【Abstract】 Objective To increase the viscosity of chitosan/glycerol phosphate（C/GP）and to improve its preparation technique in order to develop the appl ication range of C/GP. Methods Chitosan was treated by high-pressure vapor steril ization in order to prepare high viscous C/GP（HV-C/GP）and prepare C/GP by standard methods. The rheologic changes of HV-C/GP and C/GP were detected dynamically by the Gemini rheometer. The initial solution viscosity, gelation temperature and gelation time were evaluated after the viscosity of the materials were increased. Two gelation materials were placed into continuous flow thermostated cells under the same condition and harvest them at predetermined time intervals, 1st, 2nd, 5th, 10th and 25th days, then they were dried, weighed and the mass loss rate was calculated. Ultrastructure of the freeze-dried samples was visual ized by the scanning electron microscope. Results The initial viscosity of C/GP was 1.81 Pas and that of HV-C/GP was 17.24 Pas. The latter one increased 10 times as well as the former one. The gelation temperature of C/GP was 37°C and that of HV-C/GP was 34°C. There was no remarkable difference in gelation time between them. The mass loss rate of HV-C/GP at first day was 72.5% and at 25th days was 90.8%, while that of C/GP was 55.4% and 78.2%. Porous network structure was observed by the scanning electron microscope in both of them. The pore diameter of C/GP was 50-100 μm and that of HV-C/GP was 30-50 μm, which was obviously smaller than the former. Conclusion The viscosity of HV-C/GP prepared by improved technique obviously increases and the thermosensitivity has no significant changes. The degradation time of HV-C/GP in vitro lengthens. The micrographs show that the HV-C/GP gels are porous and the pore diameter are smaller than C/GP.
Objective To evaluate the effect of implantation of the complex of high viscous chitosan/glycerol phosphate with demineral ized bone matrix (HV-C/GP-DBM) in repairing cartilage defects of rabbits. Methods HV-C/ GPDBM was prepared by compounding HV-C/GP and DBM by 2:1 (W/W). Twenty-four 34-week-old New Zealand white adult rabbits, weighing 3.5-4.5 kg, were included. A bit with the diameter of 3.5 mm was used to drill 3-cm-deep holes in both sides of femoral condyle to make cartilage defects. The complex of HV-C/GP-DBM was then injected into the right holes as the experimental group and the left ones serve as the control group. The rabbits were killed at 4, 8 and 16 weeks after theoperation, respectively. The obtained specimens were observed macroscopically, microscopically and histologically. According to the International Cartilage Repair Society Histological Scoring (ICRS), the effect of cartilage repair was assessed at 16 weeks postoperatively. Results At 4-8 weeks postoperatively, in the experimental group, the defects were filled with hyal ine cartilage-l ike tissues; the majority of chitosan degradated; and the DBM particles were partly absorbed. However, in the control group, there were small quantities of discontinuous fibrous tissues and maldistributed chondrocytes at the border and the bottom of the defects. At 16 weeks postoperatively, 6 joints in the experimental group had smooth surface, and the defects were basically repaired by hyal ine cartilage-l ike tissues. The newly-formed tissues integrated well with the surrounding area. Under the cartilage, the new bone formation was still active and some DBM particles could be seen. However, the defects in the control group were repaired by fibrous tissues. The result of histological scoring of the specimens at 16 weeks showed that a total of 6 aspects including formation of chondrocytes and integration with the surrounding cartilages were superior in the experimental group to those in the control group, and there were significant differences between the two groups (P lt; 0.05). Conclusion The biodegradable and injectable complex of HV-C/GP-DBM with good histocompatibil ity and non-toxic side effects can repair cartilage defects and is a promising biomaterial for cartilage defect repair.