The role of apoptosis in the pathogenesis of Fuchs endothelial dystrophy of the cornea.

OBJECTIVE: To investigate the potential role of apoptosis in the pathogenesis of Fuchs endothelial dystrophy of the cornea. METHODS: Twenty-one corneal buttons from patients with Fuchs dystrophy and 15 control corneas were studied. Apoptosis was assessed by the in situ end-labeling of double-stranded DNA breaks, and by immunohistochemical characterization of cellular markers associated with apoptosis (Fas, FasL, Bcl-2, and Bax). Expression of Bcl-2 and Bax mRNA in the corneal stroma and endothelium was separately analyzed by a semiquantitative reverse transcriptase polymerase chain reaction. Furthermore, cultivated keratocytes generated from diseased corneal buttons and donor rims were exposed to camptothecin, an apoptotic inducer, for 6 and 24 hours. They were then examined for protein and messenger RNA (mRNA) expression of apoptotic regulatory molecules. RESULTS: DNA fragmentation was seen in the epithelium, stroma, and endothelium in 6 of 7 corneas with Fuchs dystrophy. A statistically significant difference was identified in the expression of Bax and its mRNA in the stroma, but not in the endothelium of Fuchs dystrophy corneas. Following exposure to camptothecin, keratocytes from patients with Fuchs dystrophy responded with an increased level of Bax and a low level of Bcl-2. This trend was distinctively different from the response of normal keratocytes. CONCLUSIONS: The evidence in this study points to a disease-related disturbance in the regulation of apoptosis in Fuchs dystrophy. Our findings suggest that excessive apoptosis may be an important mechanism in the pathogenesis of Fuchs dystrophy.

The role of matrix metalloproteinases in ulcerative keratolysis associated with perioperative diclofenac use.

OBJECTIVE: To investigate the role of matrix metalloproteinases (MMPs) in the pathogenesis of ulcerative keratolysis associated with topical use of generic diclofenac preoperatively and postoperatively. To characterize the inflammatory response of the cornea in this case of ulcerative keratolysis. DESIGN: Case report with clinicopathologic correlation. MAIN OUTCOME MEASURES: Corneal culture for microbial growth. Clinical and histopathologic examinations including routine histolopathologic, immunofluorescent, and immunohistochemical studies. RESULTS: Microscopic examination of the corneal button disclosed fibrinous material with neutrophils and mononuclear inflammatory cells. The corneal epithelial basement membrane was irregularly thickened and patchy. Immunohistochemical staining detected weak staining of MMP-1 and a strong presence of MMP-8 in the epithelium. MMP-8 and 9 were also present in areas of leukocytic infiltration. MMP-2 appeared in a few stromal cells. Macrophages and leukocytes were the predominant infiltrating cells. CONCLUSIONS: A nonspecific inflammatory response occurred in this case of ulcerative keratolysis. Corneal epithelial cells are capable of secreting MMP-1 and 8 and may participate in the stromal degradation and repair process of the ulcerative keratolysis associated with topical nonsteroidol antiinflammatory use.

The role of apoptosis in the early corneal wound healing after excimer laser keratectomy in the rat.

BACKGROUND: The potential role of apoptosis in corneal wound healing after excimer laser keratectomy was investigated in a rat model. METHODS: Lewis rats underwent laser keratectomy using a 193-nm excimer laser. The central corneas were ablated in three depths: group A, epithelium; group B, superficial stroma; group C, deep stroma. Eyes were collected at 1, 12, 24, and 36 h and 1 week. Cellular markers associated with apoptosis--Fas, Fas ligand (FasL), Bcl-2, and Bax were examined by immunohistochemistry. Keratocyte depletion and endothelial changes were evaluated histologically. In situ end labeling of double-stranded DNA breaks was used to demonstrate apoptosis in corneal sections. RESULTS: Keratocyte depletion was observed in 6 (50%) of 12 rats (total from groups A, B, and C) at 12 h, 11 (73%) of 15 at 24 h, 3 (20%) of 15 at 36 h, and 2 (15%) of 13 at 1 week after laser surgery. Corneal endothelial edema was observed in the ablation zone. Expression of Fas, FasL, Bcl-2, and Bax in corneal cells showed dynamics similar to that of keratocyte depletion and endothelial changes. There was less expression of apoptotic molecules in newly generated epithelial cells and more in endothelial cells of the stromal ablation groups. CONCLUSIONS: Excimer laser keratectomy triggered apoptosis of corneal keratocytes and endothelial cells. More endothelial edema was observed in the stromal ablation than in the epithelial ablation group. The expression of apoptotic molecules coincided with the period of keratocyte depletion and regeneration and of endothelial recovery, suggesting that apoptosis is a dynamic part of corneal wound healing and remodeling after excimer laser keratectomy.

Defensin gene expression in the cornea.

PURPOSE: To determine whether defensin genes are expressed in human corneas and bovine corneal keratocytes. METHODS: In situ hybridization and immunohistochemistry were used to localize defensin mRNA and protein in normal and diseased human corneas. Cultured bovine keratocytes were stimulated with IL-1alpha or TNFalpha to determine whether defensin mRNA production occurred. Reverse transcription polymerase chain reaction (RT-PCR) was performed to amplify defensin cDNA from cytokine-induced keratocytes, and Southern blots were used to verify the specificity of RT-PCR amplification products. RESULTS: Defensin mRNA and protein were not detected in normal human corneal stroma, but were readily detectable in the corneal stroma in cases of rejected transplants and postinfectious keratitis. IL-1alpha was a potent inducer of defensin gene expression in keratocytes, which began 12 h after challenge and peaked at 18 to 24 h. TNFalpha weakly induced defensin mRNA in keratocytes at about 18 h. Southern blots of the RT-PCR products probed with an oligonucleotide complementary to internal sequences of defensin demonstrated the appropriately sized products (198 bp) specific for defensin. CONCLUSIONS: This report demonstrates the presence of defensin in the human cornea and the capacity of corneal keratocytes to produce defensin mRNA in response to IL-1alpha and TNFalpha. Release of defensins by keratocytes in response to cytokines elaborated in corneal inflammation may contribute to the host defense response in microbial keratitis.

Inflammatory response in the early stages of wound healing after excimer laser keratectomy.

OBJECTIVE: To evaluate the inflammatory response and its potential role in the early stages of corneal wound healing after excimer laser keratectomy. MATERIALS AND METHODS: Lewis rats underwent excimer keratectomy using a 193-nm excimer laser. The central corneas were ablated in 3 depths: group A, epithelium; group B, superficial stroma; or group C, deep stroma. Eyes were harvested 1, 12, 24, and 36 hours, and 1 week after the rats were killed. Immunohistochemistry was used to test frozen sections with monoclonal antibodies of various inflammatory cellular markers. RESULTS: Reepithelialization was observed at 12 hours in group A, and at 24 hours in groups B and C. Regenerated epithelium covered the denuded corneal surface in groups B and C after 1 week. The expression of major histocompatibility complex II antigen was detected in infiltrating cells, corneal epithelial cells, and endothelial cells 1 hour after surgery. Only a few macrophages and Langerhans cells were in the limbus at baseline. Macrophages migrated from the limbus to the corneal ablation zone and increased 2-fold after 36 hours in all 3 groups compared with baseline. Occasional lymphocytic infiltration was identified after 25 to 36 hours. CONCLUSION: Macrophages play an active role in the wound healing after laser keratectomy and may contribute to transient corneal haze.

Anterior segment findings in AIDS patients with cytomegalovirus retinitis.

BACKGROUND: Anterior segment findings in AIDS patients presenting with cytomegalovirus (CMV) retinitis have not been specifically addressed in the American literature. METHODS: Our study evaluated 21 AIDS patients with CMV retinitis. RESULTS: Nineteen (90%) of these patients exhibited corneal endothelial deposits concurrent with CMV retinitis. The endothelial deposits were microscopic, opaque, linear flecks arranged in a reticular-like fashion. Of 42 eyes evaluated, 32 (76%) demonstrated active CMV retinitis. Corneal endothelial deposits were noted in 26 (81%) of the 32 eyes with retinitis. These corneal endothelial deposits were absent in the eyes which did not have CMV retinitis. CONCLUSION: Meticulous examination of the retina of an HIV-positive or AIDS patient who presents with reticularly arranged, linear, flecked corneal endothelial deposits should be performed to ensure that the diagnosis of CMV retinitis can be ruled out.

Increased numbers of mast cells in pterygia.

PURPOSE/METHODS: We examined the mast cells in 12 pterygium specimens of patients who underwent primary excisions and the conjunctival specimens of ten normal age-matched control subjects. RESULTS/CONCLUSION: The mean mast cell count per cubic millimeter was twice as high in the pterygium specimens as in the control specimens. Mast cell proliferation and activation may contribute to the pathogenesis of pterygium formation.

Multiple myeloma presenting with bilateral exudative macular detachments.

Bilateral exudative macular detachments were present on a 64-year-old diabetic Caucasian male who presented with bilateral blurring of vision. Besides the exudative macular detachments there was no diabetic retinopathy or congestive retinopathy, and a previous fluorescein angiogram revealed no focal leakage. Laboratory investigation, bone marrow biopsy, and a bone survey revealed the diagnosis of multiple myeloma.