软骨下骨刚度增加对关节软骨Ⅱ型胶原和MMP-1表达的影响

Influence of increased stiffness of subchondral bone on the expression of collagen typeⅡand MMP-1 in articular cartilage

目的观察增加软骨下骨刚度后关节软骨中Ⅱ型胶原和基质金属蛋白酶-1(MMP-1)的表达和分布,探讨骨关节炎发病机制。方法调整聚甲基丙烯酸甲酯(PMMA)、甲基丙烯酸甲酯(MMA)、羟基磷灰石(HA)和蒸馏水的比例,使反应温度低于40℃,制成PMMA-HA复合材料。刮除兔胫骨内侧软骨下骨后置入复合材料,对术后3、6、9、12周的关节软骨进行组织学观察,免疫组化法检测Ⅱ型胶原和MMP-1在软骨中的表达和分布,并与空白、对照组相比较。结果复合材料以HA 2g、PMMA 1g、MMA 0.7ml、蒸馏水0.3ml混合时平均最高反应温度为38.17℃,且极限强度和刚度均高于软骨下骨。实验显示关节软骨随的时间的延长出现纤维化、裂隙、变薄,软骨细胞簇集,潮线模糊或消失,Mankin分级逐渐升高;免疫组化显示Ⅱ型胶原表达增加主要在软骨移行层和深层上部,MMP-1表达以软骨表层及中上层居多,两者染色强度随观察时间的延长均逐渐升高。结论软骨下骨刚度增加后软骨中Ⅱ型胶原和MMP-1表达的量均升高,提示软骨下骨硬化对关节软骨退变的发生、发展具有十分重要的作用,可能为骨关节炎发病的主要原因之一。

Objective To observe the expression and distribution of collagen type Ⅱ and matrix metalloproteinase-1（MMP- 1） in articular cartilage after increasing the subchondral bone stiffness, and investigate the pathogenesis of osteoarthritis. Methods Adjusted the proportion among polymethylmethacrylate（PMMA）, methylmethacrylate（MMA）, hydroxyapatite（HA） and distilled water to keep the maximum reaction temperature under 40C, and obtained the composite material PMMA/HA. The composite material PMMA/HA was compressed on a mechanical material machine to determine its ultimate strength and stiffness, which were then compared with those of subchondral bone. Scraped the subchondral bone of medial tihial plateau and filled with the composite material PMMA/HA. Rabbits were sacrificed at 3, 6, 9 and 12 weeks respectively, the articular cartilages of the medial tibial plateau were harvested for histological observation, immunohistochemical analysis was adopted to record the expres sion and distribution of collagen type Ⅱ and MMP-1 in cartilage at protein level, and then compared with the blank and control groups. Results Having HA 2g, PMMA lg, MMA 0.7ml, distilled water 0. 5ml, the mean maximum reaction temperature of the composite material PMMA/HA was 38. 17 C, both the ultimate strength and stiffness of the composite material PMIVLA/HA were higher than those of subchondral bone（P〈0. 05）. With the prolonging of period, the cartilage of experimental groups underwent fibroplasias, tearing, thinning and chondrocyte clustering, and the Mankin＇s grade increased steadily. Collagen type Ⅱ expressed mainly in transition zone and upper deep zone, MMP-1 expressed predominantly in the superficial and upper intermediate layers of cartilage. Their expressions both increased steadily with time passing. Conclusions The expression of collagen type Ⅱ and MMP-1 in cartilage increases with the increase of the subchondral bone stiffness, which suggests that suhcondral bone sclerosis plays an important role in initialization and development of articular cartilage degradation and probably is one of the pathogenies of osteoarthritis.