Injectable tissue-engineered cartilage using a fibrin glue polymer
Injectable tissue-engineered cartilage using a fibrin glue polymer
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DOI：
10.1097/00006534-199906000-00001
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发表时间：
1999-06-01
影响因子：
3.6
通讯作者：
Yaremchuk, M
中科院分区：
文献类型：
--
作者：
Silverman, RP;Passaretti, D;Yaremchuk, M
本研究的目的是证明使用纤维蛋白胶聚合物生产可注射的组织工程化软骨的可行性，并确定生产固体，均匀的软骨所需的最佳纤维蛋白原和软骨细胞浓度。通过使用不同浓度的纯化猪纤维蛋白原测量纤维蛋白胶的降解速率来确定最有利的纤维蛋白原浓度。将纤维蛋白原与凝血酶（50 U/cc，溶于40 mM氯化钙中）混合以制备纤维蛋白胶。然后将猪软骨细胞悬浮在纤维蛋白原中，然后加入凝血酶。使用软骨细胞浓度为1000万、2500万和4000万软骨细胞/cc聚合物（0.4cc注射）将软骨细胞/聚合物构建体注射到裸鼠的皮下组织中。在第6周和第12周，收获新软骨并通过组织学、质量、糖胺聚糖含量、DNA含量和II型胶原含量进行分析。对照组由仅注射纤维蛋白胶（无软骨细胞）的裸鼠和仅注射悬浮在盐水中的软骨细胞的单独组（盐水中4000万个细胞/cc; 0.4-cc注射）组成。降解率最有利的纤维蛋白原浓度为50 mg/cc。新生软骨的组织学分析显示，当使用4000万软骨细胞/cc时，在6周和12周时均显示出坚实、均匀的软骨。1000万和2500万软骨细胞/cc样本显示软骨区域被残留纤维蛋白胶区域隔开。第6周时样品的质量范围为0.07 - 0.12 g，到第12周时仅略有下降。所有样本的糖胺聚糖含量范围为2.3%至9.4%;正常软骨对照组的含量为7.0%。所有样本的DNA含量范围为0.63%至1.4%，正常猪软骨的平均DNA含量为0.285%。单独的纤维蛋白胶的样品不产生软骨，并且单独的软骨细胞产生的新软骨样品与由悬浮在纤维蛋白胶中的软骨细胞产生的所有样品相比具有显著更小的质量（在6周时为0.47g，在12周时为0.46g）（p < 0.03）。凝胶电泳显示所有样品组中均存在II型胶原。本研究表明，纤维蛋白胶是一种合适的聚合物在裸鼠模型中形成可注射的组织工程软骨。当通过组织学和DNA、糖胺聚糖和II型胶原含量分析时，每cc 4000万个软骨细胞在6周和12周时产生最佳质量的软骨。
The purpose of this study was to demonstrate the feasibility of using a fibrin glue polymer to produce injectable tissue-engineered cartilage and to determine the optimal fibrinogen and chondrocyte concentrations required to produce solid, homogeneous cartilage. The most favorable fibrinogen concentration was determined by measuring the rate of degradation of fibrin glue using varying concentrations of purified porcine fibrinogen. The fibrinogen was mixed with thrombin (50 U/cc in 40 mM calcium chloride) to produce fibrin glue. Swine chondrocytes were then suspended in the fibrinogen before the addition of thrombin. The chondrocyte/polymer constructs were injected into 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, mass, glycosaminoglycan content, DNA content, and collagen type II content. Control groups consisted of nude mice injected with fibrin glue alone (without chondrocytes) and a separate group injected with chondrocytes suspended in saline only (40 million cells/cc in saline; 0.4-cc injections). The fibrinogen concentration with the most favorable rate of degradation was SO mg/cc. Histologic analysis of the neocartilage showed solid, homogeneous cartilage 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 glycosaminoglycan content ranged from 2.3 to 9.4 percent for all samples; normal cartilage controls had a content of 7.0 percent. DNA content ranged from 0.63 to 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 cartilage, and the chondrocytes alone produced neocartilage samples with a significantly smaller mass (0.47 g at 6 weeks and 0.46 g at 12 weeks) when compared with all samples produced from chondrocytes suspended in fibrin glue (p < 0.03). Gel electrophoreses demonstrated the presence of type II collagen in all sample groups. This study demonstrates that fibrin glue is a suitable polymer for the formation of injectable tissue-engineered cartilage in the nude mouse model. Forty million chondrocytes per cc yielded the best quality cartilage at 6 and 12 weeks when analyzed by histology and content of DNA, glycosaminoglycan, and type II collagen.