New cysteine protease inhibitors in human tears and milk and their medical significance are
reviewed in this paper. As protective components against bacterial infection in the eyes, we detected four
kinds of anti-bacterial proteins in normal human tears including lysozyme and three kinds of cysteine
protease inhibitors. Using our reverse zymography of normal tears, three kinds of cysteine protease
inhibitors were found to be 78kDa, 20kDa and 15kDa and were determined to be lactoferrin, Von Ebner's
Gland (VEG) protein and cystatin S, respectively. All of them belong to the cystatin super family and VEG
protein and cystatin S are well known cysteine protease inhibitors. The C-terminus area 17mer peptide, Y
679-k695, of lactoferrin showed strong homology with a common active domain of the cystatin family and
the synthesized peptide showed inhibition of cysteine proteases. Not only were disease-specific changes
found in these inhibitor profiles, but also disease-specific new inhibitors in patients tears with certain
autoimmune diseases. A 35kDa inhibitor, which was detected specifically in tears with Behcet's disease, an
typical autoimmune disease, was determined to be a lacrimal acidic proline-rich protein based on the Nterminus
sequence analysis. A65kDa inhibitor of tears with Harada's autoimmune disease was determined to
be an Ig heavy chain V-III region. In addition, lactoferrin content in Harada's disease was very low. We
found two cathepsin inhibitors in bovine milk using reverse zymography, namely lactoferrin and ?-casein.
The L 133-Q151, in the human ?-casein molecule is the active inhibitory domain. They may play an
important role in antiseptic and anti-infectious functions.
