Characterization of SV40-transfected cell strains from rabbit keratocytes

The process of corneal wound healing involves the transformation of adjacent corneal keratocytes to myofibroblast-like cells characterized by the development of prominent microfilament bundles containing α-smooth muscle- specific actin (α-SM), a contractile protein thought to be important in mediating wound contraction. Recent studies have shown that the expression of α-SM in cultured corneal keratocytes can be induced by serum and TGF(β1). To study the cellular and molecular mechanisms underlying this transformation process and to begin to identify the role of α-SM in wound contractile events, we generated immortalized rabbit corneal cell strains with extended life by using SV40 transfection. Two unique strains were isolated (TRK-36 and TRK-43). TRK-36, which appears similar to normal corneal keratocytes, maintains a stellate, keratocyte morphology when grown in the absence of serum and transforms to a myofibroblast-like cell when treated with TGF(β1) (1 ng/ml), as indicated by the induced expression of α-SM actin. TRK-43 exhibits features characteristic of myofibroblasts in that it constitutively expresses α-SM actin under serum-free conditions. Both strains show in vitro contraction of collagen gels ≤80% in 24 h in serum-containing medium. Interestingly, under serum-free conditions, TRK-43 cells showed significantly greater contraction of collagen gels compared with those of TRK-36. Overall, the establishment and further study of these cell strains may provide important insights into the molecular mechanisms underlying myofibroblast transformation.