Pure Hyaluronic Acid Research Articles

A barrier composed of chemically cross-linked hyaluronic acid (Incert) reduces postoperative adhesion formation

Objective: To test whether a barrier of chemically cross-linked pure hyaluronic acid reduces postoperative adhesion formation. Design: The material was evaluated in the murine uterine horn model using excision and electrocautery injuries and in animals who had amounts of material inserted into the abdomen to evaluate toxicity. Setting: Academic medical center. Subject(s): Mice. Intervention(s): Insertion of the barrier between uterine horns and into the peritoneal cavity. Main Outcome Measure(s): Adhesion formation at 14 days; the histology of the peritoneum, liver, and spleen at 42 days; and the number, differential count, morphology, and flow cytometry of peritoneal leukocytes 3 days postoperatively. Result(s): Fewer adhesions were present when excision injuries were separated by the barrier (12 of 28 sites [43%] versus 23 of 26 control sites [88%]), whereas the number of adhesions was unchanged after electrocautery injuries (14 of 26 sites [54%] versus 17 of 26 control sites [65%]). The uterine horn sites covered by the barrier were histologically indistinguishable from controls. No adverse impact on the peritoneum and peritoneal fluid leukocyte population was observed with barrier insertion. Conclusion(s): The use of a barrier composed of a chemically cross-linked hyaluronic acid derivative (Incert, Anika Therapeutics, Inc., Woburn, MA) reduced postoperative adhesion formation in this model without any adverse impact on the peritoneum and peritoneal leukocyte population. This barrier material shows promise in preventing postoperative adhesions and deserves clinical evaluation.

Rheology of Synovial Fluids and Substitute Polymers

Abstract Synovial fluids are highly viscous and viscoelastic solutions responsible for lubrication and damping in joints. Based on work with both healthy and arthrotically deteriorated human knee joint synovia, the rheology of these materials is discussed in terms of an entanglement network. The essential constituent of the dissolved complex is a hyaluronic acid of very high molecular mass (107), which is degraded in pathological cases with accompanying rheological deterioration. The network properties are calculated and compared with those of pure hyaluronic acid and with synovia-substituted polymers as guar gum and polyacrylamide. The network parameters reflect clearly the rheological properties and their changes in the case of arthrotic deterioration.

An approach for a rapid identification and characterization of hyaluronic acid produced by fermentation of Streptococcus equi

An approach for a rapid identification and characterization of hyaluronic acid (HA) produced by fermentation of Streptococcus equi has been developed. 125-I-labelled hyaluronic acid binding protein (HABP) was used for specific identification of HA in fermentation isolates recognized by HPLC chromatography followed by γ-radiation detection. Simultaneous determination of the molecular weight of HA was carried out also. Further confirmation of HA identification was performed by bovine testes hyaluronidase treatment and HPLC analysis of the products. The analysed sample was identified as almost pure HA with molecular weight approximately 0·8-1·2 × 10 6 Da, containing protein impurities (5·1%) and some unidentified low molecular weight products. Our approach can be used generally for a rapid identification and characterization of laboratory or industrially-produced HA during its isolation and purification.

Characterization of a cytosolic inhibitor of calf estrogen receptor binding to nuclei.

An inhibitory component that diminishes estrogen receptor (ER) binding to nuclei in vitro is present in cytosol prepared from calf uterus. The inhibitor is heat stable and resistant to enzymatic treatment with trypsin, chymotrypsin, proteinase K, deoxyribonuclease I, or ribonucleases A, T1, and U2. Results of chromatography on DEAE-cellulose and Sephadex G-150 indicate that the factor is a negatively charged macromolecule. Inhibitory activity is sensitive to sequential digestion with chondroitinase ABC, hyaluronidase, and heparinase. Approximately 70% of the inhibitory activity is destroyed by treatment with heparinase alone. Heparitinase destroys only 30% of this activity. Furthermore, the addition of pure hyaluronic acid or chondroitin sulfate to the ER-nuclei binding assay results in little inhibition, whereas addition of heparin inhibits 75% of receptor binding. Overall, these results indicate that glycosaminoglycans, present in bovine uterine cytosol, are capable of inhibiting ER-nuclei interactions. The most potent inhibitory glycosaminoglycan displays heparin-like characteristics.

The Splitting of Hyaluronic Acid by Gastric Juice and Saliva of Dogs

The splitting action on hyaluronic acid by gastric juice (18 cases) and saliva (3 cases) from dogs was studied. The determinations were performed viscometrically with pure hyaluronic acid as substrate. The pH-optimum of the activity was in a few cases determined to pH 3.4 and the activity of the samples were preliminary studied at that pH. It was found that gastric juice in 14 cases out of 18 and saliva in the 3 cases studied showed a splitting action on HyA.

Effects of X-ray Irradiation on Viscosity of Synovial Fluid

Conclusions1 . The viscosity of joint fluid is decreased by exposure to X-rays, the decrease in viscosity being directly proportional to the amount of irradiation. 2. The viscosity of pure hyaluronic acid solutions is decreased less by the action of X-rays than is the viscosity of joint fluid. 3. Joint fluid is as susceptible to the enzymatic action of bull testis hqaluronidase in the irradiated sample as in the control. S o increase in titratable acidity follows irradiation of pure hyaluronate solution, indicating that the reduction in viscosity probably does not result from oxidative degradation. 4. The possible physiological implications are discussed.

HYALURONIDASES OF BACTERIAL AND ANIMAL ORIGIN

Hyaluronidase has been investigated in various strains of pneumococci and hemolytic streptococci, and in some material of animal origin. The enzyme activity was measured by a viscosimetric method using as a substrate a fluid containing hyaluronic acid as the viscous component, and by the hydrolysis of pure hyaluronic acid into its reducing components. In pneumococci the enzyme was demonstrated in all types and in all strains tested, including smooth and rough forms of Types I, II, III, and VI. In hemolytic streptococci the enzyme from strain H44, group A, reported previously, was further investigated. In this strain, as well as in other hemolytic streptococci containing the enzyme, great variability of the enzyme concentration was found. Furthermore, the enzyme proved to be very labile, giving in the viscosimetric measurements a typical stoppage of the activity initially present. In 13 out of 14 other strains of group A organisms investigated, no enzyme was demonstrable, but the variation in activity in the enzyme-active strains renders the negative findings inconclusive. A very active enzyme, though of great variability, was found in one group C strain. The enzyme was prepared from the leech in confirmation of the work of Claude. The enzyme from testis showed a maximum at pH 4.4 in contrast to the optimum of 5.8 in pneumococcal, streptococcal, and Cl. welchii preparations. The pH curve of the testis enzyme indicated, however, a second optimum coinciding with that of the bacterial enzymes. The hydrolysis further indicated a break at about 50 per cent hydrolysis, indicating primarily the hydrolysis down to aldobionic acid units. The depolymerizing action of testis enzyme is more marked than that of pneumococcal enzyme. The results have been interpreted as due to the presence of two enzymes, one attacking the long chain molecule, the other hydrolyzing the aldobionic acid formed. The enzyme was further prepared from beef spleen. Here the hydrolysis of beta-glucuronides was compared to that of hyaluronic acid. The two actions apparently are catalyzed by two distinct enzymes. Enzyme preparations were further obtained from rabbit skin. Since hyaluronic acid has also been found in the skin, this organ may play a considerable rôle in the metabolism of hyaluronic acid. In addition to hyaluronic acid, it has been shown that hyaluronidases also hydrolyze the sulfuric acid containing polysaccharide of the cornea. This polysaccharide has previously been characterized as a natural sulfuric acid ester of hyaluronic acid. The pneumococcal enzyme preparations also attacked a polysaccharide acid prepared from submaxillary gland, which is not hyaluronic acid. However, it is believed that this hydrolysis is due to a second enzyme contained in the preparations. The testis enzyme, on the other hand, attacked chondroitinsulfuric acid and also contained a sulfatase. The depolymerizing action of hyaluronidase has been discussed. It is concluded that depolymerization and hydrolysis are probably due to the same enzyme attacking hyaluronic acid. It is suggested that the first attack of the enzyme does not cause an opening of glucosidic linkages. The available evidence indicates that the viscosity of the natural fluids is not due to macromolecules but to micellae formation, and that these micellae are depolymerized by the enzymatic reaction. It is assumed that the depolymerization is due to a primary enzyme-substrate reaction, which in itself is insufficient to open the glucosidic linkages. The latter reaction involves further steps. The relationship between hyaluronidase and "spreading factor" has been discussed anew. Though more data have been reported pointing to the identity of hyaluronidase and "spreading factor," our inability to demonstrate hyaluronidase in streptococcal material of high "spreading" potency, is still a serious obstacle to the unitarian theory. However, it seems possible that the streptococcal material may contain reversibly inactive enzyme which may be reactivated in vivo.