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Abstract: PO0637

Gelatin Methacryloyl (GelMA) as a Tuneable Biophysical Environment for the Derivation of Human Induced Pluripotent Stem Cell-Derived Kidney Organoids

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 500 Development, Stem Cells, and Regenerative Medicine

Authors

  • Clerkin, Shane, University College Dublin Conway Institute of Biomolecular and Biomedical Research, Dublin, Ireland
  • Wychowaniec, Jacek K., University College Dublin School of Chemistry and Chemical Biology, Dublin, Ireland
  • Treacy, Niall, University College Dublin Conway Institute of Biomolecular and Biomedical Research, Dublin, Ireland
  • Davis, Jessica L., University College Dublin Conway Institute of Biomolecular and Biomedical Research, Dublin, Ireland
  • Singh, Krutika, University College Dublin School of Chemistry and Chemical Biology, Dublin, Ireland
  • Brougham, Dermot F., University College Dublin School of Chemistry and Chemical Biology, Dublin, Ireland
  • Crean, John, University College Dublin Conway Institute of Biomolecular and Biomedical Research, Dublin, Ireland
Background

The translational utility of hiPSC-derived kidney organoids relies on our ability to comprehensively mimic both the biochemical and biophysical properties of the cellular milieu in vitro. An improved understanding of how the mechanical environment effects the renal progenitor niche during development and how perturbations to such biophysical environments effects cell fate are required. Within this context, Gelatin methacryate (GelMA), a derivative of collagen, represents a mechanically amendable scaffold to probe cell fate dynamics.

Methods

hiPSC-derived kidney organoids were differentiated within photo-crosslinked GelMA hydrogels of defined mechanical strengths. Hydrogels were characterised using rheological analysis and SEM. Enrichment of renal cell types in response to the various mechanical microenvironments was subsequently investigated.

Results

Rheological analysis revealed a diverse stiffness profile range from the formulated hydrogels. Hydrogels comparable to the stiffness of the gastrulation-stage embryo (G’ = 400 Pa), human kidney tissue (G’ = 2.5 kPa) and fibrotic tissue (G’ = 8-10 kPa) were generated. SEM revealed that hydrogel pore size was dependent on starting gelatin concentration in the hydrogel formulations. PCNA and cleaved caspase-3 staining of organoids embedded within scaffolds demonstrated high cell proliferation and viability in all hydrogel constructs by day 26 of differentiation. The formation of glomerular, proximal tubular and distal tubular structures, that were supported by basement membrane and interstitial cells was confirmed in all conditions using qRT-PCR and immunofluorescent analysis. Significant enrichment of MEIS1/2/3+ve interstitial cells was noted in organoids differentiated within stiffer hydrogels. Interstitial expansion and increased extracellular matrix deposition was confirmed using H&E and Masson’s trichrome staining within stiff GelMA scaffolds.

Conclusion

We propose the utility of GelMA hydrogels as faithful extracellular supports for the specification of hiPSC-derived kidney organoids. These scaffolds represent a mechanically tuneable microenvironment to investigate the effects of the biophysical milieu on renal development and disease perturbations.

Funding

  • Government Support – Non-U.S.