Quantitative analysis of immunogold labellings of collagen types I,III, IV and VI in healthy and pathological human corneas

Background: We studied the distribution of collagen types I, III, IV and VI in one healthy human cornea and in seven pathological human corneas, in which the disorders were three cases of pseudophakic bullous keratopathy (two severe, one moderate) and one case each of stage IV keratoconus, chronic ulcer, vascularized cornea and disciform keratitis. Methods: Transmission electron microscopy examinations were performed on post-embedding immunogold-labelled sections. The staining was evaluated by gold particle count in the different tissues. The presence or absence of a given antigen was determined by statistical analysis, using a d-value test. Results: Our results on healthy corneal tissues corroborate the data available from previous studies, except for collagen type VI, which we found to be absent in Bowman's layer. In pathological corneas with a collagenous layer posterior to Descemet's membrane, collagen types I, III and especially IV were detected in this collagenous layer. Collagen types I, III and VI were detected in the anterior healed stroma of other pathological corneas, except for the keratoconus cornea, in which intense collagen III staining was observed. Conclusion: The presence of collagen types I and III in the posterior collagenous layer of our pseudophakic bullous keratopathy corneas suggests that this layer corresponds to scar tissue secreted by stimulated endothelial cells.     

圆锥角膜的免疫组织化学研究

目的：了解圆锥角膜基底膜免疫组化的变化,以发现圆锥角膜可能的特异性改变,为探讨其发病机制提供依据。方法：收集圆锥角膜在我院眼科行穿透性角膜移植术的角膜片,用链霉亲和素－生物素法（SP法）检测圆锥角膜、角膜白斑、正常角膜基底膜Ⅳ型胶原、Fn、Ln表达的异同,并进行统计学分析。结果：圆锥角膜基底膜Ⅳ型胶原,Fn染色较正常角膜显著升高,但与角膜白斑比较无显著性差异。结论：圆锥角膜基底膜的改变可能与创伤愈合有关。提示圆锥角膜原发病变可能在角膜基质细胞。眼科学报2000;17：65－67。     

Altered lectin binding sites in keratoconus corneas.

We investigated the glycoconjugates in frozen sections of keratoconus corneas, using a panel of 12 biotin- or fluorescein isothiocyanate-labeled lectins. No differences between the lectin binding sites of the epithelium, endothelium and Descemet's membrane of normal and keratoconus corneas could be observed. However, in contrast to normal corneas, intense staining with peanut agglutinin (PNA) could be detected at breaks in Bowman's layer, in scar tissue and in the adjacent stroma. Furthermore, in the majority of cases binding sites for Phaseolus vulgaris erythroagglutinin (PHA-E) and increased staining with Ricinus communis agglutinin I (RCA-I) and Lens culinaris agglutinin (LCA) could also be detected in ruptures in Bowman's layer and in scar tissue. These data suggest that the scarred regions of the anterior stroma in keratoconus corneas may contain oligosaccharides with terminal D-galactose (beta 1-3)-D-N-acetylgalactosamine disaccharides (recognized by PNA), increased amounts of glycoconjugates with terminal beta-galactose residues (recognized by RCA-I), increased amounts of glycoconjugates with glucose/mannose residues (recognized by LCA), and finally, biantennary complex-type glycopeptides containing two outer galactose residues and a residue of N-acetylglucosamine (recognized by PHA-E). Since corneal scars due to causes other than keratoconus revealed lectin binding sites (particularly for PNA and to a lesser extent also for PHA-E, LCA and RCA-I) similar to those seen in scar tissue of keratoconus corneas, we conclude that it is mainly scar formation that may be responsible for the altered lectin binding sites in keratoconus.     

Explorative study of interleukin levels in the human cornea

Background: The presence of interleukins has been demonstrated in the cornea and other ocular tissues. Although pathogenic mechanisms are unknown, interleukins seem to be involved in inflammatory disorders of the cornea. The present study was undertaken to analyse concentrations of interleukin- I (IL-1) and interleukin-6 (IL-6) in human corneas with various clinical diagnoses. Methods: Immediately after keratoplasty 127 explanted human corneas with various corneal diseases were snap frozen and cryosections were prepared for histological examination. Furthermore, the protein content was measured according to the method of Bradford and the concentration of IL-1 and IL-6 were determined using a specific immunosorbent test (ELISA). Results: It was found that IL-1 and IL-6 level were clearly higher in corneas with ulcerations and distinct inflammatory signs. Lower levels of both interleukins were found in corneas with a weak expression of inflammatory signs. Conclusions: Keratitis, keratoconus with inflammatory signs, and ulcerating processes showed higher interleukin levels than corneas with non-inflammatory disorders like scar formation, corneal dystrophy and keratoconus. The results could show that, depending on the clinical diagnosis, the inflammatory status of the cornea may be evaluated by the interleukin levels determined in the corneal tissue.     

Histopathologic and electron-microscopic futures of corneal and scleral collagen fibers in osteogenesis imperfecta type III

Background: This report describes the histopathologic and electron-microscopie features of an eye from a patient with osteogenesis imperfecta type III. In particular, the diameters of corneal stromal and scleral collagen fibers were determined. Methods: The eyes of an 18-year-old white male with osteogenesis imperfecta type III were examined by light arid electron microscopy arid the pathological features were compared with an age-matched control eye. Results: The cornea was clear. The sclera had a blue color and was moderately thinned, especially at the equator. Light microscopy revealed absence of Bowman's layer. Transmission electron microscopy confirmed complete absence of Bowman's layer without evidence of scarring or inflammation. The collagen fibers of the corneal stromal lamellae were about 25% narrower than in the control, but the cornea was otherwise unremarkable ultrastructurally. The collagen fibers of the sclera were approximately 50% narrower than in the control and were much more uniform in size. Prominent portions of clastic fibers, which are usually only present in a small number in the inner portion of the sclera, were prescrit throughout the sclera. Conclusion: We propose that it is the uniformity of the scleral collagen fibers which gives the sclera translucence, producing the blue color often observed clinically in osteogenesis imperfecta. Absence of Bowman's layer of the cornea did not interfere with the stability of the cornea in this case. This appears to be the first published pathological examination of the eye in ostcogenesis imperfecta type III.     

Ex vivo phosphorus magnetic resonance spectroscopy on eye bank corneas and corneal metabolic health

Background: Since a potential exists for untoward effects on the cornea from the high magnetic fields and radio-frequency energies, and the further manipulation required for phosphorus-31 magnetic resonance spectroscopy (³¹P-MRS), we determined the effects of this technology on tissues using paired human corneas (n=4) meeting criteria acceptable for transplantation. Methods: Slit-lamp biomicroscopy, pachometry, specular microscopy, and redux fluorophotometry were performed on all corneas. One cornea of each pair was examined (<30 min) by³¹P-MRS. Following³¹P-MRS, slit-lamp biomicroscopy, pachometry, and redox fluorophotometry were again performed. Results: Data tabulated included the³¹P energy modulus (1.37±0.28), the ATP/Pi (2.92±0.59) and SP/Pi (0.76±0.04) ratios, and the intracorneal pH (7.24 ±0.09). Conclusion: Since there were no significant differences in slitlamp biomicroscopy, endothelial density and morphometry, cell counts, and pachometric and redox fluorophotometric measurements between corneas of each pair before and after³¹P-MRS. analysis, it was concluded that there was no detectable metabolic damage secondary to such analysis. This study suggests that MRS analysis of human eye-bank tissues does not damage the cornea metabolically and may provide practical evaluation of the health of the cornea at the biochemical level.     

Efficacy of iontophoresis in the rat cornea

Background: Iontophoresis can enhance penetration of drugs into tissues. We examined the extent of penetration of gentamicin into the cornea of rats during iontophoresis and the effect of varying the concentrations of gentamicin, the duration of iontophoresis and the current densities during iontophoresis. Methods: Eight groups of rats underwent corneal iontophoresis using gentamicin dissolved in agar. Low and high concentrations of gentamicin were used, as well as low and high current densities and long and short durations of iontophoresis. Control groups received topical or subconjunctival gentamicin, topical saline solution and mock iontophoresis with the agar-gentamicin mixture. The Mann-Whitney test was used for statistical evaluation. Results: Highly bactericidal concentrations of gentamicin were obtained in all the iontophoresis-treated corneas. The high concentration compared to the low concentration of gentamicin in agar significantly increased the concentration of gentamicin in the corneas, as did the longer duration of iontophoresis. However, higher current intensity did not significantly enhance the drug concentration in the cornea. Conclusion: lontophoresis with a concentrated gentamicin-agar mixture may provide a rapid increase of levels in the cornea.     

Second-harmonic imaging microscopy of normal human and keratoconus cornea

PURPOSE: The purpose of this study was to evaluate the ability of second-harmonic imaging to identify differences in corneal stromal collagen organization between normal human and keratoconus corneas. METHODS: Six normal corneas from eye bank donors and 13 corneas of patients with keratoconus, obtained after penetrating keratoplasty were examined. A femtosecond titanium-sapphire laser with 800-nm output was used to generate second-harmonic signals collected at 400 nm from central and paracentral corneal tissue blocks. Three-dimensional (3-D) data sets were collected and reconstructed to evaluate the location and orientation of stromal collagen lamellae. RESULTS: Imaging of second-harmonic signals combined with 3-D reconstruction of the normal cornea identified a high degree of lamellar interweaving, particularly in the anterior cornea. Of note was the detection of lamellae that inserted into Bowman's layer and were oriented transverse to the corneal surface, penetrating posteriorly approximately 120 mum. In keratoconus corneas, imaging second-harmonic signals identified less lamellar interweaving and a marked reduction or loss of lamellae inserting into Bowman's layer in 12 of 13 cases, particularly in regions associated with cone development without breaks in Bowman's layer or scarring. CONCLUSIONS: Compared with normal adult corneas, marked abnormalities were detected in the organization of the anterior corneal collagen lamellae of keratoconus corneas by second harmonic imaging. These structural abnormalities are consistent with the known changes in collagen organization and biomechanical strength of keratoconus.     

Histopathological variation in keratoconus.

During examination of 131 penetrating keratoplasty specimens from patients with keratoconus obtained in an 11-year period, we observed two histopathologic variants based on the appearance of Bowman's layer and the corneal epithelium. "Typical" keratoconus specimens had multiple breaks in Bowman's layer and central epithelial thinning, whereas "atypical" corneas lacked breaks in Bowman's layer and had less thinning of the central epithelium. Ninety-five corneas were from patients who underwent grafting in only one eye. Seventy-six (80%) of these corneas were "typical" and 19 corneas (20%) were "atypical" in appearance. Both variants had similar degrees of central stromal thinning. Patients with "typical" and "atypical" corneas differed demographically by race only; 49% of "typical" and 95% of "atypical" corneas were from white individuals. Thirty-six corneas were from 18 patients who underwent bilateral penetrating keratoplasty. The histologic appearance of these corneal pairs was concordant in 13 patients and discordant (one "typical" and one "atypical" cornea) in five patients. Statistical analysis indicated that this distribution is not significantly different from that predicted by chance and suggests that "typical" and "atypical" corneas are manifestations of the same disease process.     

Immunohistochemical localization of chondroitin sulfate proteoglycan and tenascin in the human eye compared with the HNK-1 epitope

Background: A previous study revealed the HNK-1 epitope in the human ciliary body beneath the ciliary epithelium. The molecules bearing this 3-sulphoglucuronic acid-containing oligosaccharide epitope in the eye remain unknown. As chondroitin sulphate proteoglycan (CSPG) and tenascin are potential candidates as bearers of the HNK-1 epitope, their distribution in the human eye was compared with that of the HNK-1 epitope. Methods: Fifty-five formalin-fixed, paraffin-embedded human eyes, including 20 normal eyes and 35 eyes with exfoliation syndrome or glaucoma, were studied immunohistochemically with monoclonal antibody (MAb) CS-56 to CSPG, MAb TN2 to tenascin, and MAbs HNK-I and VC1.1 to the HNK-1 epitope. Additionally, four frozen lens capsules with exfoliation material were studied by indirect immunofluoresence. Results: A population of dendritic cells in the inner connective tissue layer of the ciliary body and exfoliation material were immunoreactive with antibodies to the HNK-1 epitope, but no labelling for CSPG and tenascin was seen in them, including frozen sections. The inner surface of the nonpigmented ciliary epithelium was reactive for the HNK-1 epitope, and at the ora serrata also for CSPG. In some eyes with glaucoma, immunoreaction for CSPG and tenascin was seen beneath the epithelium and endothelium of the cornea. The nerve fibre layer of the retina was labelled for tenascin. In the sclera, all antibodies labelled the ground substance, and in some large blood vessels immunoreaction for CSPG and tenascin was seen subendothelially. Conclusion: Apart from the sclera, the distribution of CSPG and tenascin was different from that of the HNK-1 epitope, suggesting that this carbohydrate epitope may not be borne by these molecules in the human ciliary body.     

The corneal stroma: an inhomogeneous structure

Purpose: This study was conducted to determine the elemental composition of the human cornea. Special attention was paid to corneal stroma inhomogeneity. Methods: Seventy human corneas were examined by means of energy-dispersive X-ray analysis. Epithelium, subepithelium, middle stroma, sub-Descemet layer, Descemet's membrane and endothelium were subjected to repeated measurements. Results: In the cellular layers the phosphorus concentrations were high [0.35 mol/kg dry weight (dw) in the epithelium and 0.403 mol/kg dw in the endothelium]. Similar concentrations were found for sulphur (0.38 mol/kg dw in the epithelium). Stromal layers showed high contents of sulphur: 0.26 mol/kg dw. The phosphorus concentration was found to be higher in the subepithelium than in the middle stroma. Sulphur concentrations were highest in Descement's membrane, followed by the subepithelium and the middle stroma. Discussion: Nucleic acids and energy-containing phosphates explain the high levels of phosphorus in the cellular layers. The high sulphur concentrations may be related to the phosphoadenosinphosphosulfate and protein turnover in the epithelium. We interpret the inhomogeneous distribution of phosphorus in the stroma as a function of the density of keratocytes. An evalulation of all known sulphur-containing biochemical components of the stroma (0.217 mol sulphur/kg dw) corresponds to our measurements. In contrast to former results we find the corneal stroma to be an inhomogeneous structure.     

A SEM-study of a keratoconus and an artificially aged human cornea

The substructure of both the epithelial and endothelial surfaces of a keratoconus and an artificially aged cornea was compared with that of a healthy cornea by investigating them with a scanning electron microscope.From the depressions around the protruding centre of the epithelial surface of the keratoconus cornea, and from the whole epithelial surface of the artificially aged cornea, cells detached themselves, assuming a more or less rounded shape.The endothelial surface of both the keratoconus and the aged cornea showed areas of cells with an almost completely disintegrated cell membrane, exposing the cell contents.On the endothelial surface of the keratoconus cells were found with a missing cell-nucleus and a perforated cell membrane, due to a Kammerwasser Einbruch effect.The work was carried out at the Centre for Medical Electron Microscopy.     

Electrolytes in the cornea: a therapeutic challenge

Background: Reported here are the results of electrolyte measurements in different layers of 70 apparently normal human corneas. Methods: Samples were examined by energy-dispersive X-ray analysis under calibrated conditions in a scanning electron microscope. The method allows the simultaneous quantitative analysis of, among others, sodium (Na), chloride (Cl), phosphorus (P) and potassium (K). The results are related to the dry weight of the analyzed samples. Four distinct layers, subepithelium, middle stroma, posterior stroma and Descemet's membrane, were analysed in each cornea. Results: In the middle stroma we found concentrations of: sodium 0.609 ± 0.13, chloride 0.557 ± 0.115, potassium 0.058 ± 0.02 and phosphorus 0.038 ± 0.01 (mmol/kg dry weight). Conclusion: The collation of normal electrolyte concentrations provides reference values for future studies on changes of the corneal electrolyte composition in diseased or injured eyes. The electrolyte composition of rinsing fluids or eye drops should be adjusted to that of the corneal stroma. Phosphate buffer, for example, is not a good vehicle for topical eye treatments and should be replaced by organic buffering systems.     

DNA-based HLA class II postmortem typing: evaluation of different techniques for prospective corneal allografting

Background: The objective of this study was to establish DNA-based HLA-DR postmortem tissue typing techniques in order to improve the quality and quanity of fully HLA-typed corneas for prospective allografting. Methods: Four hundred and thirty-seven cornea donors were investigated. DNA was derived from cultivated retinal pigment epithelial cells by spin column purification or a salting out technique, and from scleral tissue by a very simple boiling method. Donors were typed by hybridization of polymerase chain reaction (PCR) products with sequence-specific oligodesoxynucleotide probes (PCR-SSOP) or by PCR with sequence-specific primers (PCR-SSP). Twenty-two of the donors were pretyped by serology. Results: We observed high concordance (96%) between the results of DNA-based postmortem typing and the serological lymphocytotoxicity test. Furthermore, the distribution of the HLA-DR specificities that were detected correlated well with the distribution in a control population. Conclusions: The results of this study demonstrate that both PCR-SSP and PCR-SSOP allow prospective allocation of HLA class II-matched corneas with high accuracy.     

圆锥角膜的共焦显微镜表现临床分级

目的 观察临床上不同阶段圆锥角膜的共焦显微镜图像，推测圆锥角膜的病理发展过程，并进行圆锥角膜的共焦显微镜分级。方法 采用共焦显微镜（Confoscan 2．0），观察24例24眼不同发展阶段（lawless分期）圆锥角膜患者的角膜共焦显微镜图像，进行分析并提出焦显微镜下的临床分分期。结果 共焦显微镜下圆锥角膜首先出现了排列规则的裂隙状暗纹，随着病情的不断进展，病变逐渐由角膜后弹力层向前发展，逐渐累及角膜的后基质和前基质，同时角膜基质细胞核拉长、排列出现紊乱，前后基质细胞失去了原有特征。急性圆锥角膜的患者角膜基质细胞出现水肿，角膜瘢痕在共焦显微镜下呈强反光的无细胞样结构。讨论 共焦显微镜下圆锥角膜的病理发展过程，是从角膜后弹力层开始，由后向前发展，逐渐累及角膜的后基质、前期基质，最后角膜破裂引起急性圆锥角膜，角膜出现水肿，愈合后遗留瘢痕。据此提出圆锥角膜共焦显微镜分期将为四个阶段：第一阶段、角膜裂隙样暗纹累及角膜后弹力层；第二阶段、角膜裂隙样暗纹累及角膜后后基质；第三阶段、角膜裂隙样暗纹累及角膜前基质；第四阶段、出现角膜基质水肿或者瘢痕。     

Orthotopic corneal transplantation in the mouse — a new surgical technique with minimal endothelial cell loss

Background: The murine model of orthotopic perforating keratoplasty is important for studying the allograft reaction, but the small dimensions cause technical difficulties. Methods: The anterior chamber of the eye of the BALB/c mouse was measured with the confocal microscope and with histological methods. Ten C3H mouse donor corneas each were separated by the conventional technique and by the newly developed underwater technique, where the opened donor eye did not lose its shape under water. The corneal endothelium was stained with trypan blue and alizarin red S. Ten BALB/c (H-2d) mice received a corneal graft taken from a C3H (H-2k) mouse by the underwater technique. Results: The 3.7-mm eye of the BALB/c mouse has a corneal diameter of 3.5 mm. The cornea has a central thickness of 170 m, the epithelium comprising 30% and the stroma 70%. While none of the corneas separated by the new underwater technique evidenced endothelial damage, a 28 ± 17.0% defect of the endothelial surface was found with the conventional technique. All transplanted corneas were clear when the lids were opened on the 2 post-operative day and clouded between the 7th and 30th days (mean 16.5 days) due to an allograft reaction. Conclusion: The newly developed underwater technique is superior to the conventional technique, since floating of the very thin donor cornea during the separation procedure prevents endothelial defects by guarding against folds. By enabling reliable keratoplasty in the mouse, this technique facilitates studies on the experimental allograft reaction.     

Expression of betaig-h3 is lower than normal in keratoconus corneas but increases with scarring.

PURPOSE: Keratoconus is a progressive ectatic disease of the cornea. Despite extensive clinical and laboratory investigations, its pathogenesis remains unclear. In this study, we examined the localization of betaig-h3, a recently described extracellular matrix protein in keratoconus corneas both in the absence and presence of subepithelial scarring. METHODS: Two normal corneas and central corneal buttons of 10 patients with keratoconus were excised during perforating keratoplasty and examined, including one case with acute corneal hydrops. In one case, keratoconus was associated with Down syndrome. Immunodetection was done with an antipeptide antibody reacting with the N-terminal part of betaig-h3. RESULTS: We found decreased betaig-h3 levels in the basal epithelial layer and keratocytes of keratoconus corneas. In the scarred corneas, however, betaig-h3 levels were increased in the basal epithelial layers and in activated keratocytes at the places of scarring. In the cornea of the patient with Down syndrome, we found an additional betaig-h3-positive zone in the anterior stroma. CONCLUSIONS: The decreased levels of betaig-h3 corneas seem to be specific for keratoconus. Considering the putative role of betaig-h3 as a cellular-attachment protein, paucity of betaig-h3 in the corneal stroma may lead to decreased mechanical stability and contribute to the development of keratoconus.     

The alterations in corneal structure at III/IV stage of keratoconus by means of confocal microscopy and ultrasound biomicroscopy before penetrating keratoplasty

PURPOSE: The aim of the study was in-real time observation and morphological evaluation of the human corneas at III/IV stage of keratoconus, using the scanning slit confocal microscope Confoscan P4 and ultrasound biomicroscopy--UBM. MATERIAL AND METHODS: The patients with keratoconus were examined according to the Amsler scale. The material consisted of 12 corneas of 11 patients (8 men, 3 women), where assessment of the corneal structure was performed with the confocal microscope ConfoScan P4 (Tomey) and ultrasound biomicroscopy--UBM Model 840 (Humphrey Instruments). The comparison of different corneal regions (central and peripheral) was evaluated. RESULTS: The confocal microscopy and UBM revealed thinning of the layers of the corneal structure and pathological changes in the central area, especially at IV stage of keratoconus. The desquamating superficial cells were elongated, arranged around the apex of the cornea. Below the Bowman's membrane a considerable disarrangement of collagen fibers reflected by bright background illumination was observed. In the posterior part of the stroma the folds were detected. The examination of the cornea showed thickening in the peripheral part, central detachment of the Descemet's membrane and the endothelium from the posterior surface of the cornea. The thickness of the cornea varied from 0.201 to 0.384 mm in the central part and 0.675 to 0.740 mm in the peripheral area. CONCLUSION: Confocal scanning microscopy combined with ultrasound biomicroscopy enables the cornea to be examined in vivo. It can be used to localize pathological changes in individual corneal layers and to assess their extent.     

Keratocyte apoptosis associated with keratoconus.

Keratoconus is an ectatic corneal dystrophy associated with stromal thinning and disruption of Bowman's layer. The purpose of this study was to explore a possible association between keratocyte apoptosis and keratoconus. Keratocyte apoptosis was evaluated in corneas of patients with keratoconus, corneas of patients with stromal dystrophies, and normal donor corneas using the transferase-mediated dUTP-digoxigenin nick and labeling (TUNEL) assay. Keratocyte apoptosis was also studied in keratoconus and normal corneas using transmission electron microscopy. TUNEL-stained keratocytes were detected in 60% of corneas with keratoconus, but only 35% of corneas with stromal dystrophies (P =0.03). The number of TUNEL-positive keratocytes detected in the keratoconus, stromal dystrophy, and normal corneas was 7+/-1 (mean+/-standard error, range 0-20), 2+/-0. 8 (range 0-9), and 0+/-0 (range 0-0) TUNEL-positive cells per section, respectively. The differences between the keratoconus and the stromal dystrophy (P =0.0097) or the normal cornea (P =0.01) groups were statistically significant. The difference between the stromal dystrophy and normal cornea groups was not statistically significant (P =0.45). The stromal dystrophy group was included to account for surgery-associated keratocyte apoptosis. No TUNEL-stained keratocytes were detected in normal corneas. Cell morphologic changes consistent with apoptosis were detected by transmission electron microscopy (TEM) in keratocytes of keratoconus corneas, but not in keratocytes in normal corneas. Chronic keratocyte apoptosis associated with ongoing epithelial injury may link risk factors associated with keratoconus such as chronic eye rubbing, contact lens wear, or atopic eye disease. Similarly, increases that have been detected in several different degradative enzymes in keratoconus corneas could be associated with chronic keratocyte apoptosis and less than perfect control of release of intracellular contents.     

The effect of high-dose methylprednisolone on laser-induced retinal injury in primates: an electron microscopic study

Background: Previously we reported an ameliorative effect of high-dose methylprednisolone in laser injury to monkey retinas. The ultrastructural modification by methylprednisolone has not been examined. Methods: Cynomolgus monkeys were given severe kgrade III) retinal laser burns and treated with an intravenous megadose of methylprednisolone. Pathologic features of the retinal lesions with or without methylprednisolone treatment were evaluated by light and electron microscopy. Results: Ultrastructurally, the treated lesions showed rapid recanalization of choriocapillaris; proliferation of retinal pigment epithelium to replace the necrotic and damaged cells, resulting in rapid re-establishment of blood retinal barrier; mild macrophagic activity; and rapid reformation of the outer limiting membrane by Mueller cells. Conclusion: A high dose of methylprednisolone affected the responses of the choriocapillaris, retinal pigment epithelium, photoreceptor cells and Mueller cells to laser injury, showing an overall beneficial effect. These modifications might be ascribed to methylprednisclone's anti-inflammatory action, protection of the microcirculation and anti-lipid peroxidation effect. Proprietary interest: none