Establishment and verification of rat models of early knee osteoarthritis

doi:10.3969/j.issn.2095-4344.1382

Abstract

Abstract:

BACKGROUND: Papain inducing knee osteoarthritis has no direct effect on collagen and chondrocyte compared with other chemical inducers.
OBJECTIVE: To evaluate the effectiveness of early osteoarthritis model of knee joint in rats by the surface morphology, radiology and histopathology of cartilage.
METHODS: Twenty adult male Wistar rats were provided by Qinglongshan Animal Breeding Farm of Jiangning District, Nanjing, and the study was approved by the Ethics Committee of Liuhe Hospital Affiliated to Yangzhou University Medical College. The rats were randomly divided into control and experimental groups. At 1, 4 and 7 days, 4% normal saline and 4% papain were injected into the rat right knee joint cavity in the control and experimental groups, respectively. Rats were killed at 10 days. The articular cartilage morphology,X-ray and three-dimensional CT images were observed before the execution. Histopathological examination was performed immediately after the execution, and the results were verified by Mankins and OARSI scores.
RESULTS AND CONCLUSION: (1) Compared with the control group, in the experimental group, the limp was obvious, the knee joint swelling on the injection side was obvious, and the hind limbs were unwilling to land. Radiological examination showed that the joint space narrowed obviously, and the value of three-dimensional CT joint space was significantly decreased [(0.680 0±0.016 3)vs.(0.558 0±0.033 9) mm, P < 0.05]. (2) Microscopic examination showed early stage inflammation characteristics at the right knee joint in the experimental group. There were significant differences in the Mankins and OARSI scores (P < 0.05). (3) In summary, early knee osteoarthritis induced by intra-articular injection of papain into the right knee joint of rats can be used in relevant clinical studies, because it provides measurable changes in joint motion, tactile abnormal pain, progressive radiodegeneration and micro-inflammation of articular cartilage, representing the evaluation of osteoarthritis, and providing a basis for studying the pathogenesis of early knee osteoarthritis.

Key words: rat model, early knee osteoarthritis, articular cartilage, papain

CLC Number:

He Qiang1, Yin Hong2, Dai Fenglei2, Zhang Bin3, Sun Xin2, Zeng Yi3, Yan Yifeng1. Establishment and verification of rat models of early knee osteoarthritis[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(27): 4338-4343.

Figures/Tables 2

2.1 实验动物数量分析实验选用Wistar大鼠20只，分为2组，实验过程无脱失，全部进入结果分析。 2.2 给药后一般情况 3次给药后实验组大鼠饮食及活动量减少，明显跛行，注射侧膝关节肿胀明显，后肢不愿着地，体质量明显减轻。对照组大鼠饮食、活动量、体质量均未见明显改变，偶有出现肢体活动障碍及跛行，注射侧膝关节出现轻微肿胀于每次注射后一两天消失。 2.3 大体观察对照组关节软骨呈灰白色，色泽光亮，软骨表面光滑平整，没有关节面裂缝及破损，没有出现软骨下骨质暴露，未见增生，见图1A；实验组软骨呈淡黄白色，色泽暗淡，软骨表面粗糙出现剥脱，有软骨缺损出现软骨下骨质暴露，有增生出现，见图1B。 2.4 X射线检查及图像观察对照组正位X射线显示膝关节间隙正常，未见狭窄，骨质边缘光滑平整，无骨赘生成，见图2A；实验组正位X射线显示膝关节间隙变窄，骨质边缘毛糙有骨赘形成，见图2B。将大鼠右腿牵拉伸直，拍摄X射线片对比关节间隙，能显示出对照组关节间隙明显大于实验组，且清晰度较高，见图2C。 2.5 三维CT检查及图像观察对照组膝关节CT显示关节间隙清晰正常，未见狭窄，关节骨质边缘光滑平整，见图3A；实验组膝关节CT显示关节间隙变窄明显，有清晰骨赘生成，骨质边缘粗糙不光滑，见图3B。从图3A和3B对比结果可见实验组与对照组相比关节间隙明显变窄，有骨赘生成，关节软骨表面粗糙不光滑。采用 SPSS 25.0统计软件来分析对照组、实验组关节间隙分析结果证明存在统计学意义[(0.680 0±0.016 3)，(0.558 0±0.033 9)mm，P=0.04 < 0.05]。 2.6 组织病理检查取标本苏木精-伊红染色后，于显微镜下观察对照组、实验组右膝关节软骨病变部位细胞，分别以Mankins和OARSI分级评估[4-6]。对照组大鼠软骨表面平整，染色清晰，软骨各层清楚可辨，潮线连续完整，细胞排列规则，层次清晰，软骨基质染色均匀，细胞核清晰。见图4A；实验组大鼠软骨表面明显粗糙不平整，表层变薄甚至破损，部分区域的细胞核固缩、坏死，钙化层细胞紊乱排列呈簇状，潮线紊乱不连续或消失，细胞排列紊乱，软骨基质染色明显减轻、不均，见图4B。Mankins评分验证了大鼠膝关节骨性关节炎的早期阶段(10 d内)2组间差异有显著性意义(P < 0.05)。以OARSI分级标准验证了大鼠膝关节骨性关节炎的早期阶段(10 d内)2组间差异有显著性意义(P < 0.05)。见表1。 "

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