Laila A. Hanninen

Morphologic effect of hyperosmolarity on rabbit corneal epithelium.

The morphologic effect of hyperosmolarity, equivalent to that seen in the tear film of patients with keratoconjunctivitis sicca (KCS), on rabbit corneal epithelium in vitro and in vivo was studied. In the in vitro studies, corneal epithelium was grown in explant cultures. Control tissue was cultured in a 307 mOsm/L medium. Epithelium cultured in the 333, 361 and 363 mOsm/L media showed decreased intercellular connections, blunting and loss of microplicae, disruptions in cell membranes and cellular swelling with decreased cytoplasmic density. In in vivo studies, corneas bathed in balanced salt solutions (BSS) concentrated to 330, 360, or 407 mOsm/L showed increased cell desquamation, and the cell changes observed at similar osmolarities in the in vitro studies. The tear film osmolarities observed in KCS are sufficient to cause the corneal epithelial changes seen in patients with this disease.

Comparative Toxicity of Intravitreal Aminoglycoside Antibiotics

We compared the toxicity of the aminoglycoside antibiotics (tobramycin, amikacin, netilmicin, and kanamycin) by ophthalmoscopy, light and electron microscopy, and electro-retinography after intravitreal injection in rabbits in doses ranging from 100 to 3,000 micrograms. The earliest manifestations of toxicity were confined to the outer retina with each drug, with lamellar lysosomal inclusions in the retinal pigment epithelium as the earliest finding. However, the aminoglycosides displayed marked differences in the threshold dose required to produce toxic reactions, permitting the following ordering of toxicity: (most toxic) gentamicin greater than netilmicin = tobramycin greater than amikacin = kanamycin (least toxic).

The influence of aphakia and vitrectomy on experimental retinal toxicity of aminoglycoside antibiotics.

To study the effect of lens and vitreous surgery on the dose threshold of aminoglycoside-induced retinal toxicity, we performed extracapsular lens extraction (Group 1) or lensectomy and vitrectomy (Group 2) on Dutch Belted rabbits. A single dose of amikacin or gentamicin ranging from 100 to 4,000 micrograms was administered intravitreally. Retinal toxicity was examined by light and transmission electron microscopy seven days after injection. Both groups showed retinal toxicity after 400 micrograms of gentamicin or 1,500 micrograms of amikacin, doses identical to those causing toxicity in intact, phakic rabbit eyes. Thus, neither surgical procedure increased the toxic threshold of injected aminoglycoside. Given the accelerated aminoglycoside clearance reported in aphakic eyes, these findings imply that the mechanism of aminoglycoside toxicity may be related to peak drug concentration rather than duration of tissue exposure.

Ocular Abnormalities in Mucolipidosis Iv

Systemic findings in a 23-year-old white man with mucolipidosis type IV included early delayed psychomotor development, mental retardation, and mild facial dysplasia. There was urinary excretion of chondroitin sulfate. Ophthalmologic examination showed corneal haze, pigmentary retinopathy, and severe optic atrophy. Light microscopy showed massively engorged superficial and intermediate epithelial cells of both the cornea and the conjunctiva. By transmission electron microscopy these contained fine granular material consistent with acid mucopolysaccharide and concentric lamellar bodies presumably representing phospholipids. This storage phenomenon was also found in macrophages, plasma cells, ciliary epithelial cells, Schwann cells, retinal ganglion cells, and vascular endothelial cells. Light microscopy also disclosed early cataract formation, marked outer retinal degeneration, and optic atrophy.

Morphologic characteristics of posterior polymorphous dystrophy. A study of nine corneas and review of the literature.

Based on their own study of nine corneas with clinically documented posterior polymorphous dystrophy and a review of the literature, the authors describe the morphologic features of this entity. Study by phase contrast light microscopy and transmission and scanning electron microscopy found that changes were primarily in the endothelium and consisted of endothelial cell degeneration and loss with focal fibroblastic and epithelial-like cell transformation. Secondary alterations of Descemets membrane were seen; they consisted of abnormal lamination with deposition of abnormal collagen material, particularly in the posterior collagen layer, and formation of guttate excrescences and pits.

Clinical and histopathologic studies of two families with lattice corneal dystrophy and familial systemic amyloidosis (Meretoja syndrome).

Lattice corneal dystrophy associated with familial systemic amyloidosis (Meretoja syndrome) has rarely been described other than in patients of Finnish origin. The authors report two North American patients with this disease who manifest blepharochalasis, lattice corneal dystrophy, open-angle glaucoma, and cranial neuropathy. In one patient, a corneal intraepithelial and subepithelial pseudodendrite was managed by superficial keratectomy, and this same patient benefited from surgical brow suspension for facial muscular weakness. In the second patient, penetrating keratoplasty was complicated by a neurotrophic persistent epithelial defect. Corneal tissue from both superficial keratectomy and penetrating keratoplasty exhibited ultrastructurally characteristic amyloid filaments and associated elastoid material. Transmission electron microscopy of conjunctiva and skin biopsies similarly revealed amyloid deposits associated with most basement membranes, the perineurium and endoneurium of most peripheral nerves, and the intima and media of arteries. By immunoperoxidase staining, the corneal amyloid deposits were positive for the amyloid P-component protein but negative for the nonimmunoglobulin amyloid A protein and prealbumin. Serum prealbumin and amyloid A related protein were normal.

Proteolytic Activation of Corneal Matrix Metalloproteinase by Pseudomonas Aeruginosa Elastase

Purified Pseudomonas aeruginosa elastase cleaved a 65 kDa gelatinase [inactive proenzyme form of matrix metalloproteinase (MMP-2)] from human corneal fibroblasts into a biologically active fragment with an approximate molecular mass of 58 kDa. However, purified pseudomonal alkaline protease did not cleave MMP-2 appreciably. Since activated MMP-2 is known to degrade native type IV, V and VII collagens, all components of the corneal basement membrane or stroma, our results suggest a new role for pseudomonal elastase in the pathogenesis of corneal infection, inflammation and ulceration.

In vitro toxicity of gentamicin to corneal epithelial cells

We report morphologic and viability changes in the rabbit corneal epithelium exposed in vitro to gentamicin sulfate. Rabbit corneal epithelial cells (3 x 10(5)) were seeded in replicate 24-well plates at their first in vitro passage, and gentamicin in concentrations of 0, 50, 250, 500, 1,000, or 5,000 micrograms/ml was added to the tissue culture medium beginning 7 days after subculture. By phase contrast microscopy, changes in cell morphologic appearance, particularly increased cytoplasmic granularity, were observed in the 5,000-micrograms/ml groups as early as 24 h after introduction of the drug. At 48 h, similar findings were observed in the 250-micrograms/ml group and at all higher concentrations. The cytoplasmic granularity was not noted in the 0- or 50-micrograms/ml groups. By electron microscopy, these observations correlated with ultrastructural findings of increased accumulations of intralysosomal bodies beginning in the 250-micrograms/ml group after 48 h of exposure to gentamicin. Furthermore, a statistically significant difference (p = 0.001) was demonstrated between the total number of viable cells in the low-dose group (50-micrograms/ml) and the high-dose groups (greater than or equal to 250-micrograms/ml) for exposure periods of 48 h or more. These findings demonstrate aminoglycoside toxicity to corneal epithelial cells in vitro similar to that seen in the human kidney and conjunctiva.

Natural history of disease in a rabbit model for keratoconjunctivitis sicca

Abstract We have continued our study of the tear film and ocular surface in our full KCS (keratoconjunctivitis sicca) rabbit model up to 52 weeks post‐operatively. Tear film osmolarity remains elevated, conjunctival goblet cell density remains decreased, and the conjunctival epithelium remains abnormal. Corneal epithelial glycogen levels decreased progressively, and at 44 weeks post‐operatively rabbits developed abnormal rose Bengal staining of the affected cornea that was shown to be associated with morphologic abnormalities at 52 weeks. Rabbits began rubbing the affected eye after the development of corneal staining. Our full KCS rabbit model demonstrates the features of the human disease.

Effects of Hypertonic Solutions on Conjunctival Epithelium and Mucinlike Glycoprotein Discharge

A simple, semiquantitative, histochemical method using periodic acid-Schiff (PAS) staining was developed to measure the mucinlike glycoprotein content of bathing solutions exposed to rabbit conjunctiva. Rabbit eyes were bathed in vivo in solutions of various tonicity (160, 260, 300, 330, and 363 mOsm/L). Eyes bathed in hypotonic or isotonic solutions (160, 260, and 300 mOsm/L) exhibited a steady-state glycoprotein secretion (50–100 μg/h). In hypertonic solutions (330 and 363 mOsm/L), glycoprotein discharge increased to 150–360 μg/h when the solutions were exchanged hourly for 6 h, but glycoprotein discharge increased to a much lower rate (100–167 μg/h) after prolonged bathing for 6 h without solution exchange. Periodic acid-Schiff-positive glycoprotein aggregates were noted more frequently on the filters of hypertonic solutions than on those of other tonicity. After eyes were bathed in hypertonic solutions, increased disruption of the surface epithelium, increased abnormal discharge of mucin granules, and decreased goblet cell density were noted in the biopsied conjunctiva. These data suggest that mucinlike glycoprotein secretion from the conjunctival goblet cells can be enhanced by the tonicity of the bathing solution, and that nonphysiological hypertonic solutions can also affect the conjunctival epithelium.