Establishment and Characterization of TGFBI R124L iPSCs Derived from a Reis–Bücklers Corneal Dystrophy Family

Reis–Bücklers corneal dystrophy (RBCD) is an early-onset TGFBI-related corneal dystrophy associated with the TGFBI R124L variant. It is characterized by recurrent corneal epithelial erosion, hyaline-like deposits involving Bowman’s layer and the anterior stroma, and progressive subepithelial corneal opacity. However, how mutant TGFBI protein (TGFBIp) accumulates in the cornea remains unclear. In this study, we established induced pluripotent stem cells (iPSCs) from two affected members of an RBCD family and one unaffected family member. The patient-derived lines retained the R124L variant, whereas the line established from the unaffected mother did not. All three lines showed pluripotency and trilineage differentiation, had normal karyotypes, and were confirmed by STR analysis to match their donors. After 4 weeks of SEAM induction, all three iPSC lines developed early SEAM-like cultures containing PAX6- and p63-positive cells, demonstrating their capacity for early differentiation toward the corneal epithelial lineage. TGFBIp was also detected in extracellular and pericellular regions by immunofluorescence. In silico analysis detected no major structural difference between WT and R124L TGFBIp in the high-confidence CROPT and FAS1 domain regions, whereas surface analysis showed local changes in electrostatic potential and molecular lipophilicity around residue 124. These lines can be used to study TGFBIp deposition and epithelial–stromal interactions in RBCD.