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Erich Knop, Nadja Knop, Tannin A. Schmidt, Sheila Morrison, Benjamin D. Sullivan, Raheleh Rahimi Darabad, David A. Sullivan; Identification of Lubricin Protein and mRNA Expression at the Human Ocular Surface. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1827.
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Lubricin plays a critical role as a boundary lubricant in articulating joints by protecting cartilaginous surfaces against frictional forces, cell adhesion and protein deposition. We hypothesize that lubricin serves an analogous role on the ocular surface, by being locally synthesized and by protecting the cornea and conjunctiva against significant shear forces generated during an eyelid blink.
Human corneal, conjunctival and control cartilage tissues were fixed with 4% paraformaldehyde, embedded in paraffin, sectioned serially, exposed to antigen retrieval buffers, and then stained for lubricin by immunohistochemistry (IHC) with a rabbit polyclonal anti-human lubricin antibody. Several negative and positive control experiments were performed. Corneal and conjunctival epithelial cells were analysed by reverse transcriptase polymerase chain reaction (RT-PCR) in order to verify the presence of the mRNA. These studies were approved by a Human Studies Committee.
Our results demonstrate that lubricin mRNA is present in and translated and expressed by human corneal and conjunctival epithelium similar to human cartilage. Protein staining appeared stronger in corneal, as compared to bulbar and tarsal conjunctival, epithelial cells. The specificity of lubricin staining was verified by employing a variety of controls. Preincubation of the anti-lubricin antibody with a pure peptide, which had served to generate the antibody, almost completely inhibited IHC staining whereas pre-incubation with a random peptide did not interfere with staining. Replacement of the antibody with either buffer alone or an irrelevant rabbit IgG antibody abolished staining completely.
Our findings support our hypothesis and demonstrate that lubricin is synthesized and expressed by human corneal and conjunctival epithelia at the ocular surface. (Supported by research grants from DFG KN 317/11 and NIH R01EY05612)
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