The use of stains in dry eye assessment (C6178)

The use of stains allows us to view these changes directly and immediately. It is, therefore, extremely important to be able to assess this damage if the diagnosis is to be made and any treatment evaluated for effectiveness. Although several techniques have evolved to assess the extent of this damage, staining the ocular surface with dyes such as fluorescein, rose bengal and lissamine green is the most practical clinically.

Fluorescein staining seems to offer the best clinical means of diagnosing the presence of corneal epithelial surface defects, particularly those of less severity than would exhibit staining with rose bengal. There is some controversy as to whether the dye is staining compromised or damaged cells, or areas where cells are missing or intracellular junctions are weak.2,3 Generally, fluorescein is fairly painless for the patient except in cases of more severe corneal damage. Although often stated to the contrary, fluorescein does stain mucus (as seen in the early case of filamentary keratitis in Figure 1).

Ever since its use by Sjögren in 1933, rose bengal has been widely used in the diagnosis of dry eye.4 Sjögren proposed that rose bengal stains dead or degenerate epithelial cells and mucus. However, later work by Feenstra and Tseng5 showed that rose bengal stains healthy cells that are not protected by an intact mucus layer. It would seem that the presence of rose bengal staining indicates a deficiency of pre-ocular tear film protection. It has yet to be shown whether these cells are healthy, dead or damaged.

The conjunctiva generally shows more staining with rose bengal than the cornea. Rose bengal staining is very uncomfortable for the patient, the degree of discomfort being proportional to the amount of agent used and the amount of damaged ocular surface cells present. As a result many clinicians use a fraction of a drop of 1 per cent rose bengal to minimise discomfort.

Lissamine green

Lissamine green is a relatively new ocular stain and in many ways is preferable to rose bengal as, in all but the most severe cases, there is little ocular discomfort.

According to Tseng,5 it does not stain healthy cells, but only dead and degenerate ones. Nor does it stain cells that are merely unprotected by an intact mucous layer, as in the case with rose bengal staining. This may be an over-simplification, as lissamine green staining can sometimes be seen where a tight fitting soft lens has been exerting pressure in the limbus, in the absence of any other signs of damage (Figure 2).

All three stains may still have a diagnostic importance. Whether the term 'vital staining' - the staining of devitalised cells - can be used completely accurately for any of the three agents is debatable.

The fluorescence of fluorescein is considerably enhanced if, in addition to the cobalt blue excitation filter with the slit-lamp illumination on maximum intensity, a yellow barrier filter is used (Wratten no12, see below) in the viewing system. This blocks the blue light necessary to cause the fluorescein to fluoresce from the image so that the green staining is more easily visualised.

Although fluorescein staining is usually associated with corneal desiccation, it may occur with other causes, such as ocular rosacea, insult from staphylococcal exotoxins, preservative toxicity (Figure 3), infections and allergies, among other causes. It is important to consider these before arriving at a diagnosis of dry eye.

According to Korb,1 the optimal technique for viewing fluorescein staining is to instil the drop, blink three times, and then to wait one to two minutes. This gives the dye enough time to penetrate the damaged epithelial cells and leach out of the tear film, so as not to mask the fluorescence of the epithelial cells with the fluorescence of the tear film itself.

Korb found that if a second drop of Fluorescein were instilled at this point, a 20 per cent increase in corneal staining was observed, often showing damage that would not have been observed if a single instillation had been used. Again the significance of this is open to debate. Sequential staining may merely show an increased cytotoxicity to fluorescein with repeat administration. In fact, a remarkably high percentage of tear normal, non-contact lens wearers show significant staining on sequential instillation.7

Lissamine green staining fades relatively quickly so should be assessed one eye at a time to prevent it being under-reported in the eye observed second. When lissamine green is used it is important to instill a volume (10-20µl) that will allow adequate staining. At least a minute and no more than four minutes shows optimum staining.8 It is important not to view with too bright a light source or the staining is bleached out. The staining is enhanced if a red filter (Wratten no 25) is used as a barrier device on the slit lamp (Figures 4 and 5).

In these photos taken after wearing a silicone hydrogel bandage contact lens, the lissamine green staining of the exposed bulbar conjunctiva can be more clearly seen when the barrier filter is used. In the case of lissamine green, the filter is blocking the green light from the stained area, making it appear darker and more visible, whereas the yellow filter used with fluorescein is blocking the illuminating blue light, enhancing the visability of the green colour in the stained areas.

These photos show how protective a silicone hydrogel contact lens can be in dry eye cases. The early silicone hydrogel lenses were relatively stiff and in some cases were poorly tolerated by patients due to discomfort. Newer products are less so and are often better accepted.

Selection of solution systems are very important in these cases as these patients do not have the tear volume to dilute the cytoxic effects of some preservatives.

Generally, in these therapeutic cases it is recommended to use a system without a preservative, or a self-disintegrating preservative such as the new Penasept low dose peroxide system, Regard or Synergy. Bandage silicone hydrogel lenses are particularly useful where there is a recurrent erosion syndrome or epithelial basement dystrophy element to the dry eye condition.

As far as digital photography is concerned a better image can be achieved by only looking at the red signal. With the ARC Optical setup used by this author it is possible to view the red signal from the camera in monochrome, producing a high quality high contrast image where the lissamine green staining appears black (Figure 6). Where such systems are not available, or where direct viewing is required, the red barrier filter system is the best answer.

Although lissamine green staining of the cornea only happens in the more severe cases of dry eye, when it does, there often appears less fluorescein staining than expected afterwards. Whether this is because they are staining different things, or the prior absorption of lissamine green reduces subsequent fluorescein uptake, will only show with further experience with these products.

The important thing with sequential comparisons is that the evaluation is always performed in the same manner, whether this is fluorescein first then lissamine green second, or visa versa, or as some US collegues do, a drop of each simultaneously. Only then can valid serial comparisons be made from each visit.

Although the assessment of corneal staining with fluorescein is the conventional means of detecting and grading compromised corneal cells, it is always difficult to standardise the judgement of the exact amount of staining present, and whether or not progression or improvement has occurred. While digital imaging has helped with sequential comparisons, these are only valid if such things as the camera sensitivity settings, slit lamp illumination settings, use of barrier filters, absolute concentration of dye in tear film and so on remain constant between patients and repeat visits.

If digital imaging is not available then grading each area of the cornea, conjunctiva and lids is the next best alternative.

Dry eye management

It may be asked, what is the significance of staining of the ocular surface?

Firstly, because the cornea is so heavily innervated, reduction in corneal staining will improve comfort levels for the patient. The longer the cornea stays healed, the stronger it becomes.

Secondly, personal experience shows that as conjunctival staining reduces, so tear quality improves, presumably due to increased goblet cell density.

A common feature of both aqueous deficient and evaporative dry eye is increased tear osmolarity. As a result, the tears 'pull' water out of the surface of the eye by osmosis, causing the dry eye symptoms that usually worsen as the day goes on. The conjunctiva is the first to suffer, with a reduction in the amount of mucus secreting goblet cells. An intact mucous film is required to hold the aqueous component of the tears in place.

Osmolarity is the concentration of dissolved particles in the tears, without regard to their size, density, electric charge or configuration. It decreases overnight due to decreased tear evaporation, and conversely increases in dry eye.9 An osmolarity of less than 312mOsm/kg is physiological, rising to 323mOsm/kg in moderate to severe dry eye.10

Lacrimal gland fluid osmolarity increases as secretory rate declines, independent of the effect of evaporation.11 With a defective lipid layer, tears evaporate faster than replaced, hence tonicity increases (Figure 8).

An electrolyte balanced, preservative-free, hypotonic tear supplement will help in all cases of dry eye syndrome, as tear osmolarity increases independently both in reduced secretion and evaporative dry eye.

Some artificial tear supplements, such as Theratears, have a low osmolarity, in an attempt to reverse that found in dry eye. If the lacrimal drainage mechanism is occluded, the effect is longer lived.12 The hypotonicity of Theratears drops has been chosen to restore the tear tonicity to a level that encourages goblet cell repopulation of the conjunctiva. This effect is greater if the patient has punctal plugs fitted as the optimised tonicity is maintained for much longer.

The aims of dry eye treatment can thus be broken down into reducing corneal and conjunctival damage (in other words, reduce staining), and improving patient comfort. This is usually done by the following means:

• Improve tear volume by aqueous supplements
• Improve quality of mucous layer - Theratears/Ilube, gel supplements
• Improve lipid layer/reduce tear evaporation - diet/omega-3 nutritional supplements, lid hygiene, hot compresses, drugs (tetracyclines), paraffin ointment and Clarymist Liposomal spray (for the polar phospholipid layer)
• Therapeutic contact lenses - reduce tear evaporation , encourage healing
• Reduce tear drainage and hence increase tear volume - punctal plugs, cautery
• Reduce inflammation - omega-3, steroids, NSAIDs, cyclosporin, anti-allergy products.

Ocular lubricants/artificial tears take many forms. The main active ingredients usually being carboxymethylcellulose (CMC), polyvinyl alcohol, hyaluronic acid, polyethylene glycol and polyvinyl-pyrrolidone. They come in preserved multi-dose and preservative-free forms. Preservative-free products are preferred as they do not suffer from the cytotoxic and allergy-causing problems suffered by their multi-dose counterparts.

The preservative-free hypotonic artificial tears this author uses are a brand called Theratears developed by Gilbard at the Schepens Eye Institute. These are unit dose, preservative-free, hypotonic, with the active ingredient CMC (0.25 per cent). CMC is negatively charged and binds to the corneal epithelial surface well. It also has a cyto-protective function against the insult from the preservatives present in contact lens disinfecting solutions.13,14 This is an important fact when bandage contact lenses are fitted to a dry eye patient to promote healing and reduce evaporation, where the cytoxic effects can be more marked due to the compromised cornea and the inability of the reduced tear volume to dilute the preservatives in the contact lens solution.

A hypotonic solution may have beneficial effects on conjunctival goblet cell populations, especially in patients fitted with punctal plugs, due to its hypotonicity. It also is a very 'slippery' product which may help corneal healing by reducing the shear effects from the upper eyelid on blinking.

Punctal plugs and cautery

If after trying to normalise the lipid, aqueous and mucous secretions as much as possible, there is still an obvious low tear volume or excessive tear drainage problem, or if the patient just wants to put their eye-drops in less often, then punctal occlusion, either with silicone plugs or cautery, may be indicated.

All plugs that are fitted deeper than the punctal opening are removed if necessary by syringing the plug through the lacrimal apparatus. It is, therefore, imperative that the canaliculii are irrigated prior to fitting to ensure this exit route is patent. If not then there would be no point in fitting the plug in any case.

This author prefers to fit conventional silicone plugs wherever possible. They are trickier to fit, requiring the punctum to be measured first for size and elasticity. Because the right size of plug is fitted tightly into the punctum, tear flow is stopped. A plug that is initially fitted tightly will function for longer. Also, the plug can be removed simply, if required.

Opinions will differ between professionals on the choice of ideal implant, and no one type is suitable for everyone. When silicone plugs will not stay in place the optometrist may refer back to the original referring ophthalmologist for punctal cautery (Figures 9 and 10).

Cautery should be used with caution as it is irreversible and carries significantly greater risks to the patient.

Lid hygiene

It has been shown that a regimen of warm compresses and lid hygiene can make a significant improvement in tear film stability.

A new product called Sterilid has been released recently which is the first antibacterial lid hygiene product. This is a foam formulation that is massaged onto the lids and lashes then rinsed off one minute later. It is a great help in those dry eye cases that have an infective lid margin element, such as in blepharitis and meibomitis.

We might also prescribe a course of fucidic acid gel antibiotic, which has both excellent anti-staphylococcal properties, and also has some anti-inflammatory capability.

The role of omega-6 and omega-3 fatty acids

If we had a predominantly inflammatory element to the problem then flaxseed/fish-oil supplementation would be a good idea, possibly adding a preservative-free NSAID eye-drop such as diclofenac, or even getting the patient's GP to prescribe a course of a low risk/low ocular penetration steroid such as fluoromethalone (FML).

A reduction in cholesterol intake to improve the quality of meibomian secretions in lipid disorders has also been suggested as a useful management option.15

This may also be a mechanism by which the omega-3 supplements help the meibomian secretions.

Unfortunately, long-term supplementation with omega-3 depletes serum levels of vitamin E, so the addition of vitamin E to nutritional supplements helps prevent this.

Evaporation reduction

Dry eye sufferers should optimise their environment to reduce tear film evaporation. They need to modify the local environment around the eye by, for example:

• Avoiding drafts
• Avoiding excessive air conditioning, central heating, airplane travel
• Using room humidifiers.

In extreme cases, the use of spectacles with tight fitting side shields or wearing swimming goggles can be beneficial to maintain high humidity levels around the eyes.15

Bandage silicone hydrogel CLs

Bandage soft silicone hydrogel contact lenses can been used to enhance corneal healing, prevent corneal dehydration and improve comfort.

In cases of recurrent erosions, the patient may wear the contact lens on an extended wear basis, to reduce overnight adhesions forming between the cornea and the tarsal conjunctiva. Evaporative dry eye is most commonly caused by meibomian gland anomalies, which can be blepharitis associated or obstructive meibomian gland dysfunction/disease.

People with large eyes are also subject to high tear evaporation (for example, patients with thyroid eye disease). This can be due to proptosis and/or lid retraction. It can be also caused by poor eyelid/eyeball alignment (for example, floppy eyelid syndrome, lagophthalmos) and other causes such as environment contact lens wear, drugs (such as BKC toxicity in eye drops or topical beta-blockers causing epithelial cell metaplasia and loss of goblet cells16, and allergy6). Activities such as driving, watching TV, using a VDU, or reading, all can increase tear evaporation as well, due to the reduced blinking that often occurs with concentration on such visual tasks. ?

The Ocular Solutions Fluorescein - Lissamine Green Barrier Filter is available at no charge to interested clinicians. Details can be found on the Ocular Solutions website: www.ocularsolutions.com Email: amatheson@ocularsolutions.com

References

1. Korb DR. The Tear Film, Its role today and in the future, p126-190, BCLA pubs Butterworth-Heinemann 2002. 
2. Feenstra R and Tseng S. Comparison of fluorescein and rose bengal staining, Ophthalmology, 1992, 99(4), 605-617.
3. Wilson et al. Corneal epithelial Fluorescein staining. J Americ Optom Assoc, 1995, 66/7, 435-441.
4. Norn M. Micropunctate vital staining of the cornea, Acta Oph, 1970 48, 108-118.
5. Feenstra R and Tseng S. What is actually stained by rose bengal, Ophthalmology, 1992, 110, 984-993.
6. Bron A, Diagnosis of dry eye. Surv Ophthalmol 45,Suppl 2, S221-S226, 2001.
7. Josephson J and Caffery B. Corneal staining characteristics after sequential instillations of fluorescein, Optom Vis Sci, 1992 69, 570-573.
8. Foulks GN and Bron AJ. Meibomian gland dysfunction: A clinical scheme for description, diagnosis, classification and grading. The Ocular Surface, 1(2003) 107-126.
9. Gilbard J. 2004, Dry Eye - Natural history, diagnosis & treatment, in Press.
10. Craig JP et al, Tear Lipid layer structure and stability following expression of the meibomian glands, Oph Phys Opt, 1995 569-574.
11. Gilbard JP, Dartt DA. Changes in rabbit lacrimal gland fluid osmolarity with flow rate, Invest Ophthalmol Vis Sci, 1982 Dec 23(6):804-6 (ISSN: 0146-0404).
12. Gilbard J and Keynon K. Tear diluents in the treatment of keratitis sicca, Ophthalmology, 1985, 92, 613-626.
13. Sulley A. 2004, Contact lens management of the marginal dry eye, in print.
14. Vehinge J, Simmons P et al. Cytoprotective properties of CMC when used prior to wearing contact lenses treated with cationic disinfecting agents, Eye and contact lens, 2003 29(3), 177-180.
15. Paugh J, Vehige J et al, A pre-application drop of CMC can reduce MPS-induced corneal staining, Optom Vis Sci 2007, vol 84, 65-71.
16. Shah S and Laiquzzaman M. Optician, June 6, 2003, p36-41.

? Andrew Matheson is a therapeutic optometrist practicing in Alresford, Hampshire. He is a director of Ocular Solutions, supplying Dry Eye and Clinical products to the profession