Download elucidate the contribution of proteins to tears. a challenge for

EDITORIAL
ARCH SOC ESP OFTALMOL 2006; 81: 187-190
ELUCIDATE THE CONTRIBUTION OF PROTEINS TO
TEARS. A CHALLENGE FOR RESEARCHERS
ELUCIDAR LA CONTRIBUCIÓN DE LAS PROTEÍNAS EN LAS
LÁGRIMAS. UN RETO PARA LOS INVESTIGADORES
MILLAR TJ1
Beneath the dominant, and almost dismissive
idea that the roles for proteins in the tears are antimicrobial and immunological, has been a persistent
undercurrent from stalwarts investigating the physical contribution of proteins to tears. It is here that
we enter the realms of surface tension and viscoelasticity and the many publications of Frank Holly,
Anthony Bron and John Tiffany. A low surface tension and a high non-Newtonian viscosity are essential for tear film stability and proper lubrication of
the ocular surface, and it appears that proteins are
imperative for these properties.
Of the many hundreds of proteins now known to
be present in human tears (1), tear lipocalin
(~18kD) has attracted special attention for its possible role in affecting surface tension and viscoelasticity. This is because of its relative abundance (1533% total tear protein), its relatively poor antimicrobial activity, and its ability to bind lipids and
hence interact with the lipid layer at the outer surface of the tear film. By comparing changes in surface tension of lipid depleted tears, and mixtures of
various tear proteins, Nagyova and Tiffany implied
that tear lipocalin was interacting with the surface
lipids to lower the surface tension (2). This idea that
proteins are interacting with the lipid layer has been
taken a step further by our group. We now believe
that proteins are an integral part of the air liquid
interface of the tear film and not simply interacting
with the lipids from below. The evidence in part has
come from comparing dynamic surface tension
changes of Meibomian oils, phospholipids, ocular
mucin, whole tears, and tear proteins in a Langmuir
trough. From this, it is clear that proteins, and not
1
Assoc. Prof. School of Natural Sciences. University of Western Sydney.
E-mail: [email protected]
Meibomian lipids alone, are needed for lowering
the surface tension to that found in whole tears.
Moreover, these proteins include the major tear proteins such as lysozyme and mucins (3,4). Enigmatically lipocalin is giving confounding results (unpublished) which are indicating that apolipocalin is
able to penetrate a lipid film, but hololipocalin (lipocalin loaded with lipids) appears to be trapped in the
aqueous. By contrast, other tear proteins unfold at
the surface and become trapped in the lipid layer. By
labelling the proteins and lipids with fluorescent
tags, we have been able to show that the presence of
proteins causes the surface layer to become gel like
rather than mobile which is the case when lipids alone form the surface film. This is comparable to clinical findings, where the surface pattern of the tear
film is stable through multiple blinks.
In terms of tear viscosity, a recent exciting, but
confounding paper by Gouveia and Tiffany (5),
indicates that pure solutions of lysozyme, lactoferrin, IgA, or tears without lipids behave as Newtonian fluids i.e. as an object is moved faster through
a fluid (shear rate) the resistance it incurs (viscosity) does not change. Whole tears (with lipids)
behave quite differently: their viscosity drops
rapidly with increased shear rate. This is known as
shear thinning or non-Newtonian behaviour, and is
thought to be important for reducing drag on the
ocular surface when blinking. In contrast with the
pure protein solutions, mixtures of pairs of the proteins above or apolipocalin alone showed shear
thinning similar to normal tears. Hololipocalin
(with lipids) showed Newtonian behaviour. At this
stage, there are no explanations for these confusing
phenomena other than it being the mixture of proteins with proteins, and proteins other than lipocalin with lipids which give tears their non-Newtonian
properties. Important here is the concept of protein
interaction in the tears rather than the role of individual proteins. This interaction of proteins has not
gone unnoticed by Ben Glasgow’s team who have
described that lipocalin interacts with both lactoferrin and lysozyme, but not albumin and consequently has suggested that there is a «functional
interdependence of these proteins in the formation
of the tear film» (6). Importantly this interaction is
ionic and is disrupted at high salt concentrations.
Speculatively, this could help explain the common
thread of increased osmolarity in dry-eye: increased
salt concentrations disrupts the protein-protein interactions leading to altered viscoelasticity and surfactant properties of the proteins in tears which in
turn leads to rapid tear breakdown.
These studies on the contribution that proteins
have to the physical nature of tears hold many exciting possibilities. Do the inflammatory proteins disrupt the physical performance of the common tear
proteins? What are the contributions of some of the
quantitatively small proteins to the physical properties of the tears, or similarly proteins that are relatively abundant, such as lacritin, but have as yet no
188
clearly defined role? I look forward to seeing the
results from intrepid young workers who have the
courage to take up the challenge of examining the
tear film as a dynamic interaction of molecules.
REFERENCES
1. Sack RA, Conradi L, Krumbolz D, Beaton A, Satbe S,
Morris C. Membrane array characterization of 80 chemokines, cytokines and growth factors in open- and closedeye tears: angiogenin and other defense system constituents. Invest Ophthalmol Vis Sci 2005; 46: 1228-1238.
2. Nagyova B, Tiffany JM. Components responsible for the
surface tension of human tears. Curr Eye Res 1999; 19: 411.
3. Tragoulias ST, Anderton PJ, Dennis GR, Miano F, Millar
TJ. Surface pressure measurements of human tears and
individual tear film components indicate that proteins are
major contributors to the surface pressure. Cornea 2005;
24: 189-200.
4. Millar TJ, Tragoulias ST, Anderton PJ, Ball MS, Miano F,
Dennis GR, et al. The surface activity of purified ocular
mucin at the air-liquid interface and interactions with meibomian lipids. Cornea 2006; 25: 91-100.
5. Gouveia SM, Tiffany JM. Human tear viscosity: an interactive role for proteins and lipids. Biochim Biophys Acta
2005; 1753: 155-163.
6. Gasymov OK, Abduragimov RA, Yusifov TN, Glasgow BJ.
Interaction of tear lipocalin with lysozyme and lactoferrin.
Biochem Biophys Res Commun 1999; 265: 322-325.
ARCH SOC ESP OFTALMOL 2006; 81: 187-190