Cystatins in human tear fluid

Abstract

The activities of cysteine proteinases which include several lysosomal cathepsins are controlled by naturally occurring inhibitory proteins termed cystatins. Cystatins occur both intracellularly and extracellularly in various tissue fluids including tears. Tears were collected by the Schirmer paper strip method from healthy volunteers who had no history or signs of external ocular disease. The tear components were extracted from the filter papers, and used to determine the apparent free cystatin activity and cystatin levels of tears, and for immunoblots. Tears were also collected using capillary tubes for the measurements of cystatins. By titrating papain, a cysteine proteinase, of known specific activity with tear fluid, relatively high levels of apparent free cystatin activity were demonstrated in tears: 28.8 +/- 3.47 (S.E.M.) pmols papain inhibited per mg tear protein (n = 9). The concentrations of cystatins in tear samples were measured by an indirect enzyme-linked immunosorbent assay (ELISA) using antibodies against human salivary cystatin S and purified cystatin S as standard. The ELISAS revealed that tears contain high levels of cystatin-like immunoreactive material, amounting to about 10% of tear proteins. In microgram cystatin S/mg protein the values were: right eye: 94.7 +/- 9.9; left eye: 115.5 +/- 14.8; n = 12. Cystatin levels of tears collected using capillary tubes were comparable: 120.7 +/- 19 micrograms/mg protein (n = 10). Immunoblots of tear fluids revealed a protein of about 14,000 molecular weight which reacted with antihuman cystatin SN monoclonal antibodies. Protein(s) of similar molecular weight were visualized using antibodies against human cystatins S and C. Less abundant additional cystatin-like immuno-reactive proteins were detected by using the two latter antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Cystatins in Tears of Patients with Different Corneal Conditions

“…Complex formation of cystatins in tears with increased amounts of cystine proteinases would protect the eye; however, the demonstration of cystatins in the papain assay would be complicated by the masking of the free cystatins. It has been demonstrated that cystatin levels measured by enzyme immunoassay in tears were much higher than that calculated from papain inhibitory assays [8]. This suggests that tears contain immunologically reactive bound cystatins in addition to free cystatins.…”

“…Cystatins firstly were discovered in chicken egg white, but are also widely distributed in mammalian tissue and can be found in body fluids including the tears [7]. Human tears contain a mixture of cystatin S, cystatin SN, and cystatin C, secreted by the lacrimal gland and not derived from corneal or conjunctival epithelial cells [8]. Cystatins in tears most likely have a protective role against the harmful effects of cysteine proteinases.…”

Changes in tear protein profile in keratoconus disease

“…The activity of cysteine proteinases is controlled by their physiological inhibitors, the cystatins, which are known to be generally present in tears. 22 Extracellular cystatins have a protective role against the damaging effects of lysosomal proteinases, which can be secreted under physiological conditions for the degradation and regeneration of tissues, as well as under pathological conditions associated with infection by bacteria or viruses, or with the corneal and conjunctival epithelium. 23 The decreased levels of cystatins that we found in KC tears is indicative of an increase in the degradation of tear proteins, which would explain the decrease that we found in the concentration of total protein in KC tears.…”

Analysis of protein composition and protein expression in the tear fluid of patients with congenital aniridia

“…This function contributes to prevention of uncontrolled proteolysis and tissue damage [29][30][31]. The expression of these proteins in tears is also known to be reduced in several pathological conditions [32][33][34][35].…”