Up until now, the diagnosis of corneal disease has been mainly based on a slit-lamp biomicroscopy examination (Figures 1A and 2A). This exam allows ophthalmologists to evaluate (1) the state of the corneal epithelium with a fluorescein test, (2) the smoothness, transparency, and approximate thickness of the cornea, and (3) the presence of pigment granules or inflammatory cells in Descemet's membrane. Slit-lamp biomicroscopy, however, cannot be used to analyze the individual layers of the cornea. For this, we need the confocal microscope.
All layers of the cornea—namely the epithelium; Bowman's membrane and subepithelial nerve plexus; anterior, central, and posterior corneal stroma; Descemet's membrane; and corneal endothelium—may be observed with the confocal microscope.1,2 Confocal biomicroscopy of normal corneas3 has established grounds for analysis of the cornea in pathological conditions, allowing for an early diagnosis of corneal inflammation, dystrophy, and degeneration; monitoring healing processes after refractive and other surgical procedures; estimation of the pathology location; the course of graft rejection after keratoplasty; and differential diagnosis of the inflammatory processes taking place in corneal structures.1,2,4-9
IN THE COURSE OF KERATITIS
In the course of keratitis, blurred structures of the corneal stroma increase reflectivity in the region of inflammation, and activated keratocytes may be visualized (Figure 1B).
The aggravation of the inflammation is estimated on the basis of the extensivity changes in the corneal stroma, the state of the corneal structures between the inflammatory regions, and the condition of the endothelial cells. During the inflammatory processes, corneal structure changes depicted in confocal images are characteristic for the etiological factor, whereas the clinical state and slit-lamp biomicroscopic examinations do not always establish an etiologic factor. Early diagnosis of keratitis is very important, because effective treatment should begin as soon as possible.1,2,4,7,10,11
Although the clinical image of fungal, acanthamoebic, or viral infection lesions tends to be characteristic, the etiologic factor is not always determined. Therefore, confocal microscopy is of the greatest diagnostic importance to differentially diagnose fungal or amoebic keratitis.
The characteristic feature of keratomycosis is the presence of light-longitudinal filamentous structures of enhanced reflectivity (ie, mycelium) (Figure 1C). This cannot be visualized with slit-lamp biomicroscopy. In cases of fungal keratitis, the confocal microscopy examination reveals changes within the corneal epithelium, strong desquamation of the superficial epithelial cells, and polymegatism and polymorphism of the basal epithelial cells with increased reflectivity. Just below the level of Bowman's membrane, atrophic nerve fibers; changes in pattern and shape of keratocytes, as well as areas without keratocytes; and collagen fiber overproduction are observed. Inflammatory cells, numerous round high-reflectivity bodies (ie, were not keratocytes but fungus-like bodies), and an aggregate of mycelium is also seen (Figure 1D).
An acanthamoebic infection is indicated by the presence of characteristic cystic formations around changes with high reflectivity and dark hyporeflective regions. In this disease, active trophozoites are also seen (Figures 2B and 2C).
In the presence of adenoviral infection, corneal structures appear differently on confocal microscopy, depending on the clinical state, stage, and individual course of the disease. On confocal microscopy, distortions were seen in the corneal epithelium at an early stage of the disease. In the late stage, we observed diffuse hyporeflective areas of scar tissue within the corneal stroma.
In some cases, confocal microscopy visualizes the presence of needle-like structures (ie, activated keratocytes that profess an active inflammatory process or immunological reaction) (Figure 2D). These structures are observed in the course of corneal inflammation, during the healing process of refractive procedures, after penetrating keratoplasty, and in some cases when visual acuity decreases—without reason—in the clinical examination of the anterior and posterior parts of the eye.
SCARS, DYSTROPHIES, DEGENERATIONS
Confocal microscopy enables us to determine corneal structures, visualizing the pathological areas and surrounding tissue.2,9,12 Anterior corneal scar, irrespective of its etiology, may significantly reduce vision and cause diplopia, glare, light sensibility, and/or discomfort. Often, decreased clarity, diplopia, or glare cannot be corrected with spectacles or contact lenses. In confocal examination, we may estimate the endothelial cells' condition, the scar penetration depth, and the state of the tissue remaining around the scar. All help us decide on a proper treatment (ie, PTK, lamellar keratoplasty, penetrating keratoplasty).
Confocal images of corneal dystrophy differ, depending on the type, severity, and clinical state of the degenerative lesion. Blurred epithelial structure, excessive exfoliation, and cystic formations within the epithelium are observed in patients with epithelial dystrophy. In some cases, blurred collagen fibers, atrophic nerve plexus, distorted nerve fibers, a distorted pattern of keratocytic nuclei within the corneal stroma, and areas of scar-like tissue with increased illumination are seen. In cases with basement membrane dystrophy, the distortion not only compromises the epithelium, basement membrane, and Bowman's membrane, but it reaches the anterior corneal stroma. In epithelial dystrophy (ie, Messman's dystrophy), round, cystic, well-scraped formations are located between the basal cell and Bowman's membrane (Figure 3A). Superficial epithelial cell desquamation with enlarged edematous cell nuclei and amorphotic hyperreflective changes in the basal cell layer are seen.
In basement membrane dystrophy (ie, Cogan's dystrophy), linear light opacities in the corneal epithelium's basal cells and folds between the basal cells and the anterior corneal stroma—caused by subepithelal dystrophic material located beneath the cells and stroma—are observed (Figure 3B).
Bowman's membrane dystrophy presents changes in Bowman's membrane, which is replaced by dystrophic material with high reflectivity and irregular shape.
In granular dystrophy (ie, Groenouw type I), reflective deposits resembling breadcrumbs are present in the corneal epithelium (Figure 3C). Confocal images of macular dystrophy (ie, Groenouw type II) show cluster deposits, in the shape of pseudorings, just beneath Bowman's membrane. Scar tissue localized in the deeper part of corneal stroma is seen (Figure 3D).
The best documented corneal structural changes are in the course of Fuchs' dystrophy,2,12,13 however, they differ depending on the stage. In the earliest stage, changes are not visible in slit-lamp biomicroscopy, but they may be visualized in confocal microscopy (Figure 4A). In the early stage of Fuchs' dystrophy, slit biomicroscopy reveals fine dark spots within the corneal endothelium, whereas in the advanced stage, the cornea has the appearance of wrought metal (Figure 4B). On confocal microscopy, diffused hyporeflective areas are seen in the early stage of the disease. Endothelial cells located beyond these areas were pleomorphic and polymegathic. In the late stage, we observe diffused hyporeflective areas surrounded by hyperreflective endothelial cells, which cannot be separately analyzed. Within the corneal stroma, collagen fibers are blurred, and background illumination is increased. In the posterior part of the stroma, dark bands are seen, and the epithelium contains cystic structures. The membranes of the basal cells are thickened, and the background illumination is increased (Figures 4C and 4D).
Chiou et al13 noticed that confocal microscopy allows a differential diagnosis between guttae (ie, characteristic for Fuchs' dystrophy, deep intrastromal corneal inflammation, posterior polymorphous dystrophy) and pseudoguttae (ie, clinical signs of keratitis).
ANTERIOR SEGMENT SURGERY
If graft rejection occurs after penetrating keratoplasty, the following are observed in the cornea: epithelial thickness diminishes, boundaries of the basal epithelial cells thicken, structures of collagen fibers blur, the number of keratocytes and inflammatory cells decrease (ie, lymphocytes and fibroblasts), and needle-like structures are seen. Even in the early phase of graft failure, the cell structure is blurred, and the basal corneal cell membranes are considerably thickened.8
After cataract surgery, confocal images make it possible to assess the probability of bullous keratopathy.
Confocal microscopy allows in vivo presentation of corneal structure changes on the cellular level. Its noninvasive character offers enormous possibilities that enable us to gradually discover the processes taking place in the corneal structures after excimer laser photoablation, both after refractive and therapeutic procedures. After refractive surgery, confocal images show morphologic appearances of the corneal structures that depend on the applied refractive procedure, postoperative period, and region of the examined cornea.2,5,6
Confocal microscopy is a young corneal diagnostic method that enables morphologic and morphometric assessment of all corneal layers. This examination may have important diagnostic meaning in (1) differentiation of the etiological factor of corneal inflammation, (2) assessment of corneal structures in the early stages of corneal dystrophy, (3) early diagnosis of bullous keratopathy, and (4) prevention of graft rejection.
In some states of corneal disease, the confocal microscope examination helps to decide whether to qualify the patient for PTK or more radical management, including corneal transplantation.
Ewa Mrukwa-Kominek, MD, PhD, is from the Department of Ophthalmology, Silesian Medical University, in Katowice, Poland, where Professor Ariadna Gierek-Lapinska is Department Head. Dr. Mrukwa-Kominek states that she has no financial interest in the products or companies mentioned. She is a member of the CRST Europe Editorial Board. She may be reached at +48 601528850; emrowka@poczta.onet.pl.
Stanislawa Gierek-Ciaciura, MD, PhD, is from the Department of Ophthalmology, Silesian Medical University, in Katowice, Poland. Dr. Gierek-Ciaciura states that she has no financial interest in the products or companies mentioned. She may be reached at +48 601898896; ciaciura@kil-okul.katowice.pl.
Up Front | May 2007
Application of Confocal Microscopy
This tool diagnoses corneal diseases by analyzing the layers of the cornea.
Ewa Mrukwa-Kominek, MD, PhD, and Stanislawa Gierek-Ciaciura, MD, PhD
