A Recoverable Curved Multilayer Cornea-on-a-Chip with DRGO-Derived Neurites

Background: In vitro corneal models that combine curved geometry, multilayer tissue organization, neural components, air-liquid interface (ALI) culture, and perfusion remain limited. Methods: We developed a recoverable curved multilayer cornea-on-a-chip (CMCoC) using a two-component polydimethylsiloxane device with a 7.8-mm-radius dome. The construct comprised an HCE-T-derived epithelial layer, human corneal stromal cells embedded in plastically compressed type I collagen, an HCEC-B4G12-containing endothelial compartment, and circumferential human induced pluripotent stem cell-derived dorsal root ganglion organoids (DRGOs). After assembly, constructs were perfused for 7 days to permit peripheral neurite extension, followed by epithelial seeding and 14 days of ALI culture with continued basal perfusion. Results: Retrieved tissues showed a visible curved profile. Immunofluorescence demonstrated the intended epithelial-stromal-endothelial organization, a multilayered epithelium, and extensive βIII-tubulin-positive DRGO-derived neurites associated with the epithelial compartment. The CMCoC had a thinner epithelial compartment than the mouse corneal reference and a higher βIII-tubulin-positive area fraction. The other three neural morphometric comparisons were not statistically significant after adjustment. Conclusions: The CMCoC combines recoverable curved tissue architecture, multicellular organization, and DRGO-derived neurites, providing a basis for future studies of corneal tissue interactions.