ABSTRACT NO.: 1
Assessment of mucin content in bottlenose dolphin tears
R. Kelleher Davis and D. A. Sullivan
Schepens Eye Research Institute, Harvard Medical School, Boston, MA, USA
Purpose: In terrestrial mammals, the tear film plays an essential role in maintaining the health of the ocular surface, protecting against toxic challenges in the external environment, thereby preserving visual acuity. This protective function is critically dependent on the integrity of the tear film structure, which is comprised of three layers: an underlying glycoprotein (mucin) foundation, a protein-rich aqueous component, and an overlying lipid layer. In a previous study, we determined that the tears of marine mammals do contain proteins, but that the tear film, examined using interferometry, appears to lack the lipid layer found in terrestrial mammals. This finding is quite significant in that it suggests that in sea mammals, an alternative component, in lieu of lipid, promotes the stability of the tear film. This led us to hypothesize that, in the absence of a lipid layer, a mucin foundation, similar to that found in terrestrial mammals, is critical to the integrity of the tear film of marine mammals. This study was carried out to determine whether dolphin tears contain mucins.
Methods: Tear samples from Atlantic bottlenose dolphins (Tursiops truncatus) and humans were analyzed for lectin binding to high molecular weight proteins. Tears from dolphins were collected via capillary suction. Tears from human subjects were collected using Wek-cel sponges. Samples were treated with or without sialidase, which removes glycosylated moieties, and then subjected to electrophoresis on sodium dodecyl sulfate–agarose gels to separate out large proteins. Protein was then transferred by Western blotting onto nitrocellulose membranes and analyzed for the binding of the lectins, peanut agglutinin (PNA) and Sambucus nigra (SNA).
Results: If mucins were present in the tears, it would be expected that PNA would bind to high molecular weight proteins from tear samples treated with sialidase, and that SNA would bind to tear samples not treated with sialidase, for PNA binds to the core of the mucin while SNA binds to glycosylated moieties. In both human and dolphin tears, those treated with sialidase did bind the biotinylated lectin, PNA, but not the SNA. Whereas, untreated tears bound the biotinylated SNA, but not PNA. This differential result confirms the presence of mucins in dolphin tears.
Conclusions: Our finding that large glycoproteins are present in the tears of bottlenose dolphins supports the hypothesis that mucins play a critical role in the protection of the surface of the eyes of marine mammals. Thanks to S. H. Ridgway for tear samples and P. Arguesso and S. Spurr-Michaud for technical advice. Supported by Arey's Pond Boat Yard Inc. and NIH grant EY05612.