Arvid Anseth

Fractionation and quantitative determination of keratan sulfate using cetylpyridinium chloride and ECTEOLA - cellulose

Abstract 1. 1.The cetylpyridinium chloride-cellulose method does not permit an accurate determination of keratan sulfate in tissue digest, due to the fact that both keratan sulfate and glycopeptides are soluble in excess cetylpyridinium chloride. 2. 2. For this reason a modification of this technique has been developed. By extraction with iso-amyl alcohol cetylpyridinium chloride is removed from an aqueous solution of keratan sulfate and glycopeptides. The water phase is subsequently subjected to fractionation on ECTEOLA-cellulose to yield one glycopeptide and one keratan sulfate fraction. 3. 3. The combined cetylpyridinium chloride and ECTEOLA procedures have been applied to the estimation of keratan sulfate in artificial mixtures as well as in tissue digests.

Studies on corneal polysaccharides *: VI. Isolation of dermatan sulfate from corneal scar tissue

Previous studies have indicated that during corneal wound healing a galactosaminoglycan accumulates in the wound area. The nature of this polymer was investigated in the present study. Corneal scars were experimentally induced by removal of the endothelium and Descemets membrane. The material isolated from the scar tissue exhibited analytical data indicating that it was a dermatan sulfate with a rather high degree of hybridity. It was also observed that the polymer was more hybridized in the early phase of scar tissue formation. It was concluded that, in the process of transformation of stromal cells to dermatan sulfate-producing cells, injury to Descemets membrane leading to impaired endothelial regeneration might play an important role.

Structure of dermatan sulfate. VI. The use of cetylpyridinium chloride-cellulose microcolumns for determination of the hybrid structure of dermatan sulfates

Abstract A variety of dermatan sulfate samples have been analyzed by the cetylpyridinium chloride-cellulose microcolumn technique before and after treatment elution with testicular hyaluronidase. It was found that “neutral magnesium chloride elution profiles” reflected the molecular-size polydispersity of the material obtained after hyaluronidase degradation. However, “acid magnesium chloride elution profiles” were not influenced by the molecular size to any large extent; instead, the proportions of l -iduronic and d -glucuronic acid as well as the sulfate content seemed to be the principal parameters involved in these elution profiles. Barium acetate-ethanol cellulose elution profiles, finally, were found to separate various dermatan sulfate samples almost entirely according to uronic acid composition. Thus, by comparing elution profiles of unknown samples to those of well-characterized dermatan sulfate polymers, sufficient data were obtained to permit a formulation of the general hybrid properties of beef-aorta and sclera dermatan sulfate as well as of the dermatan sulfate obtained from human knee-joint capsule.

Studies on corneal polysaccharides. V. Changes in corneal glycosaminoglycans in transient stromal edema.

The glycosaminoglycan content of corneal stroma during transient edema produced by removal of the endothelium was measured. It was observed that this process is associated with a considerable loss of polysaccharides. Both the keratan sulfate and the chondroitin 4-sulfate content were decreased to approximately 50% of their normal values. After healing the tissue assumed a normal appearance and the polysaccharide content returned to its original level. During this process no qualitative changes in the glycosaminoglycan pattern were observed, nor did any other polysaccharide species appear.

Studies on corneal polysaccharides: VIII. Changes in the glycosaminoglycans in some human corneal disorders*

The glycosaminoglycan pattern in human corneas afflicted with diseases was measured. In maculae corneae, keratitis and lattice degeneration a decrease in the polysaccharide content was observed. In the two latter conditions the presence of dermatan sulfate was demonstrated. The glycosaminoglycan pattern in keratoconus was normal.

Studies on corneal polysaccharides: IV. Chromatography of corneal glycosaminoglycans on ECTEOLA cellulose using formate buffers as eluting solvents

A new ECTEOLA cellulose ion exchange chromatographic procedure for the fractionation of glycosaminoglycans has been applied to corneal polysaccharides. About 80% of the total keratan sulfate was recovered in a pure state; this keratan sulfate was found to contain more than one sulfate per hexosamine moiety. The remaining 20% of the keratan sulfate contained less than one sulfate per hexosamine moiety and appeared in the chromatogram together with the bulk of the galactosaminoglycans, which were found to be chondroitin 4-sulfates containing 0·22–0·41 sulfate per hexosamine moiety.

Studies on corneal polysaccharides: VII. Changes in the glycosaminoglycans in penetrating corneal grafts

The glycosaminoglycan pattern in penetrating corneal grafts in successful and unsuccessful keratoplasties was investigated in the present study. During the early period of healing a decrease in the polysaccharide content was noticed in both conditions. After 3 months of healing transparent grafts showed a normalization of their polysaccharide pattern, whereas in the nontransparent grafts a further decrease, especially of the keratan sulfate, was observed. The nontransparent grafts also contained dermatan sulfate.

SURGICAL TREATMENT OF CORNEAL DISORDERS

During the last few years the treatment of corneal disorders has greatly improved; the principles have been stabilized and now seem to have been generally agreed upon. The technique for keratoplasty has been improved, the indications widened and settled. A certain tendency towards centralization has appeared, which has certainly contributed to the improved results. At the University Eye Clinic in Lund rather standardized indications and surgical routine have been followed during the last decade and a comparatively uniform clinical material is now available. The principal features of our experience in Lund are presented here, including present indications, surgical routine. and results.

Influence of corneal epithelium on the incorporation of 35SO4 into stromal glycosaminoglycans.

In vivo studies on 35SO4 incorporation into the polysaccharides of intact corneas and corneas lacking epithelium were performed. The isotope was injected into the anterior chamber and the activity measured on the isolated polysaccharide fractions at different times after administration. It was clearly demonstrated that the absence of epithelium strikingly decreased the sulfate incorporation in corneal polysaccharides.

Therapeutical keratoplasty in herpetic keratitis.

The introduction of IDU (1) as a specific local treatment for herpetic keratitis was a great improvement in our efforts to combat this most distressing eye infection. Several reports in the literature have confirmed the curative effect of IDU in cases with superficial engagement of the cornea. Most writers also seem to agree that IDU is less, or not at all, effective when the deeper parts of the corneal stroma are involved. The prolonged treatment which is necessary in these cases can be somewhat controversial as it has been demonstrated that IDU delays healing of corneal wounds (2). A combined local application of IDU and steroids has been proposed in advanced cases of herpetic keratitis (3 , 4), and success is sometimes achieved with this treatment. In spite of all the variations in the non-surgical therapy of herpetic keratitis tried in different parts of the world, we are always left with a group of patients resistant to all treatment, and in whom keratoplasty should be tried.