRNA-seq data during reprogramming and differentiation of iPSCs. scEntropy measures the relative order parameter of genomic transcriptions at single cell level during the process of cell fate changes, which show increase tendency during differentiation, and decrease upon reprogramming. Hence, scEntropy provides an intrinsic measurement of the cell state, and can be served as a pseudo-time of the stem cell differentiation process. Moreover, based on the evolutionary dynamics of scEntropy, we construct a phenomenological Fokker-Planck equation model and the corresponding stochastic differential equation for the process of cell state transitions during pluripotent stem cell differentiation. These equations provide further insights to infer the processes of cell fates changes and stem cell differentiation. This study is the first to introduce the novel concept of scEntropy to quantify the biological process of iPSC, and suggests that the scEntropy can provide a suitable macroscopic variable for single cells to describe cell fate transition during differentiation and reprogramming of stem cells.

scholarly journals Osteogenic differentiation of follicular stem cells on nano-Saghez scaffold containing BMP2

2019 ◽  
Vol 14 (1) ◽  
Author(s):  
Hananeh Bayat ◽  
Hassan Shahabinejad ◽  
Mohammad Bayat ◽  
Sadegh Shirian ◽  
Abdolreza Mohamadnia ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Stem Cell ◽  
Young’S Modulus ◽  
Young's Modulus ◽  
Stem Cell Differentiation ◽  
Activity Measurement ◽  
Wisdom Tooth ◽  
Morphological Studies ◽  
Gene And Protein Expression

Abstract Background Bone tissue is one of the tissues that are capable of self-regeneration. However, bone self-regeneration is defeated in the case of broad lesion of bone structure. Isolated stem cells from wisdom tooth follicles can potentially differentiate into ectodermal and mesodermal cells. Saghez is a natural substance that has been extracted from Pistacia terebinthus with unique features, such as high temperature and mechanical stability, adhesive structure, biocompatibility, and anti-neoplastic properties. Methods In this study, Saghez-encapsulated BMP2 was applied as a scaffold for wisdom tooth follicle stem cell differentiation into the osteocyte. A total of three wisdom tooth follicles were obtained for stem cell isolation. For verification of differentiation of the isolated stem cells into osteocyte and adipocyte, Oil Red and Alizarin staining were applied, respectively. Moreover, mesenchymal stem cells were distinguished by profiling their cell surface markers, includingCD73, CD90, CD44, and CD105, by flow cytometry. Saghez scaffold loaded with BMP2 factor was prepared using sol-gel method. Four experimental groups were considered in this study: cells seeded on BMP2 encapsulated in Saghez scaffold, Saghez scaffold, osteogenic medium, and DMEM medium. Results Mechanical properties of Saghez scaffold, including tensile Young’s modulus, ultimate tensile stress, compression Young’s modulus, and complex shear modulus, were 19 MPa, 32 MPa, 0.42 MPa, and 0.9 MPa, respectively. The porosity of the scaffold was 70–140 μm, and the percentage of porosity was 75–98%. The results of flow cytometry studies indicated that CD44, CD73, CD90, and CD105 were positively expressed on the membrane of the tooth follicles’ stem cell. The results indicated that the rate of differentiation of the follicle stem cells into osteocyte was the highest in the Saghez-BMP2 scaffold containing differentiation medium groups. These findings were verified by morphological studies, osteoblast and osteocalcin gene and protein expression investigations, and alkaline phosphatase activity measurement. The highest osteopontin and osteocalcin genes expression levels (1.7 and 1.9) were seen in positive control, followed by DMEM + differentiation factor (1.5 and 1.6), scaffold + BMP2 (1.2 and 1.4), DMEM + stem cell (1 and 1) and scaffold (0.4 and 0.5), and negative control respectively. Conclusion This study provides a novel system for differentiation of the stem cell into osteocytes. The results of this study suggest that loaded BMP2 in Saghez scaffold possibly acts as an osteocyte differentiator factor.

scholarly journals Using single-cell entropy to describe the dynamics of reprogramming and differentiation of induced pluripotent stem cells

2020 ◽  
Author(s):  
Yusong Ye ◽  
Zhuoqin Yang ◽  
Meixia Zhu ◽  
Jinzhi Lei
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Single Cell ◽  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Stem Cell Differentiation ◽  
Cell Level ◽  
Macroscopic Variable ◽  
Induced Pluripotent

AbstractInduced pluripotent stem cells (iPSCs) provide a great model to study the process of stem cell reprogramming and differentiation. Single-cell RNA sequencing (scRNA-seq) enables us to investigate the reprogramming process at single-cell level. Here, we introduce single-cell entropy (scEntropy) as a macroscopic variable to quantify the cellular transcriptome from scRNA-seq data during reprogramming and differentiation of iPSCs. scEntropy measures the relative order parameter of genomic transcriptions at single cell level during the process of cell fate changes, which show increase tendency during differentiation, and decrease upon reprogramming. Hence, scEntropy provides an intrinsic measurement of the cell state, and can be served as a pseudo-time of the stem cell differentiation process. Moreover, based on the evolutionary dynamics of scEntropy, we construct a phenomenological Fokker-Planck equation model and the corresponding stochastic differential equation for the process of cell state transitions during pluripotent stem cell differentiation. These equations provide further insights to infer the processes of cell fates changes and stem cell differentiation. This study is the first to introduce the novel concept of scEntropy to quantify the biological process of iPSC, and suggests that the scEntropy can provide a suitable macroscopic variable for single cells to describe cell fate transition during differentiation and reprogramming of stem cells.

scholarly journals DYRK1A Negatively Regulates CDK5-SOX2 Pathway and Self-Renewal of Glioblastoma Stem Cells

2021 ◽  
Vol 22 (8) ◽  
pp. 4011
Author(s):  
Brianna Chen ◽  
Dylan McCuaig-Walton ◽  
Sean Tan ◽  
Andrew P. Montgomery ◽  
Bryan W. Day ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Critical Role ◽  
Stem Cell Differentiation ◽  
Neural Progenitor Cell ◽  
Cellular Heterogeneity ◽  
Differentiation Potential ◽  
Glioblastoma Stem Cells ◽  
Glioblastoma Stem Cell

Glioblastoma display vast cellular heterogeneity, with glioblastoma stem cells (GSCs) at the apex. The critical role of GSCs in tumour growth and resistance to therapy highlights the need to delineate mechanisms that control stemness and differentiation potential of GSC. Dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) regulates neural progenitor cell differentiation, but its role in cancer stem cell differentiation is largely unknown. Herein, we demonstrate that DYRK1A kinase is crucial for the differentiation commitment of glioblastoma stem cells. DYRK1A inhibition insulates the self-renewing population of GSCs from potent differentiation-inducing signals. Mechanistically, we show that DYRK1A promotes differentiation and limits stemness acquisition via deactivation of CDK5, an unconventional kinase recently described as an oncogene. DYRK1A-dependent inactivation of CDK5 results in decreased expression of the stemness gene SOX2 and promotes the commitment of GSC to differentiate. Our investigations of the novel DYRK1A-CDK5-SOX2 pathway provide further insights into the mechanisms underlying glioblastoma stem cell maintenance.

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