Expression of Tear Lipocalin in Rabbit Lacrimal Fluid and Lacrimal Acinar Cells

Abstract

Abstract: : Purpose: Tear lipocalin is a widely distributed member of the lipocalin family. The concentration of tear lipocalin has been found to be decreased in tears of people with abnormal lacrimal gland functions, e.g. in kerato-conjunctivitis sicca or dry eye disease. Homologues of tear lipocalin have been found in different species including human, pig , rat, dog and horse, but the presence of tear lipocalin is yet to be established in rabbit. The presence of tear lipocalin in rabbit lacrimal fluid and isolated acinar cells was investigated. Methods: Pilocarpine-stimulated lacrimal fluid was collected from adult and pubertal New Zealand White rabbits by cannulation of the lacrimal duct using polyethylene PE 10 tubing. Tear fluid was analyzed by SDS PAGE gel electrophoresis and Western blot using a polyclonal anti human tear lipocalin antibody. A protein band of around 20 kDa was isolated from the SDS PAGE gel and the N-terminal sequence was determined. The supernatant from isolated acinar cells was analyzed by SDS PAGE gel electrophoresis and Western blot. Results: N-terminal sequence of this protein showed sufficient amino acid homology to suggest that it is a member of the lipocalin family, but it seems to be divergent in sequence from tear lipocalin of other species. The anti human lipocalin antibody recognized a 20 kDa protein in lacrimal fluid and a 40 kDa protein in the acinar cells. Conclusion: These experiments suggest that tear lipocalin is present in adult and juvenile rabbit lacrimal fluid, however, its sequence seems to be divergent from other species. Lacrimal acinar cells may secrete a larger precursor of this protein. Measurement of tear lipocalin in rabbits could be used as an index of impaired lacrimal gland function.

Keywords: lacrimal gland • protein structure/function • aging