muramidase and Thrombosis

The periosteum is a membrane that surrounds bones, providing essential cellular and biological components for fracture healing and bone repair. Tissue engineered scaffolds able to function as periosteum substitutes can significantly improve bone regeneration in severely injured tissues. Efforts to develop more bioactive and tunable periosteal substitutes are required to improve the success of this tissue engineering approach. In this work, a chemical modification was performed in chitosan, a polysaccharide with osteoconductive properties, by introducing phosphate groups to its structure. The phosphorylated polymer (Chp) was used to produce chitosan-xanthan-based scaffolds for periosteal tissue engineering. Porous and mechanically reinforced matrices were obtained with addition of the surfactant Kolliphor® P188 and the silicone rubber Silpuran® 2130A/B. Scaffolds properties, such as large pore sizes (850-1097 μm), micro-roughness and thickness (0.7-3.5 mm in culture medium), as well as low thrombogenicity compared to standard implantable materials, extended degradation time and negligible cytotoxicity, enable their application as periosteum substitutes. Moreover, the higher adsorption of bone morphogenetic protein mimic (cytochrome C) by Chp-based formulations suggests improved osteoinductivity of these materials, indicating that, when used in vivo, the material would be able to concentrate native BMPs and induce osteogenesis. The scaffolds produced were not toxic to adipose tissue-derived stem cells, however, cell adhesion and proliferation on the scaffolds surfaces can be still further improved. The mineralization observed on the surface of all formulations indicates that the materials studied have promising characteristics for the application in bone regeneration.

Topics: Adipose Tissue; Adsorption; Alkaline Phosphatase; Calcium; Cell Death; Cells, Cultured; Chitosan; Cytochromes c; Elastic Modulus; Humans; L-Lactate Dehydrogenase; Muramidase; Osseointegration; Osteogenesis; Periosteum; Phosphorylation; Polysaccharides, Bacterial; Porosity; Spectroscopy, Fourier Transform Infrared; Stem Cells; Stress, Mechanical; Thrombosis; Tissue Engineering; Tissue Scaffolds

Collagen-based materials are probably among the most used class of biomaterials in tissue engineering and regenerative medicine. Although collagen is often privileged for providing a suitable substrate on which cells can be cultured or a matrix in which cells can be dispersed, its mechanical properties represent a major limitation for clinical translation and even for handling of the obtained regenerated tissue. In this work, the combination of polysaccharides chitosan (Ch) and xanthan gum (X) was investigated as an alternative for scaffolds for soft tissue engineering. Moreover, in an attempt to reach a compromise between obtaining highly porous biomaterials while maintaining appropriate mechanical properties, a surfactant (Kolliphor® P188, K) was added to Ch-X matrices to generate pores, while silicone rubber (Silpuran® 2130A/B, S) was used to balance their mechanical properties. Addition of K (10 or 25% w/w) increased the porosity and pore-dimensions, while addition of S improved by up to 156% and 85% the elastic moduli and the elastic behavior of Ch-X-based scaffolds, under both compressive and tensile loads, respectively, at 50% strain. Relaxation tests confirmed that these materials do have a viscoelastic behavior. The presence of S increased thickness and microscale surface roughness and did not affect liquid uptake and stability, thrombogenicity, biodegradation and cytotoxicity of polysaccharide-based scaffolds. In conclusion, this work shows that Ch-X-S porous blends constitute suitable scaffolds for soft tissue engineering.

Topics: Cell Death; Chitosan; Elastic Modulus; Fibroblasts; Humans; Mechanical Phenomena; Muramidase; Polysaccharides; Polysaccharides, Bacterial; Porosity; Stress, Mechanical; Thrombosis; Tissue Engineering; Tissue Scaffolds; Water

Topics: Aminopeptidases; Animals; Arylsulfatases; Blood Urea Nitrogen; Calcium; Creatinine; Dogs; Enzymes; Glucuronidase; Graft Rejection; Kidney Transplantation; Muramidase; Potassium; Renal Artery Obstruction; Sodium; Surgical Wound Infection; Thrombosis; Transplantation, Homologous

Factors determining thrombus formation on a foreign surface were studied with the use of plastic flow chambers introduced into extracorporeal shunts. Silicone rubber shunts, joining the carotid artery and jugular vein, were implanted in dogs and remained patent for several weeks. The flow chamber geometry consisted of a 4.8 mm diameter straight tube having a 3.2 X 3.2 mm circumferential cavity in the wall. Chambers were introduced sequentially into the shunts for exposure times of 10 to 30 minutes and regulated blood flow rates of 100 to 400 ml/min. The dry weight of thrombus accumulated in the chamber (5 to 50 mg) was found to increase with exposure time up to 20 minutes and to decrease with increasing flow rate. Various components of the process of thrombus formation were altered by the administration of acetylsalicylic acid, heparin and lysozyme, used alone and in pairs. Heparin was found to be the most effective antithrombotic agent, dry weights of accumulated thrombus being on the order of 50 percent lower when compared to control values. The efficacy of heparin was found to be unaffected by the presence of aspirin and lysozyme, which themselves were not effective antithrombotic agents under the conditions of these experiments. The technique described here may provide a useful animal model for studying the influence of blood flow and different biomaterials on thrombus formation.

Topics: Animals; Aspirin; Blood Coagulation; Blood Flow Velocity; Disease Models, Animal; Dogs; Drug Synergism; Extracorporeal Circulation; Foreign-Body Reaction; Heparin; Muramidase; Platelet Aggregation; Rheology; Silicone Elastomers; Thrombosis; Time Factors