Fractionation of the water-soluble proteins from calf lens

The water-insoluble proteins from calf lens have been subdivided into a urea-soluble (USL) and a urea-insoluble (UIL) part. USL can be fractionated into four components by gel filtration on Sephadex G 200 columns equilibrated in 6 m-urea. One of the fractions contains the major cytoskeletal proteins MP42 and MP55. The major fiber plasma membrane protein MP26 remains in the UIL fraction and can be isolated by preparative SDS-polyacrylamide gel electrophoresis. Peptide mapping reveals that MP26 is definitely different from the major β-crystallin component βB_p.     

Mesothelial proteins are expressed in the human cornea

The goal of our study was to determine whether proteins typical of the human mesothelial cell phenotype, such as mesothelin, HBME-1 (Hector Battifora mesothelial cell-1) protein and calbindin 2, are expressed in the human cornea, especially in endothelial cells.Cryosections and endothelial and epithelial imprints of sixteen human cadaverous corneoscleral discs were used. The presence of proteins was examined using immunohistochemistry and Western blotting, while mRNA levels were determined by qRT-PCR.A strong signal for mesothelin was present in the corneal epithelium, while less intense staining was visible in the endothelium. Similarly, higher and lower mRNA levels were detected using qRT-PCR in the corneal epithelium and endothelium, respectively. HBME-1 antibody strongly stained the corneal endothelium and stromal keratocytes. Marked positivity was present in the corneal stromal extracellular matrix, while no staining was present in the sclera. Calbindin 2 was detected using immunohistochemistry and Western blotting in the corneal epithelium, endothelium and stroma. qRT-PCR confirmed its expression in epithelial and endothelial cells.Three proteins expressed constitutively in mesothelial cells were detected in the human cornea. The possible function of mesothelin in cell-cell contact on the ocular surface is discussed. The presence of HBME-1 protein in the endothelial layer may indicate a still unknown function that could be shared with mesothelial cells of the pleura and peritoneum. The much more pronounced occurrence of calbindin 2 in the corneal epithelium compared to fewer positive endothelial cells explains the higher turnover of epithelial cells compared to the proliferatively inactive endothelium.     

Transduction of functionally active TAT fusion proteins into cornea

A technology has recently been developed that allows for the rapid transduction of full-length functionally active proteins into intact tissue through intravenous injection and into cultured cells. This technology involves the fusion of an 11 amino acid sequence of the HIV TAT protein to the protein of interest. In the current investigation, we determined whether functionally active TAT fusion proteins could be transduced into intact corneas by topical application. TAT-beta-galactosidase was purified from bacterial cells and applied in serial dilutions (12.5-250 nm) to cultured epithelial cells for 5 or 15 min. In addition, enucleated globes and excised corneas with or without a central 3-mm epithelial debridement were incubated with TAT-beta-galactosidase for 1 or 2 hr. Excised corneas were allowed to heal in organ culture. Transduction of active beta-galactosidase was detected by incubating the cells or corneas with X-gal. TAT-beta-galactosidase was transduced into nearly all cultured cells in a concentration-dependent manner. When TAT-beta-galactosidase was topically applied to intact corneas, only the most superficial layer of epithelium was highly transduced. When the superficial layer was removed with nitrocellulose, two to four layers of cells were transduced. In corneas with a central debridement, epithelial cells at the edge of the debridement were transduced as well as the stromal cells subjacent to the debridement. Active beta-galactosidase was maintained at least 1 day in organ culture. No X-gal reaction was seen in either cells or corneas not incubated with TAT-beta-galactosidase. Functionally active proteins can be efficiently transduced into corneal epithelial and stromal cells using TAT fusion protein technology. The intact epithelium provides a barrier to penetration of TAT proteins. This barrier can be overcome by disrupting the epithelium. TAT-mediated protein transduction may be extremely useful in studies of corneal wound healing and homeostasis.     

The elimination of soluble cornea proteins through direct current

By direct current it was possible to eliminate soluble proteins from the cornea. This was proven by the decrease of the albumine-concentration which was determined by RID. Treated and untreated corneas were examined by electron microscopic investigations.     

Symmetry detection and visual attention: a "binary-map" hypothesis

Recent research suggests that human symmetry-detection mechanisms cannot simultaneously compare different colors across the axis of symmetry (Nature 399 (1999) 115). In the present study, observers were required to judge symmetry in arrays composed of elements varying not only in color, but also in size, spatial frequency and orientation. In every case, response times increased with the number of different levels of a given feature. It is proposed that this increase reflects a sequential strategy whereby coarse "binary maps" are created by attentional filtering, and the symmetry of each map is then checked. Experiment 2 required observers to detect "pseudo-symmetry" (symmetry in feature values defined relative to an arbitrary featural boundary); the ease with which this task was accomplished supported the binary map hypothesis. The results suggest that (1) symmetry detection is spatially imprecise, and (2) attentional gating can operate prior to symmetry detection in the visual pathway.     

Ultrastructural study of the posterior cornea of the dogfish "Scyliorhinus canicula L"

The cornea of Elasmobranchs remains transparent and exhibits a remarkable resistance against swelling when deprived of its endothelium. Sutural fibers, which traverse the corneal stroma anteroposteriorly, may be responsible for this property. We studied the structure of the cornea of Scyliorhinus canicula L., a galeiform Elasmobranch, with special emphasis on the presence of a corneal endothelium and on its appearance during development. Observations were made with the aid of several microscopic techniques: light and specular microscopy, transmission and scanning electron microscopy. We found that the posterior corneal surface was always covered by a fibrillar material. A structural counterpart of endothelial-like cells on the posterior cornea or in the anterior chamber was never observed in adult fishes and in embryos of different stages. In contrast, in Torpedo ocellata, a torpediniform Elasmobranch, endothelial cells were seen. The significance of these findings is discussed in context with the possibility of technical artefacts and compared with other studies.     

Recurrent erosion of the cornea.

Altogether, 80 patients aged between 24 and 73 years with recurrent erosion of the cornea have been studied and compared with a control group of 200. The patients' erosions were divisible into macroform and microform types. The macroform occurred in 10%, the microform in 56%, and both types in the same patients in 31%. The macroform was more commonly related to trauma than the microform. However, many (40%) were spontaneous in origin. The most common cause of the initial trauma was a finger nail. The recurrences occurred at around the time of waking, either just before or just after. Difficulty in opening the eye occurred in 10%. There was little evidence of precipitating factors, but eye rubbing was admitted by 10% and barbiturates were implicated in 3%. The corneae were examined in the healed state, when a high incidence (59%) were found to have superficial corneal dystrophies of the fingerprint lines, bleb, and Bietti's lacunar (map-like) types. These are considered individually, particular attention being paid to the distinction between the various types of line resembling the fingerprint line. Epithelial microcysts were also a common finding (59%) and were sometimes of the Cogan type. In only 11% of patients were there no corneal signs in the healed state. The need for careful examination of the cornea by retroillumination, using both the iris and the fundus, is stressed. The control group, in contrast, showed a very low incidence of dystrophies and cysts. Treatment was given initially with either drops or ointment and no differences in healing were found. Debridement was performed in 12 eyes as an initial treatment and also in four eyes which were not healing on medical treatment. Debridement assisted healing, but did not prevent recurrence. One eye was treated with debridement and scarification and seven with carbolization. These procedures appeared to reduce the recurrence rate. Sodium chloride ointment 5% was found useful as a prophylactic taken at bedtime, and the recurrence rate increased when it was withdrawn.