Protein kinase C subspecies in rabbit corneal epithelium: increased activity of alpha subspecies during wound healing.

Protein kinase C (PKC) has been implicated in cell proliferation and differentiation. Multiple forms of PKC have been isolated, principally from the brain where PKC is most abundant. In rabbit corneal epithelium, two distinct major peaks of PKC activity were resolved by hydroxyapatite column chromatography. Peak 2, with 65% of the total PKC activity, corresponds to alpha-PKC, based on its mobility in the column and Western blot analysis using specific monoclonal antibodies. Peak 1 did not react with either polyclonal or monoclonal antibodies to PKC alpha-, beta-, and gamma-isoforms suggesting the presence of isoforms specific to the corneal epithelium, or of another member of the PKC family. To investigate possible changes in the amounts of the various PKC subspecies during wound healing, the enzyme activities of the isolated subspecies were assayed 2, 5, and 7 days after corneal de-epithelialization. Two days after wounding, by which time the migratory limbal epithelium had covered the denuded area, total PKC activity was unchanged but alpha-PKC activity had increased to 77% of the total activity, compared with 65% in non-wounded epithelium. An increased proportion of alpha-PKC activity was also observed 5 and 7 days after wounding, during which time proliferation of epithelium continued. We hypothesize that alpha-PKC plays a role in long-term responses after injury such as gene expression and corneal epithelial proliferation. Moreover, these studies indicate that the cornea provides a good model of in vivo wound healing for PKC studies.