Sun Liguo, Li Hongguo, Qu Ling, Zhu Rui, Fan Xiangli, Xue Yingsen, Xie Zhenghong, Fan Hongbin
Institute of Orthopedic Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an 710032, China.
Department of Clinical Laboratory, Xijing Hospital, The Fourth Military Medical University, Xi'an 710032, China.
Biomed Res Int. 2014;2014:816979. doi: 10.1155/2014/816979. Epub 2014 Jun 12.
The lack of a fibrocartilage layer between graft and bone remains the leading cause of graft failure after anterior cruciate ligament (ACL) reconstruction. The objective of this study was to develop a gene-modified silk cable-reinforced chondroitin sulfate-hyaluronate acid-silk fibroin (CHS) hybrid scaffold for reconstructing the fibrocartilage layer. The scaffold was fabricated by lyophilizing the CHS mixture with braided silk cables. The scanning electronic microscopy (SEM) showed that microporous CHS sponges were formed around silk cables. Each end of scaffold was modified with lentiviral-mediated transforming growth factor- β 3 (TGF- β 3) gene. The cells on scaffold were transfected by bonded lentivirus. In vitro culture demonstrated that mesenchymal stem cells (MSCs) on scaffolds proliferated vigorously and produced abundant collagen. The transcription levels of cartilage-specific genes also increased with culture time. After 2 weeks, the MSCs were distributed uniformly throughout scaffold. Deposited collagen was also found to increase. The chondral differentiation of MSCs was verified by expressions of collagen II and TGF- β 3 genes in mRNA and protein level. Histology also confirmed the production of cartilage extracellular matrix (ECM) components. The results demonstrated that gene-modified silk cable-reinforced CHS scaffold was capable of supporting cell proliferation and differentiation to reconstruct the cartilage layer of interface.
在膝关节前交叉韧带（ACL）重建术后，移植物与骨之间缺乏纤维软骨层仍然是移植物失败的主要原因。本研究的目的是开发一种基因修饰的丝缆增强硫酸软骨素-透明质酸-丝素蛋白（CHS）混合支架，用于重建纤维软骨层。该支架通过将CHS混合物与编织丝缆冻干制成。扫描电子显微镜（SEM）显示，在丝缆周围形成了微孔CHS海绵。支架的两端用慢病毒介导的转化生长因子-β3（TGF-β3）基因进行修饰。支架上的细胞通过结合的慢病毒进行转染。体外培养表明，支架上的间充质干细胞（MSCs）增殖旺盛，并产生大量胶原蛋白。软骨特异性基因转录水平也随培养时间增加。2周后，MSCs均匀分布于整个支架。还发现沉积的胶原蛋白增加。通过胶原蛋白II和TGF-β3基因在mRNA和蛋白水平的表达，证实了MSCs的软骨分化。组织学也证实了软骨细胞外基质（ECM）成分的产生。结果表明，基因修饰的丝缆增强CHS支架能够支持细胞增殖和分化，以重建界面的软骨层。
