Elsevier

Experimental Eye Research

Volume 116, November 2013, Pages 79-85
Experimental Eye Research

Lipid order, saturation and surface property relationships: A study of human meibum saturation

https://doi.org/10.1016/j.exer.2013.08.012Get rights and content

Highlights

  • Saturation increased the phase transition temperature in human meibum by over 20 °C.

  • Saturation, more than other lipid species correlated with the phase transition temperature.

  • Saturated meibum films were ridged and elastic compared with native meibum films.

Abstract

Tear film stability decreases with age however the cause(s) of the instability are speculative. Perhaps the more saturated meibum from infants may contribute to tear film stability. The meibum lipid phase transition temperature and lipid hydrocarbon chain order at physiological temperature (33 °C) decrease with increasing age. It is reasonable that stronger lipid–lipid interactions could stabilize the tear film since these interactions must be broken for tear break up to occur. In this study, meibum from a pool of adult donors was saturated catalytically. The influence of saturation on meibum hydrocarbon chain order was determined by infrared spectroscopy. Meibum is in an anhydrous state in the meibomian glands and on the surface of the eyelid. The influence of saturation on the surface properties of meibum was determined using Langmuir trough technology. Saturation of native human meibum did not change the minimum or maximum values of hydrocarbon chain order so at temperatures far above or below the phase transition of human meibum, saturation does not play a role in ordering or disordering the lipid hydrocarbon chains. Saturation did increase the phase transition temperature in human meibum by over 20 °C, a relatively high amount. Surface pressure–area studies showing the late take off and higher maximum surface pressure of saturated meibum compared to native meibum suggest that the saturated meibum film is quite molecularly ordered (stiff molecular arrangement) and elastic (molecules are able to rearrange during compression and expansion) compared with native meibum films which are more fluid agreeing with the infrared spectroscopic results of this study. In saturated meibum, the formation of compacted ordered islands of lipids above the surfactant layer would be expected to decrease the rate of evaporation compared to fluid and more loosely packed native meibum. Higher surface pressure observed with films of saturated meibum compared to native meibum suggests greater film stability especially under the high shear stress of a blink.

Introduction

Studies over the past two centuries suggest that the functions of lipids on the surface of the tear film are to dam, lubricate, and stabilize the tear film to allow for proper refraction, prevention of evaporation, degradation of mucinic clots, provide antibacterial effect, and suppress exposure to UV rays (Murube, 2012). The major source of tear lipids are the meibomian glands and to a lesser degree the glands of Zeis and Moll (Murube, 2012). The composition (Borchman et al., 2007b, Butovich, 2008, Nagyova and Tiffany, 1999, Pucker and Nichols, 2012, Wollensak et al., 1990) and physical properties (Borchman et al., 2007a, Borchman et al., 2007b, Nagyova and Tiffany, 1999) of tear lipids are slightly different compared to meibum lipids. The biophysical characteristics (Borchman et al., 2011, Borchman et al., 2010b, Foulks et al., 2010) and thus the composition of meibum changes with age (Borchman et al., 2010a, Borchman et al., 2012a, Oshima et al., 2009) and meibomian gland dysfunction (Borchman et al., 2012b, Borchman et al., 2010c, Foulks and Borchman, 2010, Foulks et al., 2013, Foulks et al., 2010, Green-Church et al., 2011, Oshima et al., 2009, Shrestha et al., 2011).

Meibum lipids undergo a broad phase transition between 25 °C and 40 °C that is centered near 30 °C (Borchman et al., 2011, Borchman et al., 2007a, Borchman et al., 2010b, Borchman et al., 2007b). The phase transition range is close to the temperature range experienced by the surface of the cornea when the ambient temperature changes from −20 °C to 40 °C (Geiser et al., 2004, Morgan et al., 1995, Shellock and Schatz, 1992). The phase transition at physiological temperature may have functional significance since lipids that are at their phase transition temperature inhibit the rate of evaporation more effectively than lipids that are above or below their phase transition (Rantamäki et al., 2013). Meibum lipids are said to undergo a gel to liquid crystalline phase transition because unlike melting, meibum lipids are not completely solid (74% ordered) and in a gel state below the transition temperature and the lipid hydrocarbon chains are packed orthorombically (Borchman et al., 2011, Borchman et al., 2007a, Borchman et al., 2007b). Above the transition temperature meibum lipids are not completely liquid (73% disordered) and in a liquid crystalline state (Borchman et al., 2011, Borchman et al., 2007a, Borchman et al., 2007b). Meibum disorder was measured by quantifying the percentage of gauche rotamers (conformation) in the hydrocarbon chains which cause kinks in the hydrocarbon chains leading to the disruption of positive van der Wall's interactions between hydrocarbon chains. Because the temperature of the eye surface is near to the phase transition temperature of meibum, elevating the temperature above physiological temperature causes the meibum to be more disordered. This is relevant because meibum lipid hydrocarbon chain disorder was indirectly related to the delivery of lipid to the eyelid margin (Borchman et al., 2007a). The conformational changes observed in the hydrocarbon chains of meibum lipid with increasing temperature suggest that the observed therapeutic increased delivery of meibum (Nagymihalyi et al., 2004) with eyelid heating, one of the oldest treatments for dry eye (Harrison and Lawlor, 1998, Mori et al., 1999, Thygeson, 1946), could be related to the increased disorder in the packing of the hydrocarbon tails.

Tear film stability decreases with age (Bacher, 2010, Cho and Yap, 1993, Craig and Tomlinson, 1998, Guillon and Maissa, 2010, Henderson and Prough, 1950, Isenberg et al., 2003, Lavezzo et al., 2008, Lawrenson et al., 2005, Maissa and Guillon, 2010, Mantelli et al., 2007, Mohidin et al., 2002, Ozdemir and Temizdemir, 2010, Sforza et al., 2008, Sun et al., 1997) however the cause(s) of the instability are speculative. Some correlations between the biophysical properties of meibum and age have been made. The meibum lipid phase transition temperature and lipid hydrocarbon chain order at 33 °C decreases with increasing age (Borchman et al., 2010b). It is reasonable that stronger lipid–lipid interactions could stabilize the tear film since these interactions must be broken for tear break up to occur. One would expect water penetration through the lipid layer, necessary for evaporation to occur, would be inhibited by stronger lipid–lipid interactions. Meibum from infants with a very stable tear film are more saturated than those of adults (Borchman et al., 2010a, Borchman et al., 2012a), and lipid saturation is related to the phase transition temperature (Borchman et al., 2011, Borchman et al., 1999, Borchman et al., 2004) which is also related to lipid order (fluidity) at physiological temperature for many natural and synthetic membranes (Borchman et al., 2011, Borchman et al., 2010b).

Tear film stability not only decreases with age as discussed above, but also decreases with meibomian gland dysfunction (Foulks and Borchman, 2010, Green-Church et al., 2011) and like age-related changes, the causes are speculative. The lipid phase transition temperature and order of meibum from donors with meibomian gland dysfunction are higher compared to normal age matched donors (Borchman et al., 2011, Foulks et al., 2013, Foulks et al., 2010). The rise in order is not due to hydrocarbon chain saturation changes as it is with age since lipid saturation does not change with meibomian gland dysfunction (MGD) (Borchman et al., 2012b, Oshima et al., 2009), or aqueous-deficient dry eye (Joffre et al., 2008).

In this study, meibum from a pool of adult donors was saturated catalytically and the influence of saturation on meibum hydrocarbon chain order was determined by infrared spectroscopy and the influence of saturation on the pressure/area curves was determined using Langmuir trough technology.

Section snippets

Materials

Silver chloride windows for infrared spectroscopy were obtained from Crystran Limited, Poole, United Kingdom. Oleyl oleate (OO) and platinum(IV) oxide were obtained from the Sigma Chemical Company (St. Louis, MO).

Diagnosis of normal status

Normal status was assigned when the patient's meibomian gland orifices showed no evidence of keratinization or plugging with turbid or thickened secretions and no dilated blood vessels were observed on the eyelid margin. Normal donors did not recall having dry eye symptoms. Written

Human material

Human donors of meibum were recruited from the members and associates of Professor Borchman's laboratory at the Kentucky Lions Eye Center, Louisville Kentucky. Meibum from four donors was pooled: 37M, 56F, 58M and 67M where the numbers are the donors age, M is male, F is female. All donors were Caucasian.

Human meibum lipid phase transitions and the infrared CH stretching region

The CH2 stretching bands were predominant in the infrared spectra of meibum due to the large number of CH2 groups in the lipid hydrocarbon chains (data not shown) (Borchman et al., 2011,

Discussion

Saturation of native human meibum did not change the minimum or maximum values of ṽsym, so at temperatures far above or below the phase transition of human meibum, saturation does not play a role in ordering or disordering the lipid hydrocarbon chains. Saturation did increase the phase transition temperature in human meibum by over 20 °C, a relatively high amount. Because the phase transition of meibum is close to the physiological temperature of the corneal surface (Geiser et al., 2004, Morgan

Acknowledgments

Supported by the Kentucky Lions Eye Foundation, and an unrestricted grant from Research to Prevent Blindness Inc. DD is a Department of Physiology fellow, University of Louisville, Louisville, KY. GWC is a KBRIN fellow, University of Louisville, Louisville, KY.

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