Influence of joint distraction height on cartilage repair in rabbit models of knee oteoarthritis

doi:10.3969/j.issn.2095-4344.1242

Abstract

Abstract:

BACKGROUND: Joints distraction has been shown to activate cartilage cell regeneration, which has been gradually used in the treatment of osteoarthritis. However, the related basic research of this technology is weak. Especially the influence of different joint distraction height on cartilage repair is little known.
OBJECTIVE: To explore the influence of different joint distraction height on cartilage repair in rabbit knee osteoarthritis.
METHODS: New Zealand white rabbits, aged 5-6 months (provided by Experimental Animal Center of Shanxi Medical University) were randomized into mode and blank control groups. A model of rabbit knee osteoarthritis was made by intra-articular injection of 4% papain. The blank control group underwent intra-articular injection of normal saline. Afterwards, the rabbit models were randomized into modeling, normal joint space distraction, 1.5×joint space distraction and 2.0×joint space distraction groups. The rabbits in the latter three groups, the joint distraction was carried out by placing Ilizarov External Fixator(3/4). The blank control and modeling groups received no treatment. At 6 weeks after surgery, animals of each group were sacrificed for gross observation and macroscopically analyzed using ICRS system and Mankin scoring system. The expression levels of interleukin 1β and matrix metalloproteinase 13 in cartilage tissue were detected by immunohistochemical staining.
RESULTS AND CONCLUSION: (1) The articular cartilage in the blank control group was smooth and intact. In the modeling group, the defect of articular cartilage was obvious. The defect area in the normal joint space distraction group showed cartilage regeneration, but did not completely fill the defect area. In the 1.5×joint space distraction group, the regenerated cartilage was completely filled the defect area, and histological observation showed transparent cartilage. The regenerated cartilage the 2.0×joint space distraction group was completely filled in the defect area, and histological observation showed transparent cartilage. The ICRS score and Mankin score in each distraction group were significantly higher than those in the modeling group (P < 0.001), and the scores in the 1.5× and 2.0×joint space distraction groups were significantly higher than those in the normal joint space distraction group (P < 0.001). (2) The expression levels of interleukin 1β and matrix metalloproteinase 13 in cartilage tissue in each distraction group were significantly lower than those in the modeling group (P < 0.01), and the levels in the 1.5×and 2.0×joint space distraction groups were significantly lower than those in the normal joint space distraction group (P < 0.001). (3) In summary, the regeneration of cartilage can be better achieved by 1.5× and 2.0×joint space distraction groups than the normal joint space distraction.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: joint distraction, cartilage tissue regeneration, joint distraction height, Ilizarov technology, cartilage repair, knee osteoarthritis, mechanical stress, bone defects

CLC Number:

R454.4
R363

Shi Huanan, Zhang Yonghong, Peng Ruijian, Li Xiaohui, Guo Xiujuan. Influence of joint distraction height on cartilage repair in rabbit models of knee oteoarthritis[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(19): 2972-2979.

Figures/Tables 3

2.1 实验动物数量分析新西兰大白兔2只用于验证造模，其余均进入结果分析，无脱落。 2.2 膝关节软骨形态验证造模情况左膝关节X射线：可见空白对照组关节形态正常，未见明显骨赘形成(图2A)；模型组关节间隙变窄，软骨下骨硬化，可见明显的骨赘形成(图2B)。大体观：可见空白对照组软骨呈半透明状，无软化，表面光滑、无裂纹，关节液清亮，滑膜未见明显增生(图2C)；模型组软骨色泽灰暗，负重区可见明显磨损，深达软骨下骨，滑膜增生明显(图2D)。苏木精-伊红染色：可见空白对照组软骨表面光滑整齐，四层结构清晰，中层细胞呈柱状排列(图2E)；模型组软骨病变明显，四层结构破坏，可见全层缺失(图2F)。番红O-固绿染色：可见空白对照组软骨基质呈深红色，潮线清晰，蛋白聚糖含量正常(图2G)；模型组软骨缺损，潮线可见(图2H)。 2.3 关节牵伸后6周各组软骨形态大体观结果空白对照组软骨呈半透明状，无软化，表面光滑、无裂纹，关节液清亮，滑膜未见明显增生(图3A)；模型对照组软骨厚度变薄，软化，表面粗糙，可见多发裂纹，局部深达软骨下骨，滑膜增生明显(图3B)；正常关节间隙牵伸组软骨缺损区可见软骨再生，再生软骨质软，表面欠光滑，再生软骨未完全填充软骨缺损区(图3C)；1.5倍正常关节间隙牵伸组再生软骨完全填充且高出缺损区，再生软骨颜色苍白、质软，表面欠光滑(图3D)；2.0倍正常关节间隙牵伸组再生软骨完全填充缺损区，再生软骨颜色苍白、质软，表面欠光滑(图3E)。2.0倍正常关节间隙牵伸组软骨修复ICRS评分明显高于模型对照组(P < 0.001)，3个实验组评分均低于空白对照组(P < 0.01)，见图4。正常关节间隙牵伸组分值显著低于1.5倍和2.0倍正常关节间隙牵伸组(P < 0.001)，1.5倍和2.0倍正常关节间隙牵伸组无明显差异(P > 0.05)。 2.4 关节牵伸后6周各组软骨苏木精-伊红染色、番红O-固绿染色显微镜观察结果空白对照组软骨表面光滑整齐，四层结构清晰，中层细胞呈柱状排列，番红O-固绿染色显示糖胺多糖均匀表达(图5A)；模型对照组可见明显软骨缺损区，番红O-固绿染色显示糖胺多糖不表达(图5B)；正常关节间隙牵伸组软骨缺损区再生软骨无明显四层结构，以浅层纤维组织为主，深层为较薄的透明软骨，可见成团分布的同源细胞群，番红O-固绿染色显示浅层糖胺多糖不表达，深层可见其表达(图5C)；1.5倍正常关节间隙牵伸组软骨表面欠光滑，可见软骨组织的四层结构，各层软骨细胞排列不整齐，可见软骨裂隙，深层可见多个成团分布的同源细胞群，番红O-固绿染色显示糖胺多糖在软骨表层以下表达(图5D)，但不均匀；2.0倍正常关节间隙牵伸组软骨表面欠光滑，可见软骨组织的四层结构，各层软骨细胞排列不整齐，深层可见多个成团分布的同源细胞群，番红O-固绿染色显示糖胺多糖在软骨全层表达，欠均匀，以深层为主，表层含量低(图5E)。各牵伸组Mankin骨关节病组织评分明显低于模型组(P < 0.001)，显著高于空白对照组(P < 0.001)，其中正常关节间隙牵伸组分值显著高于1.5倍和2.0倍正常关节间隙牵伸组(P < 0.001)，1.5倍和2.0倍正常关节间隙牵伸组间无明显差异(P > 0.05)，见图6。 2.5 关节牵伸后6周各组白细胞介素1β、基质金属蛋白酶13在软骨组织的表达光镜下可见各组白细胞介素1β、基质金属蛋白酶13的表达情况：空白对照组仅见软骨表层极弱表达；模型对照组软骨全层基质、软骨细胞胞浆高表达，呈深棕色，阳性结果较空白对照组显著增强；正常关节间隙牵伸组再生软骨全层表达，以表、中层显著；1.5倍正常关节间隙牵伸组再生软骨以表层表达为主，中层可见散在表达；2.0倍正常关节间隙牵伸组软骨表层仍可见表达，阳性结果高于空白对照组。见图7，8。经Image Pro Plus 6.0图像分析软件分析累积吸光度值，白细胞介素1β、基质金属蛋白酶13累积吸光度值越大，则阳性结果越高。各牵伸组累积吸光度值明显低于模型对照组(P < 0.001)，正常关节间隙牵伸组吸光度值显著高于1.5倍和2.0倍正常关节间隙牵伸组(P < 0.001)，1.5倍和2.0倍正常关节间隙牵伸组间无明显差异(P > 0.05)。各组白细胞介素1β累积吸光度值见图9，基质金属蛋白酶13累积吸光度值见图10。"

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