Gene expression of proteases and protease inhibitors in the human ciliary epithelium and ODM-2 cells

Complementary DNAs (cDNAs), corresponding to the human proteinases cathepsins D and O and proteinase inhibitors alpha 2-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 alpha 1-antitrypsin in vitro; and (iii) whether alpha 1-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, alpha 2-macroglobulin, alpha 1-antitrypsin and PP5/TFPI-2 transcripts, Western blot analysis and immunoprecipitation experiments with cathepsin D and alpha 1-antitrypsin antibodies indicated that metabolically labeled ciliary body explants and/or ciliary epithelial cells in vitro with S-35-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 alpha-phorbol didecanoate (PDBu), elicited upregulation (up to 5-fold) of transcription, synthesis and secretion of alpha 1-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, alpha 1-antitrypsin, can be modulated in response to phorbol ester. (C) 1997 Academic Press Limited.