Wydanie 3/2011
str. 64

Laserowa skaningowa mikroskopia konfokalna i spektralna optyczna koherentna tomografia – przyżyciowe zastosowanie i obrazowanie zmian morfologii rogówki w dystrofii rogówki Schnydera

In Vivo Laser Scanning Confocal Microscopy and Spectral Optical Coherence Tomography – Application and Morphological Changes in Schnyder Corneal Dystrophy

Dyrda Agnieszka A.1,2, Janiszewska Dominika2, Wylęgała Edward2

1 Department of Ophthalmology, Hospital de la Esperanza, Parc de Salut Mar, Barcelona
  Head: Professor Miguel Castilla Cespedes, MD, PhD
2 Oddział Okulistyczny Okręgowego Szpitala Kolejowego w Katowicach
  Head: Professor Edward Wylęgała, MD, PhD


Summary: Purpuse: Description of the clinical use of laser scanning confocal microscopy and spectral optical coherence tomography in the case of a patient with Schnyder corneal dystrophy.
Design: Case report.
Material and Methods: 68-year-old patient was examined ophthalmologically before and after penetrating keratoplasty of the left eye. The examination included visual acuity, slit-lamp biomicroscopy, B-scan ultrasonography, in in vivo laser scanning confocal microscopy (HRT3/Rostock Cornea Module; Heidelberg Engineering GmbH, Dossenheim, Germany) and spectral optical coherence tomography (SOCT Copernicus Plus: Optopol, Zawiercie, Poland).
Results: Using confocal microscopy and optical tomography allowed the confirmation of the diagnosis of Schnyder corneal dystrophy before obtaining material for histopathological examination. The studies mentioned before had highlighted the characteristic image of the disease and localization of lipid deposition. Application of these modern methods of imaging the cornea also facilitated the choice of surgical method (penetrating keratoplasty), and helped us to understand the pathophysiology of this rare disease of the cornea.
Conclusions: The confocal laser microscopy and optical coherence tomography, new, non-invasive technologies, proved to be useful tools in the diagnosis and conduct of patients with Schnyder corneal dystrophy.

Słowa kluczowe: dystrofia rogówki Schnydera, dystrofie rogówki, laserowa skaningowa mikroskopia konfokalna, spektralna optyczna koherentna tomografia, przyżyciowe zastosowanie.

Keywords: Schnyder corneal dystrophy, corneal dystrophies, laser scanning confocal microscophy, spectral optic coherence tomography, in vivo use.


Schnyder corneal dystrophy (SCD) previously called Schnyder crystalline corneal dystrophy (SCCD) is a rare autosomal, dominantly inherited bilateral disease affecting both sexes with equal probability. The causative gene is localized on chromosome 1p36 (1) and called UBIAD1 (UbiA prenyltransferase domain-containing protein1) (2,3). This gene is involved in cholesterol metabolism. This disorder is characterized by abnormal deposition of cholesterol, phospholipids and recently suggested sphingomyelin mainly in the cornea(4-6), however systemic association such as genu valgum (4%) and hypercholesterolemia (66%) is also observed (7).
Tipically the bilateral ring-shaped or disciform central opacities are found in clinical exam, which usually consist of fine, polychromatic, needle-shaped or rectangular crystals, but sometimes disciform opacity can be present without any evidence. In fact, 46% of SCD eyes have no subepithelial crystals (3), which may delay a diagnosis until the fourth decade, when normally recognised in the first, second decade of life (5). Consequently, Weiss (8) and then the International Committee for the Classification of Corneal Dystrophies (9) removed “crystalline” from the nomenclature.
The diagnosis of SCD is usually based on clinical findings. Histochemical and electron microscopy studies are utilized in a diagnosis confirmation, but the corneal tissue is available only after surgical intervention.
In vivo corneal confocal microscopy allows to perform noninvasive real-time spatial sectioning of living corneal tissues at the cellular level (10,11). The clinical usefulness of this method has been documented in studies of both normal human corneas (12) and corneas affected by dystrophy (13,14), including SCD (15). In vivo laser scanning confocal microscopy (LSCM) (HRT3/Rostock Cornea Module; Heidelberg Engineering GmbH, Dossenheim, Germany) permits more detailed in vivo layer-by-layer observations of corneal microstructure with an axial resolution of nearly 4 micrometers, much better than that obtained with conventional white-light confocal microscopes [for instance, 10 micrometers axial optical resolution with ConfoScan 2 (Nidek Technologies, Vigonza, Italy)] and incomparably better than that with biomicroscopy. The possibility of in vivo evaluation of corneal morfology gives us also spectral optical coherence tomography (SOCT).
The purpose of this case report is to present the results of noninvasive real-time studies: confocal microscopy and spectral optical coherence tomography and their utilization in handling SCD case.

Case
A 68-year-old man was referred to our clinic with the diagnosis of Schnyder corneal dystrophy and with the symptoms of photophobia, glare and low visual acuity (VA) more affected in photopic condicions. VA in a lit room was 0.04 in the right eye and 0.02 in the left eye which did not improve with correction and corneal sensation was reduced. Slit-lamp biomicroscopy revealed bright, refractile crystals in the anterior stroma of the central and paracentral cornea bilaterally. Patient had severe panstromal disk-like opacity affecting pupillary axis, prominent arcus (senilis) lipoides in both eyes and mid-peripherial corneal haze that fills in the area between the corneal opacity and the peri­pherial arcus (Fig. 1). Fundus examination was not possible because of medias opacity. US B exam showed retina attached in both eyes. A LSCM with a diode laser of 670 nm wavelenght (HRT3/RostockCorneaModule: Heidelberg Engineering, Drossenheim, Germany) was also performed obtaining two-dimensional confocal images of the different corneal layers. Normal epithelial cells, large accumulations of hiperreflective rectangular and needle-shaped material in the subepithelial and anterior stroma were observed. The basal epithelial/ subepithelial nerve plexus and fibres were absent and the keratocytes of mid and posterior stroma were not visualized properly probably because of hiperreflection of anterior structures. Due to the same reason the endothelial cells were not seen (Fig. 2). Subsequently, the corneas of the patient were scanned with central anterior asterisk scan patterns, by a high-speed, high-resolution, spectral optical coherence tomography (SOCT Copernicus Plus: Optopol, Zawiercie, Poland). The images obtained indicated that the crystalline deposits were localized within the anterior stroma and reflectivity of Descemet membrane and endothelial cells was increased (Fig. 3).
The serum lipid levels showed hypercholesterolemia (HDL 66 mg/dl) and hypertriglyceridemia (TG 184 mg dl) although the patient was on medical treatment from the moment of the myocardial infarction. He did not refer the genu valgum nor family history of corneal disease.
The patient underwent penetrating keratoplasty (PKP) in the left eye. Immediate postoperative evaluation reveled: VA 0.06 in the left eye and the slit-lamp examination showed corneal button to be transparent with a few Descement folds. No evidence of epithelial defect was observed (Fig. 4). After 2 months follow-up the VA is 0.1 and BCVA is 0.3 in the left eye and the corneal graft remained transparent (Fig. 5).

Discussion
Weiss (7) reported that patients with SCD older than 39 years had evidence of diffuse stromal haze occuppying the whole diameter of the cornea, which causes significant decrease of VA as seen in case of our 68-year-old patient. Cholesterol crystals may affect VA by diffraction of light, resulting in glare and photophobia, major complaints of our patient.
Morphological evaluation on SCD by in vivo LSCM and SOCT serve to clarify the clinical findings of this dise­ase. The glare and the photopic visual acuity diminished is caused by light dispersion on the deposits of crystalline material in the anterior stroma. LSCM and SOCT highlighted in our case the morphological changes at the level of Bowman’s membrane and anterior stroma with an accumulation of crystalline deposits, which is consistent with the result of the previous histological study of SCD (16-18). The reduced corneal sensation postulated by Wiess (7) found an explication by revealing that the basal epithelial/subepithelial innervation is destroyed in older patients. As our patient belongs to the group with the most advanced corneal changes and the most prominent symptoms according to Weiss classification based on corneal findings with patient’s age (5), we observed the alteration of nerve structures and we found the corneal sensation decreased.
Other finding from in vivo LSCM study is that the shape and numbers of crystalline deposits in the anterior stroma differ among patients despite quite similar clinical appearance (18). Whether an association between deposit size and numbers and genotype exist, needs further confocal and mutiational analysis using a large number of patients with SCD as suggested in Kobayashi study (19).
Unfortunately, there is no local or systemic medical treatment available to stop the progression of corneal lipid deposition resulting in cornea opacification. Recognized surgical methods are phototherapeutic keratectomy (PTK), penetrating keratoplasty (PKP) and deep anterior lamellar keratectomy (DALK). The choice of the treatment depends on the case severity and the depth of lipids deposits. In the early stages when we have only subepithelial distribution of crystalline material we can use the PTK to obtain visual acuity improvement. It is considered to prevent accumulation of crystaline deposits in the anterior stroma better then anterior keratectomy (14) and compared with PKP and DALK is a less invasive method, but is effective only when the crystals are located in superficial cornea (20). The choice between PKP and DALK is also based on the depth of corneal affectation. Weiss (5) showed that, contrary to what is usually assumed, SCD affected the entire thickness of the corneal stroma in the majority of elderly patients, although the crystals were always subepithelial. Dissolved cholesterol or lipid or both have even been found in the basal epithelial cells and endothelial cells (3). Affectation of endothelium makes the selection of DALK impossible. This is the reason why study of corneal morphology in vivo is so important. The analysis of corneal structures helps to make the right decision on the surgery method and confocal microscopy and OCT contribute far more information than biomicroscopy, even if the resolution of the images of the more posterior structures is decreased because of the high reflectivity of the more anterior structures caused by intense crystalline deposition (14). We opted for PKP as the best choice for our patiens because of SOCT (hiperreflectivity of posterior complex: Decemet membrane and endothelium) and LSCM (increase reflectivity of stroma) images strongly suggested the entire cornea affectation. The choice of PKP among elderly patients is concerned with the results of Weiss study which demonstreted that the incidence of PKP depends on patients age and that among elderly patients amounts to 77% (7).
In conclusion, in vivo laser confocal microscopy and spectral optical coherent tomography, new emerging non-invasive technologies, proved to be a useful tool in SCD handling.

References:
1. Weiss JS, Kruth HS, Kuivaniemi H et al.: Mutations in the UBIAD1 gene on chromosome short arm 1, region 36, cause Schnyder crystalline corneal dystrophy. Invest Ophthalmol Vis Sci 2007, 48, 5007-5012.
2. Orr A, Dube MP, Marcadier J et al.: Mutations in the UBIAD1 gene, encoding a potential prenyltransferase, are causal for Schnyder crystalline corneal dystrophy. PLoS ONE 2007, 2, e685.
3. Weiss JS: Visual morbidity in thirty-four families with Schnyder crystalline corneal dystrophy (an American Ophthalmological Society thesis). Trans Am Ophthalmol Soc 2007, 105, 616-648.
4. Rodrigues MM, Kruth HS, Krachmer JH, Willis R: Unesterified cholesterol in Schnyder’s corneal crystalline dystrophy. Am J Ophthalmol 1987, 104, 157-163.
5. Weiss JS: Schnyder’s dystrophy of the cornea: a Swede-Finn connection. Cornea 1992, 11, 93-101.
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7. Weiss JS: Schnyder corneal dystrophy. Curr Opin Ophthalmol 2009, 20, 292-298.
8. Weiss JS: Schnyder crystalline dystrophy sine crystals: recommendation for a revision of nomenclature. Ophthalmology 1996, 103, 465-473.
9. Weiss JS, Møller H, Lisch W et al.: The IC3D classification of the corneal dystrophies. Cornea 2008, 27, S1- -S42.
10. Cavanagh HD, Petroll WM, Alizadeh H et al.: Clinical and diagnostic use of in vivo confocal microscopy in patients with corneal disease. Ophthalmology 1993, 100, 1444 -1454.
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12. Kobayashi A, Sugiyama K: In vivo corneal confocal microscopic findings of palisades of Vogt and its underlying limbal stroma. Cornea 2005, 24, 435-437.
13. Kobayashi A, Sugiyama K, Huang AJ: In vivo confocal microscopy in patients with central cloudy dystrophy of Francois. Arch Ophthalmol 2004, 122, 1676-1679.
14. Kobayashi A, Sakurai M, Shirao Y et al.: In vivo confocal microscopy and genotyping of a family with Thiel-Behnke (honeycomb) corneal dystrophy. Arch Ophthalmol 2003, 121, 1498-1499.
15. Vesaluoma MH, Linna TU, Sankila EM et al.: In vivo confocal microscopy of a family with Schnyder crystalline corneal dystrophy. Ophthalmology 1999, 106, 944-951.
16. Freddo TF, Polack FM, Leibowitz HM: Ultrastructural changes in the posterior layers of the cornea in Schnyder’s crystalline dystrophy. Cornea 1989, 8, 170-177.
17. Garner A, Tripathi RC: Hereditary crystalline stromal dystrophy of Schnyder. Br J Ophthalmol 1972, 56, 400--408.
18. Weller RO, Rodger FC: Crystalline stromal dystrophy: histochemistry and ultrastructure of the cornea. Br J Ophthalmol 1980, 64, 46-52.
19. Kobayashi A, Fujiki K, Murakami A et al.: In vivo Laser Confocal Microscopy Findings and Mutational Analysis for Schnyder’s Crystalline Corneal Dystrophy. Ophthalmol 2009, 116, 1029-1037.
20. Koksal M, Kargi S, Gurelik G et al.: Phototherapeutic Keratectomy in Schnyder Cristalline Corneal Dystrophy. Cornea 2004, 23, 311-313
 



Fig. 1. Slit-lamp biomicroscopy – the right and left cornea with severe panstromal central disk-like opacity, prominent arcus lipoides and mid-peripherial corneal haze. The anterior stroma of the central and paracentral with crystallline deposits.
Ryc. 1. Lampa szczelinowa – rogówki prawa i lewa z dyskoidalnym centralnym zmętnieniem śródmiąższowym, wyraźnie zaznaczoną obwódką starczą i przymgleniem wypełniającym przestrzeń między zmętnieniem centralnym a obwódką starczą. Osady krystaliczne znajdują się w przedniej części istoty właściwej centralnej i paracentralnej rogówki.
 







Fig. 2. In vivo laser scanning confocal microscopy – a. normal basal epithelial cell layer,
b. subepithelial stroma, c. anterior stroma, d. midstroma with numerous rectangular and needle- shaped crystals and increased background intensivity, e. keratocyte nuclei undetectable in mid and posterior stroma.
Ryc. 2. In vivo laserowa skaningowa mikroskopia konfokalna – a. prawidłowa warstwa nabłonkowa, b. podnabłonkowa część istoty właściwej, c. przednia istota właściwa, d. środkowa istota właściwa z dużą liczbą kryształów czworokątnych i w kształcie igły oraz zwiększoną intensywnością tła, e. keratocyty z niewidocznymi jądrami w częściach istoty właściwej – środkowej i tylnej.



Fig. 3. Spectral optical coherence tomography – hiperreflecivity of crystalline deposits localized within the anterior stroma and hyporeflecivity of more profund layers of the both corneas, hiperreflectivity of Descemet membrane.
Ryc. 3. Spektralna koherentna tomografia optyczna – hiperrefleksyjne pasmo złogów w przedniej istocie właściwej oraz hiporefleksyjność głębszych warstw obu rogówek, hiperrefleksyjność błony Descemeta.


Fig. 4. Slit- lamp biomicroscopy – the left eye in the 12th day after PKP.
Ryc. 4. Lampa szczelinowa – lewe oko w 12. dobie po keratoplastyce drążącej.


Fig. 5. Slit- lamp biomicroscopy – the left eye in the 2nd month after PKP.
Ryc. 5. Lampa szczelinowa – lewe oko 2 miesiące po keratoplastyce drążącej.

 


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