Soluble glycoproteins of the lacrimal sac: role in defense with special reference to prolactin-inducible protein (PIP).

Purpose: Glycoproteins play an important role in human mucosal defenses and immunity-related cell-to-cell interactions. The aim of the present study is to investigate the presence and patterns of lacrimal sac glycoproteins involved in defense mechanisms with a special reference to prolactin-inducible protein (PIP). Methods: The study was performed on healthy lacrimal sacs obtained from exenteration samples immediately after surgery and frozen at -80 degrees for subsequent analysis. Four lectins namely Concanavalin A (Con A), Dolichos lablab lectin (DLL), Wheat Germ agglutinin (WGA), and Momordica charantia lectin (MCL) were purified by affinity chromatography. Soluble proteins extract of the lacrimal sac was subjected to chromatography on lectin-affigel columns. Eluted samples from each of the lectin coupled-affigels were analyzed by 10% SDS-PAGE under reducing conditions and the protein bands were visualized using Coomassie blue stain. The protein gel bands were further subjected to mass spectrometry for glycoprotein analysis. Results: Mass spectrometry identified several glycoproteins from the lacrimal sac extracts, with known roles in defense mechanisms. The number of such glycoproteins identified were 9 each from Con A and DLL-I affinity eluted gel bands and 8 and 14 from MCL and WGA affinity eluted gel bands, respectively. Interestingly, PIP was detected in significant proportions in all the eluted gel bands with WGA showing the highest expression. Conclusions: This study is the first step towards the lacrimal sac glycoprotein profiling. PIP could be a major lead for further work on the etiopathogenesis of lacrimal drainage obstructions.

Lysosomal enzymes and mannose 6-phosphate receptors in the lacrimal drainage system: Evidence and its potential implications.

PURPOSE: To investigate the presence and patterns of lysosomal enzymes and mannose 6-phosophate receptor (MPRs) in human lacrimal drainage system. METHODS: The study was performed on healthy lacrimal sacs and nasolacrimal ducts obtained from exenteration samples immediately after surgery and frozen at -80°C for subsequent analysis. Soluble proteins' extract was used for enzyme assays, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (PAGE), native PAGE, activity staining, and western blot analysis. Membrane proteins were separately assessed for detection of mannose 6-phosphate receptors, MPR 46. Sepharose gels, 4-methylumbelliferyl substrates, and antibodies against common lysosomal enzymes and MPRs were used. Enzyme assays were carried out in triplicate to ascertain the results. RESULTS: Differential lysosomal enzyme activities were documented, and among them acid phosphatase and ß-hexosaminidase were found to be high. Western blot analysis using enzyme antibodies and subsequent activity staining confirmed strong signals for moderately expressed enzymes such as fucosidase, glucuronidase, and mannosidase. Membrane extracts demonstrated the presence of MPR 46, which indicates the possible roles of cation-dependent MPRs in lysosomal targeting in human lacrimal drainage system. CONCLUSION: This study provides a proof of principle for the presence of differential lysosomal activity and mannose 6-phosphate ligand transport receptors in human lacrimal drainage system and hypothesizes the potential implications of their dysfunctions.