Biomechanical Characterization of Retinal Pigment Epitheliums Derived from hPSCs Using Atomic Force Microscopy

Area of Science:

• Ophthalmology
• Cell Biology
• Biophysics

Background:

• The retinal pigment epithelium (RPE) is vital for photoreceptor survival and homeostasis.
• RPE dysfunction causes retinal degeneration, leading to vision loss (e.g., Age-related macular degeneration, Retinitis Pigmentosa).
• Regenerative medicine using human pluripotent stem cells aims to replace RPE cells, but their biomechanical properties are poorly understood.

Purpose of the Study:

• To characterize the structural and mechanical properties of RPE cells during epithelial formation.
• To investigate the relationship between RPE biomechanics and epithelial development.
• To assess the utility of Atomic Force Microscopy (AFM) for RPE characterization in cell therapy.

Main Methods:

• Cultured RPE cells from four cell lines at different stages of epithelial formation.
• Utilized immunofluorescence to identify apical markers and assess epithelial characteristics.
• Measured transepithelial resistance and cytokine secretion to confirm apico-basal polarity.
• Employed Atomic Force Microscopy (AFM) to scan the apical surface of living and fixed RPE cells.

Main Results:

• RPE monolayers exhibited softening at the apical cell center and stiffening at the cell borders during epithelial formation.
• AFM revealed apical protrusions dependent on the actin network, indicative of microvilli formation.
• Confirmed epithelial characteristics including increased transepithelial resistance and polarized cytokine secretion.

Conclusions:

• RPE cell biomechanical properties change significantly during epithelial development.
• AFM provides valuable insights into RPE structural and mechanical characteristics.
• These findings are essential for understanding RPE function and improving RPE-based cell therapies for retinal diseases.