Endre A. Balazs

The Use of Sodium Hyaluronate (Healon®) in Human Anterior Segment Surgey

The safety and efficacy of sodium hyaluronate (Healon) was evaluated in a wide spectrum of anterior segment surgical procedures. Healon is not only safe, but actually facilitates the outcome of surgery. Healon placed intracamerally before cataract removal results in significantly decreased endothelial cell loss. In filtering procedures, use of intracameral and subconjunctival Healon promotes superior bleb formation while still maintaining chamber depth postoperatively. Corneal grafts performed over intracameral Healon receive maximal endothelial protection and manifest striking postoperative clarity.

Tapping Mode Atomic Force Microscopy of Hyaluronan: Extended and Intramolecularly Interacting Chains

The extracellular matrix polysaccharide hyaluronan has been examined by tapping mode atomic force microscopy. High molecular weight hyaluronan was deposited on mica from dilute aqueous solution and imaged in air. Long unbranched chains could be observed and were found to be compatible with the known covalent structure of hyaluronan. In addition, chains with evidence of intramolecular association were observed. In the simplest cases, the association took the form of loops stabilized by antiparallel double-stranded (probably double-helical) segments. In other cases, the polarity of the associated regions could not be determined. Extensive intramolecular association in long hyaluronan chains resulted in a fenestrated structure of the same type as that formed by intermolecular association at higher concentrations.

1H NMR of glycosaminoglycans and hyaluronic acid oligosaccharides in aqueous solution: The amide proton environment☆

The exchangeable amide protons of hyaluronic acid (HA) oligosaccharides and a higher-molecular-weight segment dissolved in H2O at pH 2.5 or 5.5 were examined by H NMR spectroscopy at 250 MHz. The HA segment preparation showed a single amide resonance, near the chemical shift for the amide proton of the monosaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose (beta-GlcNAc). Smaller HA oligosaccharides showed two or three separate amide proton resonances, corresponding in relative peak area to interior or end GlcNAc residues. The interior GlcNAc amide resonance occurred at the same chemical shift as the single resonance of the HA segment. For the end GlcNAc residues, linkage to D-glucuronopyranose (GlcUA) through C1 resulted in an upfield shift relative to the beta-anomer of GlcNAc, whereas linkage through C3 resulted in a downfield shift relative to the corresponding anomer of GlcNAc. These chemical-shift perturbations appeared to be approximately offsetting in the case of linkage at both positions. The amide proton vicinal coupling constant (ca. 9 Hz) was found to be essentially independent of chain length, residue position, or solution pH. These data favor a nearly perpendicular orientation for the acetamido group with respect to the sugar ring, little affected by linkage of GlcNAc to GlcUA. No evidence for the existence of a stable hydrogen bond linking the amide proton with the carboxyl(ate) oxygen of the adjacent uronic acid residue was found. The amide proton resonances for chondroitin, chondroitin 4-sulfate, and dermatan sulfate were compared to that of HA. The chemical shifts of these resonances deviated no more than 0.1 ppm from that of HA. A small dependence on the identity of the adjacent uronic acid residue was noted, based on the observation of two resonances for dermatan sulfate.

Age-related changes in the vitreus and lens of rhesus monkeys (Macaca mulatta)☆

Biomicroscopic observations of the lens and vitreus, and biochemical analyses of vitreus and aqueous humor, were carried out on free-ranging rhesus monkeys (Macaca mulatta) ranging in age from 1 to 19 years. No senile cataracts were observed. After an initial decrease, the thickness of the lens increased in animals above 4 years of age, concomitant with a decrease in the depth of the anterior chamber. Three patterns of branching of the hyaloid arteriole and the capsulo-pupillary vascular system on the posterior surface of the lens were found. Six stages of vitreus patterns are described showing development of vitreous tracts and liquid pockets and corresponding closely to similar stages observed in human vitreus. The concentration and the molecular size of Na-hyaluronate and the protein concentration in the vitreus showed no significant change with age. The amount of liquid vitreus which could be collected showed a significant increase with age. Biochemical analysis of the aqueous humor revealed no significant differences in Na-hyaluronate or protein concentration in young and old adults.

Exogenous Na-hyaluronate in the anterior chamber of the owl monkey and its effect on the intraocular pressure.

Exogenous, ultrapure (sterile, pyrogen-free), non-inflammatory fraction of Na-hyaluronate (NIF-NaHA) was introduced into the anterior chamber of owl monkeys (Aotus trivirgatus), replacing approximately 48% or 77% of the aqueous humor and creating post-injection intraocular pressures (IOPs) below normal (5-10 mmHg) or above normal (40-60 mmHg), respectively. Five different molecular weight samples (MW 1.7, 3.4, 3.7, 4.5 and 4.9 X 10(6)) were used. All solutions contained 1% NIF-NaHA and, because of the varying molecular weights, the viscosities of the solutions ranged between 10 000 and 930 000 cSt. The IOP and the rate of export of the exogenous NIF-NaHA from the anterior chamber were measured. All solutions caused an increase in the IOP, and the maximum level occurred at 4 hr after injection. In all cases, the IOP returned to normal 24 hr after injection. The highest and most persistent increase in IOP was observed after the injection of the solution with the lowest viscosity (10 000 cSt). The smallest increases in IOP over the post-operative value were observed after replacement of the aqueous humor using those samples with viscosities of 10 0000 to 300 000 cSt. The turnover (export rate) of injected NIF-NaHA depends for the most part on the viscosity of the injected solution. With increasing viscosity the rate constant, and therefore the half-life, of the injected NIF-NaHA decreases. The volume fraction of the viscous solution replacing the aqueous humor is also a determining factor in establishing the turnover rate. The molecular weight of the injected NIF-NaHA did not change during that time (48 hr) in which a sufficient amount of sample for analysis could be obtained. No evidence was found for the presence of any kind of hyaluronic acid-degrading agent in the anterior chamber.

Retinal degeneration in rhesus monkeys, Macaca mulatta. Survey of three seminatural free-breeding colonies

The ophthalmoscopic investigation of 162 seminatural, free-breeding rhesus monkeys is reported. The age (ranging from 1 month to 21 years), the maternal lineage and sibling relationships of these monkeys were known. Age-related changes in the fundus appearance, macula, retinal vessels, remnants of the hyaloid arteriole and choroidal patterns are described. Two major fundus pathologies were observed: a macular pigmentary anomaly and paramacular drusen-like lesions. These appeared separately or in combination, unior bilaterally. Less frequent pigmentary anomalies were observed outside the macula. The macular pigmentary anomalies and drusen-like lesions affect both sexes (male: 44%; female: 56%). Both pathologies were observed in fairly severe form in very young animals (1 to 3 years), but the frequency increases with age. In the old adult group (16–21 years) one or both eyes of 83% of the animals were affected. Since the same pathology with a some-what lower frequency was also found in a genetically unrelated group of (12+-year-old) monkeys living in cages in a completely different environment, and in a genetically-related group living on a different diet, environmental and nutritional causes in etiology were excluded. The hereditary nature of the disease is highly probable, but the mode of inheritance could not be determined from the data available. The fundus pathology described resembles in many respects some human hereditary retinal degenerations, and therefore it is hoped that further studies of this disease will shed some light on the etiology and pathophysiology of pigmentary and drusen-like disorders in humans.

The fine structure of the hyaloid arteriole in bovine vitreous

Abstract The purpose of this study is the descriptive analysis of the morphology of the bovine fetal and neonatal hyaloid arteriole as seen with the transmission electron microscope. Gross and minute details of intra- and extracellular formations in the wall of this arteriole are noted during its developing and atrophying stages. During the last period of the first trimester and through mid-gestation the endothelial cells and the primitive myocytes have the ultrastructural characteristics of actively seretory cells. It is presumed that they are involved in the synthesis of the elaborate lamellar systems of the extracellular matrix. Observations are made on the appearance and the staining characteristics of the elaborate multi-layered basal lamina-like membranes located between the intima and muscularis and between the several muscle cell layers. The composition and eventual fate of the non-degenerated portion of the hyaloid arteriole is discussed and its relation to the wall of Cloquets canal is postulated.

Temperature effect on the dynamic rheological characteristics of hyaluronan, hylan A, and synvisc

ABSTRACT This study examines the effect of temperatures in the range 25-65 °C on the dynamic rheological behavior of aqueous salt solutions containing hyaluronan, hylan A or Synvisc. Increasing temperature significantly reduced the modulus and complex viscosity for all three samples. This change is qualitatively understood to result from the increasing population of higher energy conformations as the temperature is raised, resulting in a decrease in the persistence length of the polymer chain. The data were found to be compatible with the viscosities predicted using a general relation proposed by Matsuoka and Cowman for polymer solutions.

Isolation and Characterization of Ichthyosan from Tuna Vitreous

Ichthyosan has been prepared from tuna vitreous. Glucuronic acid was found to account for the total uronic acid content of the macromolecule, while the hexosamine content was a mixture of N-acetyl-glucosamine and N-acetyl-galactosamine. When ichthyosan was gel filtered on Sepharose 2B or Sephacryl S-300, using sodium or calcium chloride, the elution profile of the column gave only one peak indicating no separation between glucosamine and galactosamine containing fractions. Similar results were obtained when ichthyosan was chromatographed on DEAE-cellulose using a salt gradient both in the presence and absence of 7.0 M urea. When ichthyosan was gel filtered in 4.0 M guanidine-HCL and subsequently chromatographed on DEAE-Sephacel or DEAE-cellulose, three well separated fractions were present. The two major fractions (II and III) were characterized as chondroitin and hyaluronic acid respectively; while fraction I representing about 3-5% of the total polysaccharide content of ichthyosan was identified as a keratan-like molecule. The same pattern was obtained when ichthyosan was digested with proteolytic enzymes and subsequently chromatographed on DEAE-cellulose or DEAE-Sephacel. Based on these findings it is concluded that in ichthyosan chondroitin, hyaluronate and keratan-like molecular chains are bound to proteins in non-covalent linkages.

Photoparamagnetism in solid nucleic acid-cationic dye complexes.

USING electron paramagnetic resonance (EPR) techniques, we have demonstrated that when complexes between cationic dyes and the naturally occurring anionic polysaccharides (heparin and hyaluronic acid) or the synthetic polyanion sodium poly(ethylene sulphonate) are exposed to light, the complexes become paramagnetic. The EPR spectra showed no hyperfine splitting and the intensity of the signal was not temperature-dependent. The action of the light promotes electron transfer within the complex1. We report here that the solid complexes between DNA, RNA and a range of cationic dyes exhibit a similar type of behaviour.