Role of keratan sulphate (sulphated poly -N-acetyllactosamine repeats) in keratoconic cornea, histochemical, and ultrastructural analysis.

AIMS: Keratan sulphate (KS) is the predominant glycosaminoglycan (GAG) present in the corneal stroma where it is thought to regulate collagen fibril diameter. In this study we investigated the distribution of KS in normal and keratoconic corneas. METHODS: Four normal, one mild, and four severe keratoconic corneas were used for the study. Distribution of keratan sulphate proteoglycans (KS-PG) was investigated using a primary monoclonal antibody (5-D-4) that recognizes disulphated disaccharides in the poly-N-acetyllactosamine repeats of KS. The immuno-reactivity of 5-D-4 was analyzed by immunohistochemistry and immuno-electron microscopy. RESULTS: Immuno-histochemistry showed diffuse 5-D-4 staining in keratoconic cornea compared to the punctuate staining in normal corneas. In the single cornea with mild keratoconus, immunogold microscopy revealed a very high density of KS-PG staining, especially in the posterior stroma, compared to severe keratoconic and normal cornea. The amount of KS-PG in the stroma in severe keratoconus was slightly less compared to the normal cornea. In the mild keratoconic cornea, a higher quantity of KS-PG was present around the keratocytes. In severe keratoconic corneas, a higher quantity of KS-PG was present within the keratocytes compared to normal cornea. CONCLUSIONS: The finding of an altered expression of KS in our keratoconic corneas, in particular the strong expression of KS in keratocytes, is in keeping with reports of an altered expression of proteoglycan metabolism in keratoconus. KS-PG plays an important role in stromal collagen fibril assembly and a dysregulation of KS-PG synthesis or catabolism could explain changes in collagen fibril spacing and diameter, which we have reported elsewhere.

Optic nerve sheath diameter, intracranial pressure and acute mountain sickness on Mount Everest: a longitudinal cohort study.

OBJECTIVE: To investigate the association of optic nerve sheath diameter (ONSD), as a correlate of intracranial pressure (ICP), with acute mountain sickness (AMS). DESIGN: Longitudinal cohort study of mountaineers from sea level to 6400 m. SETTING: Mount Everest (North side). PARTICIPANTS: 13 mountaineers (10 men, 3 women; aged 23-52 years) on a British expedition to climb Mount Everest. INTERVENTIONS: ONSD was measured ultrasonically, 3 mm behind the globe using B scans recorded with an OTI-Scan 3D scanner (Ophthalmic Technologies, Canada). Serial binocular scans were recorded at sea level, and 2000, 3700, 5200 and 6400 m. All ONSDs were measured by a blinded observer. MAIN OUTCOME MEASURES: ONSD, AMS score (using the Lake Louise scoring system), heart rate, and oxygen saturation levels. RESULTS: All results were analysed by regression analysis with adjustment. ONSD was positively associated with increasing altitude above sea level (0.10 mm increase in ONSD per 1000 m, 95% CI 0.05 to 0.14 mm) and AMS score (0.12 mm per score, 95% CI 0.06 to 0.18 mm); further associations were found with resting heart rate (0.29 mm per 20 beats/min, 95% CI 0.17 to 0.41 mm) and oxygen saturations (0.20 mm per 10% decrease, 95% CI 0.11 to 0.29 mm). CONCLUSIONS: ONSD increases at high altitude, and this increase is associated with more severe symptoms of AMS. Given the linkage between ONSD and ICP, these results strongly suggest that intracranial pressure plays an important role in the pathophysiology of AMS.

Basal Tear Osmolarity as a metric to estimate body hydration and dry eye severity.

The osmolarities of various bodily fluids, including tears, saliva and urine, have been used as indices of plasma osmolality, a measure of body hydration, while tear osmolarity is used routinely in dry eye diagnosis, the degree of tear hyperosmolarity providing an index of disease severity. Systemic dehydration, due to inadequate water intake or excessive water loss is common in the elderly population, has a high morbidity and may cause loss of life. Its diagnosis is often overlooked and there is a need to develop a simple, bedside test to detect dehydration in this population. We hypothesize that, in the absence of tear evaporation and with continued secretion, mixing and drainage of tears, tear osmolarity falls to a basal level that is closer to that of the plasma than that of a tear sample taken in open eye conditions. We term this value the Basal Tear Osmolarity (BTO) and propose that it may be measured in tear samples immediately after a period of evaporative suppression. This value will be particular to an individual and since plasma osmolarity is controlled within narrow limits, it is predicted that it will be stable and have a small variance. It is proposed that the BTO, measured immediately after a defined period of eye closure, can provide a new metric in the diagnosis of systemic dehydration and a yardstick against which to gauge the severity of dry eye disease.

A hypothesis to suggest that light is a risk factor in glaucoma and the mitochondrial optic neuropathies.

The authors propose that light entering the eye interacts with retinal ganglion cell (RGC) axon mitochondria to generate reactive oxygen intermediates (ROI) and that when these neurons are in an energetically low state, their capacity to remove these damaging molecules is exceeded and their survival is compromised. They suggest that in the initial stages of glaucoma, RGCs exist at a low energy level because of a reduced blood flow at the optic nerve head and that in the mitochondrial optic neuropathies (MONs), this results from a primary, genetic defect in aerobic metabolism. In these states RGCs function at a reduced energy level and incident light on the retina becomes a risk factor. Preliminary laboratory studies support this proposition. Firstly, the authors have shown that light is detrimental to isolated mitochondria in an intensity dependent manner. Secondly, light triggers apoptosis of cultured, transformed RGCs and this effect is exacerbated when the cells are nutritionally deprived. Detailed studies are under way to strengthen the proposed theory. On the basis of this proposal, the authors suggest that patients with optic neuropathies such as glaucoma or at risk of developing a MON may benefit from the use of spectral filters and reducing the intensity of light entering the eye.

Clinical staining of the ocular surface: mechanisms and interpretations.

In this article we review the mechanism of ocular surface staining. Water-soluble dyes are excluded from the normal epithelium by tight junctions, the plasma membranes and the surface glycocalyx. Shed cells can take up dye. A proportion of normal corneas show sparse, scattered time-dependent, punctate fluorescein uptake, which, we hypothesise, is due to a graded loss of the glycocalyx barrier, permitting transcellular entry into pre-shed cells. In pathological staining, there is little evidence of 'micropooling' at sites of shedding and the term 'punctate erosion' may be a misnomer. It is more likely that the initial event involves transcellular dye entry and, in addition, diffusion across defective tight junctions. Different dye-staining characteristics probably reflect differences in molecular size and other physical properties of each dye, coupled with differences in visibility under the conditions of illumination used. This is most relevant to the rapid epithelial spread of fluorescein from sites of punctate staining, compared to the apparent confinement of dyes to staining cells with dyes such as lissamine green and rose bengal. We assume that fluorescein, with its lower molecular weight, spreads initially by a paracellular route and then by transcellular diffusion. Solution-Induced Corneal Staining (SICS), related to the use of certain contact lens care solutions, may have a different basis, involving the non-pathological uptake of cationic preservatives, such as biguanides, into epithelial membranes and secondary binding of the fluorescein anion. It is transient and may not imply corneal toxicity. Understanding the mechanism of staining is relevant to the standardisation of grading, to monitoring disease and to the conduct of clinical trials.

Topical application of serotonin or the 5-HT1-agonist 5-CT intraocular pressure in rabbits.

PURPOSE: The effect of serotonin (5-hydroxytryptamine: 5-HT) and other agonists on rabbit intraocular pressure (IOP), pupil size, and the breakdown of blood-aqueous barrier were evaluated. METHODS: Serotonin and various other agonists were applied topically to the rabbit eye, and intraocular pressure was followed over the next 3 hours using a Digilab 30D pneumatometer. RESULTS: It was demonstrated immunohistochemically that topical 5-HT reached the anterior chamber within 1 hour. Serotonin raised the IOP in a dose-dependent manner over a period of up to 4 hours, with a maximum reached between 30 minutes and 1 hour. A similar effect was observed with the 5-HT1-agonist 5-carboxamidotryptamine (5-CT). Neither tryptamine, 5-hydroxytryptophan, melatonin, gepirone, nor 5,6/5,7-dihydroxytryptamine caused any changes in IOP. Serotonin did not cause a change in pupil size or a breakdown of the blood-aqueous barrier, nor did the aqueous cAMP change significantly after topical 5-HT administration. CONCLUSIONS: The data presented suggest a role for 5-HT in the control of IOP. Previously demonstrated receptors on the iris-ciliary body and the effect of the 5-HT1-agonist 5-CT suggest that the rise in IOP may be caused partly or entirely by an increase in aqueous secretion mediated by 5-HT1-like receptors. Whether or not 5-HT has a role in altering aqueous outflow resistance remains to be seen. An effect of serotonin on other aspects of aqueous dynamics or on the extraocular muscles to cause a change in IOP cannot be excluded.

Tear meniscus changes during cotton thread and Schirmer testing.

PURPOSE: To elucidate the effect of the cotton thread test (CT-T) and Schirmer test (S-T) on the tear reservoir by evaluating the radius of tear meniscus curvature. METHODS: The radii (R) of the central lower tear menisci were measured by a newly developed video meniscometer in 11 eyes of 11 normal volunteers (6 men, 5 women; mean age, 27.7 +/- 3.6 years [SD]) and 9 eyes of 9 patients with tear deficiency and severe dry eye in whom the puncta had been therapeutically occluded (9 women; mean age, 50.6 +/- 10.4 years). In this dry eye group, the absence of reflex tearing, coupled with the absence of lacrimal drainage due to punctal occlusion allowed more precise observation of the removal of tears from the meniscus. A 1-minute CT-T was performed, followed after an interval of 10 minutes by a 1-minute S-T. Tear meniscus curvature was documented before (R:(0)) and during the tests at 30 seconds (R(30)) and 60 seconds (R:(60)). RESULTS: In the normal group, respective R values (CT-T; S-T; mean +/- SD mm) were R(0) (0.26 +/- 0.11; 0.26 +/- 0. 07), R(30) (0.27 +/- 0.16; 0.20 +/- 0.13), and R(60) (0.29 +/- 0.15; 0.23 +/- 0.21); and in the dry eye group, respective R: values (CT-T; S-T) were R(0) (0.59 +/- 0.23; 0.51 +/- 0.19), R(30) (0.52 +/- 0.25; 0.22 +/- 0.09), and R(60) (0.51 +/- 0.19; 0.21 +/- 0.08). It was demonstrated in the dry eye group that R was diminished more by the S-T than by the CT-T in the time course of the measurement (P = 0.01). In the dry eye group alteration of R occurred within the first 30 seconds, and in this group significant correlation was found between R(0) and the S-T result (r = 0.67; P = 0.05), and between R(60)- R(0) and the S-T result (r = -0.81; P = 0.01). Also, there was a significant correlation between R(60)- R(0) and the S-T result in the normal group (r = 0.71; P = 0.02). There were no significant correlations between R(0) or R(60)- R(0) and the CT-T results in either group. CONCLUSIONS: These studies afford some insight into the dynamics of the Schirmer test, suggesting that wetting is influenced by the negative hydrostatic pressure within the tear meniscus. With the protocol used, no conclusion could be drawn about the relation between meniscus radius and wetting of the cotton thread.

Morphologic characteristics and chemical composition of Christmas tree cataract.

PURPOSE: Christmas tree cataract consists of highly refractile multicolored "needles" crisscrossing the lens fibers of the deep cortex. The fact that the colors vary according to the angle of the incident light, and that in retroillumination only a dim outline of the cataract is seen, would suggest that Christmas tree cataract is a diffractive phenomenon. This study was performed to unravel the ultrastructure and chemical composition of the Christmas tree needles. METHODS: Eight lenses from donor eyes and four extracapsularly extracted lenses with Christmas tree cataract were investigated by scanning and transmission electron microscopy. The chemical composition was studied with energy-dispersive x-ray microanalysis and Raman microspectroscopy. RESULTS: Scanning electron microscope examination showed that the needles are smooth, rectangular, plate-like elements bordered by membranes and amorphous material and running crisscross through the lens. In the specimens for transmission electron microscopic examination, the needles proved to be largely dissolved, but the remains showed regular spacings of approximately 5 nm. Material identical in spacing and electron density was found in neighboring cells bound to a reticular membranous network originating from the fiber-limiting membranes. Energy-dispersive x-ray and Raman microanalysis showed that the needles have a high sulfur content and pronounced S-S, CS-SC, and C-S vibrations. The cytoplasm adjacent to the needles and reticular meshwork had an elevated Ca++ content. CONCLUSIONS: It is concluded that cystine is the most likely candidate for the Christmas tree needles and that the needles probably are formed as the result of an age-related aberrant breakdown of crystallins induced by elevated Ca++ levels.

Heterogeneity in ultrastructure and elemental composition of perinuclear lens retrodots.

PURPOSE: To unravel the cataractogenic process(es) leading to the birefringent lenticular bodies known as perinuclear retrodots. METHODS: Ten human lenses containing biomicroscopically verified perinuclear retrodots were systematically screened and analyzed using scanning electron microscopy and energy dispersive x-ray microanalysis to verify their ultrastructure and elemental composition. RESULTS: Three types of retrodots were distinguished, different in size, ultrastructure, and origin. Two of them contained calcium phosphate, the third probably contained calcium oxalate. All three types were separated from surrounding normal fibers and the crystalline inclusions were sequestered within membrane-lined bodies. CONCLUSIONS: Because of these observations and data found in the literature it is postulated that elevated free calcium is the initiating factor in the formation of retrodots, trapped by either oxalate or phosphate and sequestered in the retrodots. It is suggested that the oxalate is derived from ascorbate because of impaired protection against oxidative stress in the older lens. Phosphoric acid is believed to be released by calcium-induced hydrolysis of membrane phospholipids.

Assessment of meibomian gland function in dry eye using meibometry.

OBJECTIVE: To study meibomian gland function in dry eyes using meibometry. METHODS: Forty-two patients with clinically diagnosed dry eye that was reclassified as meibomian gland dysfunction (MGD [n = 12]), aqueous-tear deficiency (AD [n = 10]), MGD combined with AD (n = 2), "incomplete" dry eye (n = 12), and non-dry eye (6 eyes) were compared with 41 healthy control subjects. The following 2 techniques of meibometry were applied: direct meibometry (DM) measuring lipid imprints using the Meibometer, and integrated meibometry (IM) using image-scanning and computer densitometry. Tear film lipid layer thickness was assessed using interference microscopy. RESULTS: Imprints were homogeneous for all subjects except those with MGD. Mean+/-SE readings on results of DM were 127.24+/-24.4 for MGD, 306.4+/-9.2 for AD, 248.6+/-13.2 for incomplete dry eye, and 268.5+/-6.3 for controls, showing lower values in the MGD group relative to all others (P<.001). Results of IM gave similar results (P<.001, P =.01, and P<.001, respectively). Lipid layers appeared lower for the MGD group than others. CONCLUSIONS: Compared with controls, lid lipid levels are reduced in patients with MGD, and increased in women with AD. Lipid layer thickness is increased in women with AD compared with patients with MGD. Both meibometric techniques may be useful for evaluating MGD. Although DM requires special equipment (the Meibometer), it provides a record of immediate diagnostic value. Although IM is less effective than DM, it offers visual documentation of the lipid imprint, which may itself be of diagnostic value, and uses equipment available in many laboratories.

Deposits and proteoglycan changes in primary and recurrent granular dystrophy of the cornea.

OBJECTIVE: To investigate the origin and distribution of granular deposits in the corneas of 3 patients with granular dystrophy, 1 of whom had previously received a lamellar keratoplasty in which the granular dystrophy had recurred. METHOD: Corneal tissue from 2 patients with primary granular dystrophy (patients 1 and 2) and from a patient with recurrent granular dystrophy (patient 3) was examined. Corneal graft tissue was fixed in (1) 3% glutaraldehyde in sodium cacodylate buffer, (2) 2.5.% glutaraldehyde in sodium acetate buffer containing cuprolinic blue, and (3) 4% paraformaldehyde in phosphate-buffered saline. RESULTS: In patient 1 (aged 48 years), electron-dense granular structures were observed in epithelium, Bowman layer, and throughout the stroma. Bowman layer was absent in several places. Patient 2 (aged 78 years) showed similar features except with more deposits in the stroma. In patient 3 (aged 48 years), granular structures were heavily deposited in the epithelium; there were also some deposits in the posterior (host) stroma, some of which were associated with partially degenerated keratocytes. Bowman layer appeared normal. In all 3 patients, the intracellular or extracellular granular structures were surrounded by fine fibrillar material and abnormal proteoglycans. Electron-lucent spaces within the corneal stroma contained large quantities of abnormal proteoglycan filaments that were attached in part to collagen fibrils. CONCLUSIONS: Results from patient 3 support an epithelial origin for the deposits, presumably from keratoepithelin, aggregated with other proteins. The role of keratocytes is less clear, although the presence of deposits in the stroma of all 3 patients, some associated with keratocytes, suggests that these cells might produce granular material in addition to abnormal proteoglycans.

Diagnosis of dry eye.

Dry eve disease is characterized by symptoms, ocular surface damage, reduced tear film stability, and tear hyperosmolarity. There are also inflammatory components. These features can be identified by various kinds of diagnostic tests (symptom questionnaires, ocular surface staining, tear break-up time, and osmometry), although there may not be a direct correlation between the number or severity of symptoms and the degree of ocular surface damage or tear deficiency. Once the diagnosis of dry eye disease has been established, further tests can be used to classify the condition into tear-deficient or evaporative dry eve. The two forms of dry eye are not mutually exclusive and often co-exist. The optimal diagnosis of dry eye disease, therefore, depends on the results of several tests, and this article suggests an appropriate order for performing these tests at a single clinic visit.

Superficial lines and associated disorders of the cornea.

In a clinical classification of fine superficial corneal lines, we recognized five major types of line: mare's tail lines, fibrillary lines, fingerprint lines, lacunar dystrophy or map-like changes, and tram lines. The fingerprint lines were further classified into several subgroups. There was a relationship between certain lines, in particular, fingerprint lines and lacunar dystrophy or map-like change with the occurrence of Cogan's microcystic dystrophy and recurrent erosion.

Ocular infection with orf virus.

Orf virus infection of the pericanthal eyelid skin and conjunctiva was diagnosed by electron microscopy. Orf is a paravaccinia virus responsible for contagious pustular dermatitis in sheep and goats. It occasionally affects humans. The patient was a farmwife who had recently handled lambs. She had a raised ulcerating lesion at the inner canthus, follicular conjunctivitis, and lymphadenopathy. Treatment with 35% idoxuridine in dimethylsulfoxide on the skin and 0.5% idoxuridine ointment on the conjunctiva led to resolution after three weeks.

Epithelial deposition of gold in the cornea in patients receiving systemic therapy.

Epithelial gold deposits were demonstrated biomicroscopically in 13 of 15 patients receiving gold therapy for rheumatoid arthritis. Only one of six further patients who had received gold therapy in the past showed such deposits. Deposits appeared with a cumulative dose of sodium thiomalate exceeding 100 mg. The earliest appearance was after seven months of therapy and deposits were visible in one subject as long as nine months after therapy had stopped. The density of deposit does not appear to increase over the whole of the dose range, possibly as a result of the effect of epithelial turnover. No symptoms attributable to the deposit were encountered in any patient.

Clinical and biochemical heterogeneity in gyrate atrophy.

Two patients had milder variants of the usual form of gyrate atrophy. Although the appearance of the chorioretinal degeneration was identical, the progression of the clinical signs and symptoms were slower with night blindness, cataracts, and decreased vision occurring at an older age. Additionally, the level of hyperornithinemia, although still increased above normal, was lower than that usually found in such patients (between 448 and 676 microM). Additionally, one of our patients responded to high dosage vitamin B6 with a 27% reduction in plasma ornithine. The difference in the quantitation of the increase in plasma ornithine, the difference in responsiveness to vitamin B6, and the difference in the clinical consequences of this underlying biochemical abnormality, indicated a variant form of gyrate atrophy, suggesting the presence of genetic heterogeneity.

Cavernous-dural fistula with secondary angle-closure glaucoma.

PURPOSE: To report a rare case of angle-closure glaucoma, secondary to the rapid development of a choroidal effusion, in a patient with a long-standing cavernous-dural shunt. METHODS: Case report. Investigations included computed tomographic scan, magnetic resonance imaging, and carotid angiography. RESULTS: The development of the choroidal effusion occurred because of partial thrombosis of the ipsilateral superior ophthalmic vein and cavernous sinus. Drainage of the choroidal effusion resolved the angle-closure glaucoma. CONCLUSIONS: The combination of worsening signs and evidence of thrombosis indicates impending resolution of a cavernous-dural shunt. However, if a choroidal effusion causes angle-closure glaucoma, prompt surgical drainage should be considered to prevent permanent peripheral anterior synechiae formation, with the expectation that the effusion will not recur.

Lacrimal streams: the demonstration of human lacrimal fluid secretion and the lacrimal ductules.

The clinical demonstration of the ductular orifices of the human lacrimal gland is reported. Lacrimal fluid secretion can be shown after instillation of 2% sodium fluorescein. Ductular orifices are visible on biomicroscopy. In keratoconjunctivitis sicca the lacrimal fluid streams appear normal, or to be diminished or absent.

Corneal toxicity from vinblastine solution.

The case is reported of a male physician who accidentally sprayed some vinblastine solution into his eyes. The resulting lesions are described.

West Indian punctate keratopathy in a Caucasian.

We present a patient with the picture of corneal white spots, very similar in morphology to those described in native West Indians, occurring here in a Caucasian. Their aetiology remains obscure.