Injectable tissue-engineered cartilage using a fibrin glue polymer

The purpose of this study was to demonstrate the feasibility of using a fib rin glue polymer to produce injectable tissue-engineered cartilage and to d etermine the optimal fibrinogen and chondrocyte concentrations required to produce solid, homogeneous cartilage. The most favorable fibrinogen concent ration was determined by measuring the rate of degradation of fibrin glue u sing varying concentrations of purified porcine fibrinogen. The fibrinogen was mixed with thrombin (50 U/cc in 40 mM calcium chloride) to produce fibr in glue. Swine chondrocytes were then suspended in the fibrinogen before th e addition of thrombin. The chondrocyte/polymer constructs were injected in to the subcutaneous tissue of nude mice using chondrocyte concentrations of 10, 25, and 40 million chondrocytes/cc of polymer (0.4-cc injections). At 6 and 12 weeks, the neocartilage was harvested and analyzed by histology, m ass, glycosaminoglycan content, DNA content, and collagen type II content. Control groups consisted of nude mice injected with fibrin glue alone (with out chondrocytes) and a separate group injected with chondrocytes suspended in saline only (40 million cells/cc in saline; 0.4-cc injections). The fib rinogen concentration with the most favorable rate of degradation was SO mg /cc. Histologic analysis of the neocartilage showed solid, homogeneous cart ilage when using 40 million chondrocytes/cc, both at 6 and 12 weeks. The 10 and 25 million chondrocytes/cc samples showed areas of cartilage separated by areas of remnant fibrin glue. The mass of the samples ranged from 0.07 to 0.12 g at 6 weeks and decreased only slightly by week 12. The glycosamin oglycan content ranged from 2.3 to 9.4 percent for all samples; normal cart ilage controls had a content of 7.0 percent. DNA content ranged from 0.63 t o 1.4 percent for all samples, with normal pig cartilage having a mean DNA content of 0.285 percent. The samples of fibrin glue alone produced no cart ilage, and the chondrocytes alone produced neocartilage samples with a sign ificantly smaller mass (0.47 g at 6 weeks and 0.46 g at 12 weeks) when comp ared with all samples produced from chondrocytes suspended in fibrin glue ( p < 0.03). Gel electrophoreses demonstrated the presence of type II collage n in all sample groups. This study demonstrates that fibrin glue is a suita ble polymer for the formation of injectable tissue-engineered cartilage in the nude mouse model. Forty million chondrocytes per cc yielded the best qu ality cartilage at 6 and 12 weeks when analyzed by histology and content of DNA, glycosaminoglycan, and type II collagen.