In this work, a systematic study of the viscoelastic properties of hydrogels based on polyethylene glycol diacrylate (PEG-DA) is presented. In addition to artificial PEG-DA-based hydrogels, natural hydrogels in the form of human articular cartilage were examined. Specimens were (unconfined) compression tested under static and dynamic load. Besides this, instrumented indentation tests with different indenter geometries (cylindrical, spherical) and load ranges (macro-and nano-indentation) were carried out and relaxation tests for the determination of moduli and relaxation time were performed. Tensile tests completed the list of measurement techniques. The measured initial moduli of the evaluated hydrogels range from 10 4-10 7 Pa. Spherical indentation was used in testing human articular cartilage in phosphate buffered saline (PBS). Cartilage samples were measured shortly after explantation, being stored at room temperature. The influence of freezing and shock-freezing was evaluated. It turned out that freezing has a massive impact on sample properties, especially on the stress relaxation time and the ratio of initial to equilibrium modulus.
Cartilage regeneration methods have been examined in various animal models. The major limitation of those studies is the biological difference between human and animal cartilage. We propose an in vivo model for human chondrocytes in a human cartilage defect environment. Human full-thickness (2-4 mm) articular cartilage discs (diameter 10 mm) attached to 3-6 mm subchondral bone, were obtained from human femur heads. Chondral defects (diameter 4 mm) were set within the cartilage disc without violating the subchondral bone. Human chondrocytes were isolated, cultivated for three passages and then suspended at a concentration of 10(7) cells/ml. The defect was completely filled with the cell suspension (approximately 30 microl) and then covered with a thin sheet of human periosteum, fixed with fibrin sealant. Discs were implanted subcutaneously in the backs of nude mice for 5 and 8 weeks. Controls were uncovered discs filled with cell suspension and covered discs without cells. Histological evaluation revealed a gradient of differentiation from the cartilage lateral side to the centre of the defect. A proteoglycan-rich matrix was formed with some chondron-like structures at the border of native cartilage, whereas fibrous tissue was built in the centre of the defect. After 8 weeks the areas of differentiating cells enlarged compared to 5 weeks, indicating the progress of cartilage repair. The control discs without cells or cover showed no chondrogenesis. Interestingly, uncovered discs filled with cells showed comparable areas of differentiating cells at the defect surface but lack of fibrous tissue in the middle. The histological results were supported by MRI measurement.
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