Extrahepatic synthesis of plasminogen in the human cornea is up-regulated by interleukins-1α and-1β

The avascular cornea has limited access to plasma proteins, including plasminogen, a protein that is synthesized by the Fiver and supplied to most tissues via the blood. Recent experiments by others using plasminogen-deficient mice revealed the importance of plasmin, the active form of plasminogen, for the maintenance of the normal cornea and for corneal wound healing [Kao, Kao, Bugge, Kaufman, Kombrinck, Converse, Good and Degan (1998) Invest. Ophthalmol. Vis. Sci. 39, 502-508; Drew, Kaufman, Kombrinck, Danton, Daugherty, Degen and Bugge (1998) Blood 91, 1616-1624]. In the present experiments, plasmin was identified as a major serine proteinase in the human cornea. The major plasminogen and plasmin forms on non-reducing zymograms and Western blots had M-r values of 76 x 10(3) and 85 x 103, with minor forms of M-r 200 x 10(3), 135 x 10(3), 68 x 10(3) and 45 x 10(3). Angiostatin-like peptides with M(r)s of 48 x 10(3), 45 x 10(3) and 38 x 10(3) were observed which bound to lysine-Sepharose and reacted with anti-plasminogen monoclonal anti- bodies directed towards kringle domains 1-3 of plasminogen. The cornea contained 1.1 +/- 0.15 mu g of plasminogen + plasmin/cornea, or 0.54 +/- 0.05 mu g of plasminogen + plasmin/mg of protein. Cornea conditioned medium contained nine times the amount of plasminogen + plasmin that could be extracted from the cornea. These data suggested that corneal cells, unlike most extrahepatic cells, synthesize plasminogen. The synthesis of plasminogen by the cornea was confirmed by immunoprecipitation of metabolically labelled plasminogen, sequencing of its cDNA obtained by reverse transcriptase-PCR and inhibition of protein synthesis. Interleukins-la and -1 beta stimulated corneal plasminogen synthesis 2-3-fold; however, interleukin-6 decreased corneal plasminogen synthesis by approx. 40 % at early times after addition of the cytokine. By 24 h of culture, no differences were noted in the presence and absence of interleukin-6. Thus the cornea can synthesize plasminogen and regulate its synthesis in response to its environment, including cytokines induced in the cornea by injury and inflammation. Therefore the cornea can control the amount of plasminogen, the precursor of both plasmin and angiostatin.