APL Bioengineering (Sep 2023)

3D photopolymerized microstructured scaffolds influence nuclear deformation, nucleo/cytoskeletal protein organization, and gene regulation in mesenchymal stem cells

  • Francesca Donnaloja,
  • Manuela Teresa Raimondi,
  • Letizia Messa,
  • Bianca Barzaghini,
  • Federica Carnevali,
  • Emanuele Colombo,
  • Davide Mazza,
  • Chiara Martinelli,
  • Lucia Boeri,
  • Federica Rey,
  • Cristina Cereda,
  • Roberto Osellame,
  • Giulio Cerullo,
  • Stephana Carelli,
  • Monica Soncini,
  • Emanuela Jacchetti

DOI
https://doi.org/10.1063/5.0153215
Journal volume & issue
Vol. 7, no. 3
pp. 036112 – 036112-21

Abstract

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Mechanical stimuli from the extracellular environment affect cell morphology and functionality. Recently, we reported that mesenchymal stem cells (MSCs) grown in a custom-made 3D microscaffold, the Nichoid, are able to express higher levels of stemness markers. In fact, the Nichoid is an interesting device for autologous MSC expansion in clinical translation and would appear to regulate gene activity by altering intracellular force transmission. To corroborate this hypothesis, we investigated mechanotransduction-related nuclear mechanisms, and we also treated spread cells with a drug that destroys the actin cytoskeleton. We observed a roundish nuclear shape in MSCs cultured in the Nichoid and correlated the nuclear curvature with the import of transcription factors. We observed a more homogeneous euchromatin distribution in cells cultured in the Nichoid with respect to the Flat sample, corresponding to a standard glass coverslip. These results suggest a different gene regulation, which we confirmed by an RNA-seq analysis that revealed the dysregulation of 1843 genes. We also observed a low structured lamina mesh, which, according to the implemented molecular dynamic simulations, indicates reduced damping activity, thus supporting the hypothesis of low intracellular force transmission. Also, our investigations regarding lamin expression and spatial organization support the hypothesis that the gene dysregulation induced by the Nichoid is mainly related to a reduction in force transmission. In conclusion, our findings revealing the Nichoid's effects on MSC behavior is a step forward in the control of stem cells via mechanical manipulation, thus paving the way to new strategies for MSC translation to clinical applications.