Changes in the Transcriptome Profiles of Human Amnion-Derived Mesenchymal Stromal/Stem Cells Induced by Three-Dimensional Culture: A Potential Priming Strategy to Improve Their Properties

Int J Mol Sci. 2022 Jan 13;23(2):863. doi: 10.3390/ijms23020863.


Mesenchymal stromal/stem cells (MSCs) are believed to function in vivo as a homeostatic tool that shows therapeutic properties for tissue repair/regeneration. Conventionally, these cells are expanded in two-dimensional (2D) cultures, and, in that case, MSCs undergo genotypic/phenotypic changes resulting in a loss of their therapeutic capabilities. Moreover, several clinical trials using MSCs have shown controversial results with moderate/insufficient therapeutic responses. Different priming methods were tested to improve MSC effects, and three-dimensional (3D) culturing techniques were also examined. MSC spheroids display increased therapeutic properties, and, in this context, it is crucial to understand molecular changes underlying spheroid generation. To address these limitations, we performed RNA-seq on human amnion-derived MSCs (hAMSCs) cultured in both 2D and 3D conditions and examined the transcriptome changes associated with hAMSC spheroid formation. We found a large number of 3D culture-sensitive genes and identified selected genes related to 3D hAMSC therapeutic effects. In particular, we observed that these genes can regulate proliferation/differentiation, as well as immunomodulatory and angiogenic processes. We validated RNA-seq results by qRT-PCR and methylome analysis and investigation of secreted factors. Overall, our results showed that hAMSC spheroid culture represents a promising approach to cell-based therapy that could significantly impact hAMSC application in the field of regenerative medicine.

Keywords: 3D priming; MSC spheroids; MSC therapeutic properties; RNA sequencing; human amnion-derived mesenchymal stromal/stem cells; regenerative medicine.

MeSH terms

  • Amnion / cytology*
  • Biomarkers
  • Cell Culture Techniques
  • Cell Differentiation
  • Cell Separation
  • Cells, Cultured
  • Computational Biology / methods
  • Epigenesis, Genetic
  • Gene Expression Profiling
  • Gene Expression Regulation
  • Gene Ontology
  • Humans
  • Immunophenotyping
  • Mesenchymal Stem Cells / cytology
  • Mesenchymal Stem Cells / metabolism*
  • Molecular Sequence Annotation
  • Regenerative Medicine
  • Transcriptome*


  • Biomarkers