天然高分子复合人工肌腱组织细胞外基质的构建与表征

Preparation and Characterization of an Extracellular Matrix of Artificial Tendon Tissue from Natural Macromolecules

摘要:

胶原与壳聚糖是两种具有较好生物相容性和一定力学强度的天然高分子, 可在肌腱组织工程中用于细胞外基质的构建, 但二者单独使用时各有不足。本研究利用二者性能上的互补, 在一定的外力场作用下, 采用EDC/NHS对2种天然高分子材料进行共价交联, 获得具有一定空间取向和力学强度的多孔支架, 然后引入细胞黏附因子RGD进行表面修饰, 构建了具有较好组织相容性和细胞亲和性及适当降解速率的人工肌腱组织细胞外基质。对基质材料的力学性能、亲水性、体外降解速率等的检测和显微观察结果显示: 所构建的多孔支架材料柔软富有弹性, 抗拉强度达: 15.0 Mpa, 相应形变为: 7.33%; 孔隙率: 79.4%; 吸水率: 772%; 保水率: 206%; 在RPM1640培养液(含10%胎牛血清)和人血清中, 3周总降解率分别为: 4.13%和37.2%, 其降解速率可与肌腱修复周期相吻合, RGD修饰后材料对3T3-L1细胞具有较好的亲和性。有望成为理想的人工肌腱组织和人造皮肤细胞外基质, 或整形手术的软组织填充材料。

Abstract:

Collagen and chitosan are well natrual polymers to be used as extracellular matrix on tissue engineering because of their biocompatibility, certain mechanical strength and biodegradability. But there are some disadvantages when they are used to construct extracellular matrix respectively. This experiment utilized their complementary performances to prepare a composite extracellular matrix of artificial tendon tissue that had adequacy mechanics strength and good biocompatibility, cell affinity, biodegradability. Collagen and chitosan were covalently crosslinked using EDC and NHS to obtain a porous scaffold material that the porous was oriented under an external force. Then RGD peptide was covalently attached to scaffold material surface to improve its affinity with cells. The microstructure of scaffold material was observed under microscope and scanning electron microscope. Simultaneously, the physical performance, hydrophilicity, ecto-degradation rate and cell compatibility of scaffold material were measured in the experiments. The results showed that this scaffold material was soft and stretchy. Its tensile strength was 15.0 MPa, corresponding shape extension was 7.33%, and its porosity was 79.4%. Its water absorption rate and water retention rate were 772% and 206% respectively. Its degradation rate in RPM1640 culture mediun with 10% fetal bovine serum and in human serum were 4.13% and 37.2% respectively after three weeks. These degradation rates are suitable for the rehab course of injured tendon. Moreover the degradation rates can be controlled by adjusting technological conditions and degree of cross linking. Significantly higher affinity with 3T3-L1 cell was detected on the scaffold material modified by RGD peptide. The various physical performances of this complex scaffold material are appropriate for constructing extracellular matrix of artificial tendon tissue or artificial skin. Moreover, it could be used as soft tissue slurry for plastic surgery.