Gene Expression of Proteases and Protease Inhibitors in the Human Ciliary Epithelium and ODM-2 Cells

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Complementary DNAs (cDNAs), corresponding to the human proteinases cathepsins D and O and proteinase inhibitors α2-macroglobulin and PP5/TFPI-2, have recently been isolated and identified from a subtractive human ciliary body library. In the present study we determined: (i) their pattern of expression in the human eye; (ii) the ability of the ciliary body and/or ciliary epithelial cells to synthesize and secrete cathepsin D and α1-antitrypsin in vitro; and (iii) whether α1-antitrypsin expression in cultured ciliary epithelial cells is modulated by protein kinase C activation. Northern analysis demonstrated that the ciliary body expresses high levels of cathepsins D and O, α2-macroglobulin, α1-antitrypsin and PP5/TFPI-2 transcripts. Western blot analysis and immunoprecipitation experiments with cathepsin D and α1-antitrypsin antibodies indicated that metabolically labeled ciliary body explants and/or ciliary epithelial cells in vitro with35S-methionine, synthesize and secrete these proteins. Cultured nonpigmented ciliary epithelial ODM-2 cells, in response to phorbol-12-myristate 13-acetate (PMA), but not to the non-protein kinase C binding phorbol ester 4 α-phorbol didecanoate (PDBu), elicited up-regulation (up to 5-fold) of transcription, synthesis and secretion of α1-antitrypsin. These results provide in vitro evidence that the ciliary epithelium synthesizes and secretes a selective group of proteinases and proteinase inhibitors detected also in aqueous humor. The expression of at least of one of the proteinase inhibitors, α1-antitrypsin, can be modulated in response to phorbol ester.