Purified bovine ocular mucin and HA have no surface activity. However, despite having no surface activity in their own right, ocular mucins are likely to be present at the surface of the tear film, where they cause an increase in surface pressure by causing a compression of the lipids (a reorganization of the lipids) and alter the viscoelastic properties at the surface.

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Purpose Tear lipocalin (Tlc) is a major lipid binding protein in tears and is thought to have an important role in stabilizing the Meibomian lipid layer by transferring lipids to it from the aqueous layer or ocular surface, or by adsorbing to it directly. These possible roles have been investigated in vitro using human Tlc. Methods Tlc was purified from human tears by size exclusion chromatography followed by ion exchange chromatography. Three additional samples of the Tlc were prepared by lipidation, delipidation, and relipidation. The lipids extracted from the purified Tlc were analyzed by HPLC-MS followed by fragmentation. Adsorption of these different forms of Tlc to a human Meibomian lipid film spread on the surface of an artificial tear buffer in a Langmuir trough were observed by recording changes in the pressure with time (∏-T profile) and monitoring the appearance of the film microscopically. These results were compared with similar experiments using a bovine Meibomian lipid film. Results The results indicated that Tlc binds slowly to a human Meibomian lipid film compared with lysozyme or lactoferrin, even at 37°C. The adsorption of Tlc to a human Meibomian lipid film was very different from its adsorption to a bovine Meibomian lipid film, indicating the nature of the lipids in the film is critical to the adsorption process. Similarly, the different forms of Tlc had quite distinct adsorption patterns, as indicated both by changes in ∏-T profiles and the microscopic appearance of the films. Conclusions It was concluded that human Tlc was capable of adsorbing to and penetrating into a Meibomian lipid layer, but this process is very complex and depends on both the types of lipids bound to Tlc and the lipid complement comprising the Meibomian lipid film.

Human meibomian lipids form highly compressible, non-collapsible, multilayered liquid films. These lipids have surfactants that allow them to spread across an aqueous subphase. Their surfactant properties are unaffected by increasing divalent cations or hyperosmolarity but are sensitive to temperature. Cooling of meibomian lipids enhances their surfactant properties.

PurposeSebum may contribute to the composition of the tear film lipid layer naturally or as a contaminant artifact from collection. The aims of this study were to determine: if sebum changes the rheology of meibum surface films; if the resonance near 5.2 ppm in the 1H-NMR spectra of sebum is due to squalene (SQ); and if sebum or SQ, a major component of sebum, interacts with human meibum.MethodsHuman meibum was collected from the lid margin with a platinum spatula. Human sebum was collected using lipid absorbent tape. Langmuir trough technology was used to measure the rheology of surface films. Infrared spectroscopy was used to measure lipid conformation and phase transitions. We used 1H-NMR to measure composition and confirm the primary structure of SQ.ResultsThe NMR resonance near 5.2 ppm in the spectra of human sebum was from SQ which composed 28 mole percent of sebum. Both sebum and SQ lowered the lipid order of meibum. Sebum expanded meibum films at lower concentrations and condensed meibum films at higher concentrations. Sebum caused meibum to be more stable at higher pressures (greater maximum surface pressure).ConclusionsPhysiological levels of sebum would be expected to expand or fluidize meibum making it spread better and be more surface active (qualities beneficial for tear film stability). Sebum would also be expected to stabilize the tear film lipid layer, which may allow it to withstand the high shear pressure of a blink.

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The lipid layer of the tear film is associated with preventing evaporative loss. The ability of human Meibomian lipids to reduce evaporation in vitro was tested. Films of human or animal Meibomian lipids or mixtures of cholesterol and phosphatidylcholine were spread on the surface of either artificial buffer or on whole tears and placed on a mass balance that was enclosed in a sealed chamber. The temperature was adjusted to 37 degrees C and gas flow was controlled. Increasing the amounts of Meibomian lipids gave a very small reduction in evaporation. It was concluded from these in vitro experiments that prevention of evaporation from the tear film is not due to the Meibomian lipids alone, but is more likely to be due to a complex interaction between components of the aqueous and the Meibomian lipids.

This is the first-ever report of the antimicrobial role of human meibomian lipids. These lipids possess antimicrobial properties against both Gram-positive and Gram-negative bacteria and are involved in the innate host defense of tears in protecting the ocular surface against microbial pathogens.