生物型人工韧带的制备及体外检测

背景：目前临床上修复膝关节叉韧带断裂的材料有自体膑腱（BTB）、自体半腱肌和股薄肌，以及同种异体组织移植等，但这些方法都存在一定的局限性。因此，人们研制人工韧带希望能够代替自体和异体移植。目的：探讨以猪肌腱韧带为原料经新的组织处理技术构建生物型人工韧带的可行性。设计：新型生物材料试验。单位：中山大学附属第三医院骨科。材料：肌腱韧带的来源成年猪由中山大学动物中心提供，雌雄不拘。S-520型扫描电镜为日本日立公司产品；LWK-10B微控电子拉力试验机为广州实验设备厂产品。方法：实验于2004-01／2005—06在中山大学动物中心完成。以猪肌腱韧带为原料应用环氧化物作为交联剂加上专门的除抗原技术及蛋白修饰技术制作生物型人工韧带。将来源于6周龄山羊的骨髓基质细胞体外培养后，移植于人工韧带支架中培养3周，应用扫描电镜观察人工韧带形态结构及细胞相容性：应用力学测试分析生物型人工韧带的生物力学特点。主要观察指标：人工韧带形态结构、细胞相容性的生物力学特点。结果：①人工韧带的形态结构：生物型人工韧带的外观与正常肌腱外观相似。组织学检查显示韧带为无细胞的胶原纤维。电镜检查显示韧带为走向一致的发状纤维样物紧密平行排列，表面有棉絮样物附着，未见细胞。②人工韧带细胞相容性：扫描电镜观察，异种骨髓基质细胞于支架中培养3周后，细胞呈球形黏附在韧带材料表面及孔隙侧壁，并可见细胞分泌的基质沉积于细胞周围，但基质分泌量较少。③力学测试结果：人工韧带直径约5mm，最大拉力为927．19N，抗拉强度47．22N／mm，速度100mm／min，而正常山羊的ACL直径约5mm，最大拉力为807．50N，抗拉强度41.13N／mm。结论：应用新的组织处理技术研制的生物型人工韧带有效?

Preparation of artificial biological ligament and its detection in vitro

BACKGROUND： Currently, the materials used in clinical practice to repair cruciate ligament of knee joint contain auto-graft bone- mid 1/3 patella tendon-bone （B-T-B）, auto-semitendinous muscle, gracilis muscle and allogenic tissue graft. All of them are limited to a certain degree in clinical application. Therefore, people hope to consistently develop artificial ligaments to take the place of auto- and allografts. OBJECTIVE： To investigate the feasibility to construct artificial biological ligament （ABL） by applying a novel biochemical technique using porcine tendon as the raw material. DESIGN： Research of new biological material. SETTING： Department of Orthopedics, Third Affiliated Hospital of Sun Yat-sen University. MATERIALS： Adult pigs of either gender were provided by the Animal Center of Sun Yat-sen University. Scanning electron microscope （SEM, S-520） was provided by Hitachi, Japan, and micro-controlled electron tension-testing device （Model LWK-10B） by Guangzhou Experimental Devices Factory. METHODS： The experiment was performed at the Animal Center of Sun Yat-sen University from January 2004 to June 2005. ABL was established by means of treating porcine tendon with epoxy cross-linking fixation, diversified antigen minimization process, mechanic enhancement modification and surface activating process. Under aseptic condition, a 6-month-old goat＇s bone marrow was abstracted, and then the bone marrow matrix stem cells were cultured in ABL stent for 3 weeks. Scanning electron microscope was used to observe structure and compatibility of artificial ligament, and mechanics test was used to analyze biomechanics characteristics of ABL. MAIN OUTCOME MEASURES： Structural features, cell compatibility and biomechanics characteristics of ABL. RESULTS： 1. Structural features of ABL- The appearance of ABL was similar to that of the normal human ligament. Histological examination showed that the ABL was collagen fibers with no cells. Electron microscope examination revealed that the ABL