title: Single-cell differences in matrix gene expression do not predict matrix deposition.
abstract: Mesenchymal stem cells (MSCs) display substantial cell-to-cell heterogeneity, complicating their use in regenerative medicine. However, conventional bulk assays mask this variability. Here we show that both chondrocytes and chondrogenically induced MSCs exhibit substantial mRNA expression heterogeneity. Single-molecule RNA FISH to measure mRNA expression of differentiation markers in single cells reveals that sister cell pairs have high levels of mRNA variability, suggesting that marker expression is not heritable. Surprisingly, this variability does not correlate with cell-to-cell differences in cartilage-like matrix production. Transcriptome-wide analysis suggests that no combination of markers can predict functional potential. De-differentiating chondrocytes also show a disconnect between mRNA expression of the cartilage marker aggrecan and cartilage-like matrix accumulation. Altogether, these quantitative analyses suggest that sorting subpopulations based on these markers would only marginally enrich the progenitor population for 'superior' MSCs. Our results suggest that instantaneous mRNA abundance of canonical markers is tenuously linked to the chondrogenic phenotype at the single-cell level.
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Allison J
authors.initials: AJ
authors.lastname: Cote
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.|Department of Orthopaedic Surgery, McKay Orthopaedic Research Laboratory, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.|Translational Musculoskeletal Research Center, Philadelphia VA Medical Center, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Claire M
authors.initials: CM
authors.lastname: McLeod
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.|Department of Orthopaedic Surgery, McKay Orthopaedic Research Laboratory, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Megan J
authors.initials: MJ
authors.lastname: Farrell
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Patrick D
authors.initials: PD
authors.lastname: McClanahan
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Margaret C
authors.initials: MC
authors.lastname: Dunagin
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Arjun
authors.initials: A
authors.lastname: Raj
authors.affiliation: Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.|Department of Orthopaedic Surgery, McKay Orthopaedic Research Laboratory, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.|Translational Musculoskeletal Research Center, Philadelphia VA Medical Center, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Mauck
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002417:Cattle; D002454:Cell Differentiation / Q000502:physiology; D019902:Chondrocytes / Q000502:physiology*; D005109:Extracellular Matrix; D005786:Gene Expression Regulation / Q000502:physiology*; D017404:In Situ Hybridization, Fluorescence; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D012333:RNA, Messenger
chemical_list: D015415:Biomarkers; D012333:RNA, Messenger