体外构建组织工程骨-软骨复合组织

In vitro fabrication of tissue engineering osteochondral composites

背景:设计一体化、具有过渡结构的双层支架材料,复合软骨细胞、骨髓间充质细胞,有利于新生的骨与软骨组织之间形成良好界面。目的:模仿自然骨-软骨基质构建复合支架,以软骨细胞和骨髓间充质干细胞为种子细胞,体外观察复合组织的成软骨及成骨能力。方法:制备明胶-硫酸软骨素-透明质酸及明胶-陶瓷化骨多孔复合支架,构建自然骨-软骨基质复合支架,复合兔软骨细胞与骨髓间充质干细胞,分未成骨诱导与成骨诱导两组培养,并进行MTT、糖胺多糖含量、碱性磷酸酶活性检测,以及苏木精-伊红染色检测。结果与结论:未成骨诱导与成骨诱导两组骨髓间充质干细胞增殖及糖胺多糖含量差异无显著性意义。未成骨诱导组碱性磷酸酶活性缓慢上升,成骨诱导组诱导后碱性磷酸酶活性迅速上升,14d时达到稳定状态。两组苏木精-伊红染色结果无明显区别,均已形成含有双层组织的类似骨-软骨样组织,其间可见未降解支架形态,但由于基质形成不完善及支架未完全降解,此种结构不成熟,细胞分布不均匀,支架内部可见散在无细胞区域。证实采用两种细胞与双层结构的支架经体外分层复合能够形成组织工程骨软骨复合组织。

BACKGROUND： To design an integrated and biphasic scaffold that is cocultured with chondrocytes and bone marrow mesenchymal cells is beneficial to form good interface between bone and cartilage tissue. OBJECTIVE： To construct a bilayered scaffold based on imitating natural osteochondral matrix that is cocultured with chondrocyte and bone marrow mesenchymal stem cells as seed cells so as to observe the chondrogenic and osteogenic capacity of the composite tissue. METHODS： Gelatin-chondroitin sulfate-hyaluronic acid and gelatin-ceramic bone porous scaffolds were prepared to be cocultured with chondrocytes and bone marrow mesenchymal stem cells for osteogenic induction or non-osteogenic induction. MTT, glycosaminoglycan, alkaline phosphatase and hematoxylin-eosin staining were detected. RESULTS AND CONCLUSION： MTT results showed the bone marrow mesenchymal stem cell proliferation and glycosaminoglycan contents had no statistical difference between the osteogenic induction and non-osteogenic induction groups. Alkaline phospbatase activity increased gradually in the non-osteogenic induction group, but increased rapidly in the osteogenic induction group that became stable at 14 days. No significant difference was found in the results of hematoxylin-eosin staining between the two groups. Osteochondral-like tissues were found in the two groups with non-degradable scaffolds. Cell-free regions were scattered within the scaffolds owing to the incomplete matrix formation and scaffold degradation. This study proved that these two kinds of seed cells cocultured with the bilayer scaffold in vitro can form tissue engineering osteochondral composite tissues.