Engineered extracellular matrices with controlled mechanics modulate renal proximal tubular cell epithelialization

PLoS One. 2017 Jul 17;12(7):e0181085. doi: 10.1371/journal.pone.0181085. eCollection 2017.

Abstract

Acute kidney injury (AKI) is common and associated with significant morbidity and mortality. Recovery from many forms of AKI involves the proliferation of renal proximal tubular epithelial cells (RPTECs), but the influence of the microenvironment in which this recovery occurs remains poorly understood. Here we report the development of a poly(ethylene glycol) (PEG) hydrogel platform to study the influence of substrate mechanical properties on the proliferation of human RPTECs as a model for recovery from AKI. PEG diacrylate based hydrogels were generated with orthogonal control of mechanics and cell-substrate interactions. Using this platform, we found that increased substrate stiffness promotes RPTEC spreading and proliferation. RPTECs showed similar degrees of apoptosis and Yes-associated protein (YAP) nuclear localization regardless of stiffness, suggesting these were not key mediators of the effect. However, focal adhesion formation, cytoskeletal organization, focal adhesion kinase (FAK) activation, and extracellular signal-regulated kinase (ERK) activation were all enhanced with increasing substrate stiffness. Inhibition of ERK activation substantially attenuated the effect of stiffness on proliferation. In long-term culture, hydrogel stiffness promoted the formation of more complete epithelial monolayers with tight junctions, cell polarity, and an organized basement membrane. These data suggest that increased stiffness potentially may have beneficial consequences for the renal tubular epithelium during recovery from AKI.

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism
  • Apoptosis / drug effects
  • Cell Nucleus / metabolism
  • Cell Polarity
  • Cell Proliferation / drug effects*
  • Cells, Cultured
  • Collagen Type IV / chemistry
  • Collagen Type IV / metabolism
  • Cytoskeleton / drug effects
  • Epithelial Cells / cytology
  • Epithelial Cells / metabolism
  • Extracellular Signal-Regulated MAP Kinases / antagonists & inhibitors
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Flavonoids / pharmacology
  • Focal Adhesion Protein-Tyrosine Kinases / metabolism
  • Focal Adhesions / drug effects
  • Humans
  • Hydrogel, Polyethylene Glycol Dimethacrylate / chemistry
  • Hydrogel, Polyethylene Glycol Dimethacrylate / pharmacology*
  • Kidney Tubules, Proximal / cytology
  • Microscopy, Fluorescence
  • Phosphoproteins / metabolism
  • Phosphorylation
  • Stress, Mechanical*
  • Tight Junctions
  • Transcription Factors
  • Vinculin / metabolism

Substances

  • Adaptor Proteins, Signal Transducing
  • Collagen Type IV
  • Flavonoids
  • Phosphoproteins
  • Transcription Factors
  • YAP1 (Yes-associated) protein, human
  • Vinculin
  • Hydrogel, Polyethylene Glycol Dimethacrylate
  • Focal Adhesion Protein-Tyrosine Kinases
  • Extracellular Signal-Regulated MAP Kinases
  • 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one