Molecular characteristics of some commercial high-molecular-weight hyaluronans.

Commercially available hyaluronan (HA) samples were investigated by the method of size exclusion chromatography (SEC). The fractions eluted from the SEC column were on-line molecularly characterized by using a multi-angle laser light scattering (MALLS) photometer. Along with the SEC-MALLS technique, the high-molecular-weight HA biopolymers were (off-line) analyzed by capillary viscometry.

In vitro cytotoxicity testing of coladerm membrane.

At present, biodegradable and biocompatible membranes based on collagen and glycosaminoglycans play an important role in substitutive medicine. Modern biomaterials use a chemically modified collagen-based matrix for implants with programmable biodegradability as a substitute of buccal mucosa, skin, cartilage, etc. Besides the requirements for biocompatibility and biodegradability, the membranes must be also non-toxic. Therefore, cytotoxicity testing of these materials in vitro is an integral part of introducing newly developed types of membranes into clinical practice. As a biological model for the tested COLADERM membrane, cell cultures from human embryonic fibroblasts (B-HEF-2) were used for both cytotoxicity testing as well as in tests to assess the ability of cells to proliferate on this membrane. Along with the ability of cells to grow on the surface and inside the membrane, immunohistochemical examination and scanning electron microscopy (SEM) were performed as well. The obtained results have shown that the COLADERM membrane is non-toxic with suitable structural and biological properties for clinical application as a substitute of buccal mucosa following surgical ablation of malignant tissues from the oral cavity.

Biocompatibility studies of modified collagen/Hyaluronan membranes after implantation.

Two varieties of collagen/sodium hyaluronan membranes were used as dermal substitutes in a biocompatibility implantation study on rats. In order to improve especially the physical and mechanical properties of the material, the membranes were chemically modified using a combination of hexamethylenediisocyanate (HMDC) as a crosslinker and polyoxy-ethylene (POE) as a spacer. According to both macroscopic and microscopic histological observations, the membranes were well accepted by the surrounding host tissue in all the animals. No major differences in relation to the outgrowth of the material by host tissue have been observed between the implant varieties A and B. The most important finding was that no pathological changes or important alterations of the host tissues were detected.

Aggregation Properties of Sodium Hyaluronate with Alkanediyl-alpha,omega-bis(dimethylalkylammonium Bromide) Surfactants in Aqueous Sodium Chloride Solution.

Aggregation properties of sodium hyaluronate (NaHA) with alkanediyl-alpha,omega-bis(dimethylalkylammonium bromide) surfactants (referred to as dimeric surfactants) in aqueous sodium chloride solutions have been studied as a function of surfactant chemical structure. Surface tension measurements indicate the unusual parabolic dependence of surface tension vs log surfactant concentration with a surface tension minimum at concentration c(min). The increase of surface tension above c(min) may be related to the formation of clusters consisting of NaHA chain and dimeric surfactants at the air-water interface and in the bulk. From light scattering measurements, molecular weight, hydrodynamic radius, and second virial coefficient have been calculated. The simple calculation of the ratio of positive charge of dimeric surfactant unit per one negatively charged hyaluronate disaccharidic unit in NaHA-surfactant complex reveals that there is a slight excess of positive surfactant charges per one negatively charged disaccharidic unit in the region around c(min) and the NaHA-surfactant complex is not far from electroneutrality. The nonlinear behavior of viscosity vs surfactant concentration in the NaHA-dimeric surfactant system depends on surfactant chemical structure. The behavior is concerned with the size increase due to complex growth and with the size shrinkage above c(min). A model describing the behavior of NaHA-surfactant complex in the bulk and at the interface is suggested. Copyright 2000 Academic Press.

Biocompatibility Studies of a New Biosynthetic Dermal Substitute Based on Collagen/Hyaluronan Conjugate.

The use of glycosaminoglycans (GAGs) as the second macromolecular component with collagen for the membrane preparation is based on the idea to create a biodegradable scaffold for new tissue. A hybrid collagen/hyaluronan membrane with specific bubble macrostructure was designed to serve as a synthetic dermal substitute. The objectives were to explain the two questions: what is the local biological response to the implanted membrane, and what is its reconstruction rate in tissue. The histological study proved low irritability, good compatibility, ingrowth of autologous tissue starting on day 7 post implantation, and resorption within four weeks. The major part of the implant was replaced by autologous tissue at the end of three weeks post implantation. The overall local biological response to implant was very good.

Resorption and calcification of chemically modified collagen/hyaluronan hybrid membranes.

The objectives of this study were to characterize collagen/hyaluronan hybrid membrane before and after different chemical crosslinking, as well as to investigate the impact of the treatment method on resorption and calcification processes by in vivo tests. The kinetics study of resorption of collagen/hyaluronan membranes showed that after 28 days the membranes were completely resorbed. The modification of the membranes with glutaraldehyde comparing to the similar modification of pericardial tissue conducts higher calcification. Treatment with hexamethylene diisocyanate together with poly(oxyethylene) improved mechanical properties but the resorption process was similar. Additional treatments with diphenylphosphorylazide and 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide (EDAC) resulted in prolongation of the resorption time. Calcification was suppressed in case of treatment with EDAC.

[Results of application of hydroxyapatite and hydroxyapatitecollagen.]. / Výsledky pouzitia hydroxyapatitu a hydroxyapatitkolagénu na výpln kostných defektov.

The authors present their experience of the application of hydroxyapatite and a composite-hydroxyapatitecollagen in 2 groups of patients. In the first group they evaluate the mediumterm results in 10 girls and 8 boys of average age of 12,5 years after the filling of bone defects in case of bone cysts, aneurysmatic cysts, fibrous cortical defect and eosinophilous granuloma. Hydroxyapatite was applied in the form of rolls, small bricks, strips. The first signs of healing were evident as early as after 4 months, after 36 months osteointegration was practically finished. In the second group of 1995-2000 period they applied hydroxyapatite in 15 patients of average age of 45 years. In this group they applied the mentioned biomaterial also in 6 patients with instabilities and primary and secondary (metastasis) bone tumours on the spine where they also achieved after a shorter follow-up of 6-8 months good osteoinductive and conductive qualities of the biomaterial. The authors point out the two biggest contributions in this respect: filling of bone cavities with an unusually big amount of hydroxyapatite (up to 70 g) and good, even though only short-term results after resection and filling of malignant bone tumours. Key words: hydroxyapatite, collagen, osteoinduction, osteoconduction, bone defects.

Hydroxyapatite-collagen-hyaluronic acid composite.

The new composite biomaterial made from hydroxyapatite and collagen conjugated with hyaluronic acid has been studied. The structure evaluation of the composite showed more dense arrangement due to the formation of collagen hyaluronic acid conjugate, and particles of inorganic component are closely anchored in the structure. Mechanical properties and model swelling of this composite are compared with properties of hydroxyapatite-collagen composite of similar material without hyaluronic acid. Higher cohesivity of the new biomaterial can give new possibilities for application as bone implant material. The test of contact cytotoxicity showed a very good biocompatibility of the biomaterial.

[Use of membranes based on collagen as tissue substitutes]. / Vyuzitie membrán na báze kolagénu v substitucnej medicíne.

Collagen, as biocompatible and bioactive non-specific polymer, occupies prominent position in the field of tissue engineering. The advanced biomaterials prefer collagen as a matrix for biodegradable implants as are biosynthetic skin substitutes, tendon and ligament substitutes, but also cartilage. Many studies notice the application of chemically modified collagen membranes for hernia and anastomoses reparations, vascular prosthetic materials, or for collagen tubes used in regeneration of peripheral nerves. Besides of defining main problems of tissue engineering, the paper analyses the developmental tendency of these biomaterials in the world, and compares it with the situation in Slovak Republic.

Properties of collagen and hyaluronic acid composite materials and their modification by chemical crosslinking.

This article describes properties of composites of collagen-hyaluronic acid shaped to layered materials. According to the results, interactions of these two polymers are very strong. The properties can be influenced by chemical crosslinking using glyoxal and starch dialdehyde. The different behavior during enzymatic degradation by collagenase and in swelling experiments is discussed in relation to material composition. The valuable properties of the composites observed in this study show the possibility of their use as biomaterials.

Modification of layered atelocollagen: enzymatic degradation and cytotoxicity evaluation.

Two kinds of layered atelocollagen materials cross-linked with hexamethylene diisocyanate (HMDIC), starch dialdehyde and glyoxal were enzymatically treated by bacterial collagenase. Evaluating collagenase digestion assay for these material showed progressive differences, particularly in the group of samples cross-linked with HMDIC. This should offer the possibility of programmed enzymatic degradation. These materials may be toxicologically acceptable as proven by the short-term test used for cytotoxicity evaluation.

[Atelocollagen/hydroxylapatite composite material as bone defect filler in an experiment on rats]. / Kompozit atelokolagén-hydroxylapatit ako výpln kostných defektov v pokuse na potkanoch.

Many types of biomaterials are used as skeletal bone fillers in reconstructive surgery. An attention is paid to hydroxyapatite due to its high biocompatibility with the surrounding tissue. The paper deals with the testing of new collagen/hydroxyapatite composite material applied to the bone defect on os parietale of rats. The composite material was prepared from the bovine atelocollagen dispersion and the dispersion of hydroxyapatite. Collagen serves as a matrix in which the particles of hydroxyapatite are anchored. The composite presents the advantage that after the saturation with physiological solution it is compact and can be shaped. The composite material was implanted in the form of plate into six male Wistar rats to the ground bed on the surface of os parietale. The implants were taken out after four moths. The macroscopic finding of soft tissue, bones and implants gave evidence about good healing without any undesirable reaction. This was also confirmed by the histological observations. Collagen was resorbed and the rest of the material strongly adhered to the bone. The marked osteocytes were present in the zone of the newly formed bone and the dividing line between new and old bone was clear. The experimental results give preconditions to the clinical use of this new composite implant material the structural improvement of which is in progress. (Fig. 2, Ref. 12.)

Modification of layered atelocollagen by ultraviolet irradiation and chemical cross-linking: structure stability and mechanical properties.

Physical and chemical modifications of atelocollagen materials have been carried out to improve their physical, chemical and biological properties. We have studied the influence of physical modification using ultraviolet irradiation and of chemical modification using hexamethylene diisocyanate on the physical properties of layered atelocollagen materials. The study evaluated the structural stability of cross-linked materials using swelling experiments. Influence on mechanical properties is also discussed.

Depolymerization reactions of hyaluronic acid in solution.

Samples of microbial sodium hyaluronate were degraded by heating, ultrasonication ultraviolet (UV) and gamma-ray irradiation and enzymatic treatment. The weight-average molecular weight, Mw, of hyaluronate in 0.15 M NaCl and 0.06 M Na2HPO4 was determined by gel filtration with UV detection. The Mw of the degraded samples varied from 8 x 10(4) to 1.38 x 10(6). Depolymerization processes can be described by linear relationship (1/Mw)2 = f(t) in the case of ultrasonic treatment and by non-linear relationships in the cases of heating and UV irradiation at 257 nm. Gamma-ray irradiation and enzymatic treatment caused chemical degradation and depolymerization to oligosaccharides, respectively.

[Biomaterials based on collagen]. / Biomateriíaly na báze kolagénu.

The review deals with collagen which is an essential natural polymer for preparing of biomaterials used in different fields of medicine. Besides discussing the structure of the basic polymer, the antigenicity of collagen materials as well as the bioactive properties of the collagen implants are described. The substantial part of the paper is oriented to the bioengineering processes of the preparation of the collagen matrix and composite structures. For these the physical and chemical modification of collagen are used. The processes of the modification are shown to be important in the fixing of the structure, improving the mechanical properties and influencing and directing the resorption of collagen materials in organism. Two types of materials are subdued to a detailed discussion. One which is combined with glycosoaminoglycanes and serves as the temporary skin cover and the second type, the structure of which, in the combination with hydroxylapatite imitates bone. (Tab. 4, Ref. 22.)