Connexins and cadherins in the cell-cell junctions of corneal fibroblasts and myofibroblasts

Purpose: In the normal cornea, keratocyte interaction is achieved by gap junctions and by tight junctions. After wounding, fibroblasts and α-smooth muscle actin expressing myofibroblasts arise in the cornea. We have used corneal fibroblasts and myofibroblasts to investigate the molecular basis of their cell-cell interaction and for insights into their respective roles in wound healing. Methods: Using Western blot analysis and immunofluorescent microscopy, we determined the relative expression and localization of junctional proteins: connexins and cadherins, and cadherin associated, actin-binding proteins (catenins). Results: In cultured corneal fibroblasts, the gap junction protein, connexin43, was highly expressed and localized to dense maculae; no cadherin was present at the cell-cell contacts. Cultured myofibroblasts showed the opposite pattern: cadherins were highly expressed and localized at the cell-cell contacts; less connexin43 was present and was primarily cytoplasmic. Myofibroblast cadherin, as identified with a pan-cadherin antibody, has molecular mass of 135 kD and weakly reacted with an N-cadherin monoclonal antibody. In addition, cadherin associated cytoplasmic proteins α-, and β-catenins co-localized with cadherin at the cell-cell borders of the myofibroblasts while γ-catenin or plakoglobin did not. Conclusions: The presence of connexin43, rather than cadherin, at the cell-cell borders of corneal fibroblasts supports a primary communication role of junctions in confluent corneal fibroblasts. In contrast, the presence of cadherin at the cell-cell borders of myofibroblasts may provide a site for insertion of actin filaments hence supporting the actin based force generation for effective wound closure.