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Burkhardt S. Schuett, Thomas J. Millar, Chendur Palaniappan; Interaction Of Surfactant Protein (sp) And Keratin With Meibomian Lipid Films. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4246.
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It is likely that proteins contribute to the lipid layer of the tear film. Lung SPs have been detected in tears and keratin is likely to be released from the meibomian ducts. Therefore, the interaction of these proteins with meibomian lipid films has been investigated.
Bovine lung SP-B and SP-C were purified using a combination of extraction and Sephadex LH-20 chromatography. SP-B, SP-C and human keratin (Sigma) were spread either alone or mixed with human meibum on a Langmuir trough at 35°C and pressure-area (Π-A) isocycle profiles were recorded. The penetration of keratin into a spread meibomian lipid layer was also monitored. Measurements were accompanied by fluorescence microscopy of the spread films during the Π-A isocycles.
Lung SP films were very surface active with a maximum pressure of 45mN/m using as little as 1µg protein, but hysteresis on expansion was less pronounced than observed for meibum. The maximum pressure (Πmax) of 12mN/m of pure meibum doubled and tripled if co-seeded with small amounts of SP-B and SP-C (1 or 2μg). Despite this increase in Πmax, no increase in hysteresis was observed. This differs from results seen with other proteins. Keratin alone (1μg) gave a Πmax of 28mN/m indicating that it was also strongly surface active. Its Π-A profiles showed more hysteresis and elasticity compared with pure meibomian films. Keratin (1µg) injected into the subphase penetrated a spread meibomian lipid film as indicated by a gradual increase in pressure, 10mN/m, with time. Penetration of higher concentrations of keratin (7x) was faster and gave an increase in equilibrium pressure (16mN/m). Mixed with meibomian lipids, keratin caused a dramatic increase in hysteresis in the Π-A profile which was in stark contrast to SPs. Micrographs taken after penetration of keratin into the meibomian layer during Π-A profiles illustrate a significantly stiffer film. At medium pressures (8-18 mN/m) liquid-crystal patches appeared which were squeezed out at higher pressures. These are all features not seen in pure meibomian lipid films.
The results of this study strongly indicate that even small amounts of keratin and tear SPs have a strong effect on the surface activity of meibum and may play a role in normal lipid films.
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