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In order to maintain tissue organization, many animal cells are surrounded by structural molecules that make up the extracellular matrix (ECM). Together, the molecules in the ECM maintain the structural integrity of tissue as well as the remarkable specific properties of certain tissues.
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The extracellular matrix or ECM holds cells together to form a tissue and allows the cells within the tissue to communicate. ECM comprises proteins such as fibronectin, collagen, laminin, etc. The most abundant protein in this space is collagen. Collagen fibers are interwoven with carbohydrate-containing protein molecules called proteoglycans. ECM allows cell migration and provides a structural scaffold at cell adhesion that anchors the cell when the extracellular matrix proteins interact with...
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A Rapid, Scalable Method for the Isolation, Functional Study, and Analysis of Cell-derived Extracellular Matrix
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Single-cell differences in matrix gene expression do not predict matrix deposition.

Allison J Cote1, Claire M McLeod1,2,3, Megan J Farrell1,2

  • 1Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Nature Communications
|March 4, 2016
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Summary
This summary is machine-generated.

Mesenchymal stem cells (MSCs) show significant variation in gene expression, even between related cells. This heterogeneity means common markers don't reliably predict their ability to produce cartilage for regenerative medicine.

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Area of Science:

  • Biomedical Engineering
  • Stem Cell Biology
  • Regenerative Medicine

Background:

  • Mesenchymal stem cells (MSCs) exhibit significant cell-to-cell heterogeneity, which poses challenges for their application in regenerative medicine.
  • Traditional bulk assays obscure this cellular variability, limiting a deeper understanding of MSC function.

Purpose of the Study:

  • To investigate the extent of mRNA expression heterogeneity in chondrocytes and chondrogenically induced MSCs at the single-cell level.
  • To determine the relationship between mRNA expression of differentiation markers and functional cartilage matrix production in single cells.
  • To assess the predictability of functional potential using combinations of canonical markers.

Main Methods:

  • Single-molecule RNA fluorescence in situ hybridization (smFISH) to quantify mRNA expression in individual cells.
  • Analysis of mRNA variability in sister cell pairs to assess heritability of marker expression.
  • Transcriptome-wide analysis to identify potential predictive marker combinations.
  • Quantitative assessment of cartilage-like matrix production in relation to aggrecan mRNA levels.

Main Results:

  • Both chondrocytes and MSCs display substantial mRNA expression heterogeneity, with high variability observed even between sister cells, indicating non-heritable marker expression.
  • This mRNA variability did not correlate with differences in cartilage-like matrix production.
  • Transcriptome-wide analysis revealed that no specific marker combinations could reliably predict functional potential.
  • De-differentiating chondrocytes showed a dissociation between aggrecan mRNA levels and actual matrix accumulation.

Conclusions:

  • Instantaneous mRNA abundance of canonical markers is poorly linked to the chondrogenic phenotype at the single-cell level.
  • Sorting cells based on current marker expression would offer only marginal enrichment for superior MSC subpopulations.
  • The heterogeneity in MSCs and the disconnect between mRNA levels and function necessitate alternative strategies for cell selection in regenerative medicine.