Rat tail static compression model mimics extracellular matrix metabolic imbalances of matrix metalloproteinases, aggrecanases, and tissue inhibitors of metalloproteinases in intervertebral disc degeneration.
Rat tail static compression model mimics extracellular matrix metabolic imbalances of matrix metalloproteinases, aggrecanases, and tissue inhibitors of metalloproteinases in intervertebral disc degeneration.
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DOI：
10.1186/ar3764
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发表时间：
2012-03-06
影响因子：
4.9
通讯作者：
Nishida K
中科院分区：
文献类型：
--
作者：
Yurube T;Takada T;Suzuki T;Kakutani K;Maeno K;Doita M;Kurosaka M;Nishida K
椎间盘内细胞外基质（ECM）的纵向降解机制尚不清楚。我们的目的是通过静态压缩模型阐明椎间盘退变中分解代谢和合成代谢基因表达谱及其平衡。将48只12周龄雄性Sprague-Dawley大鼠尾部置入带弹簧的ilizarov型装置，以1.3 MPa静态加载56 d。收集实验负载和远端未负载的对照盘，通过实时逆转录聚合酶链反应（PCR）信使RNA定量分析分解代谢基因[基质金属蛋白酶(MMP)-1a, MMP-2, MMP-3, MMP-7, MMP-9, MMP-13，一种带有血小板反应蛋白基序的分解素和金属蛋白酶(ADAMTS)-4和ADAMTS-5]，抗分解代谢基因[金属蛋白酶组织抑制剂(TIMP)-1， TIMP-2和TIMP-3]， ECM基因[凝集蛋白-1，胶原型1-α1，以及促炎细胞因子基因[肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1α、IL-1β和IL-6]。免疫组化检测MMP-3、ADAMTS-4、ADAMTS-5、TIMP-1、TIMP-2和TIMP-3蛋白表达水平和分布。同样研究了MMP-和聚集酶切割的聚集蛋白新表位的存在，以评估聚集蛋白的溶解活性。定量PCR显示所有MMPs和ADAMTS-4均上调，ADAMTS-5未上调。TIMP-1和TIMP-2基本不变，TIMP-3下调。凝集蛋白-1和2型胶原-α1表达下调，1型胶原-α1表达上调。尽管TNF-α升高，但il几乎没有上调。免疫组织化学显示，在髓核中，MMP-cleaved aggrecan neoepitope的免疫阳性细胞百分比从7天到56天增加，MMP-3免疫阳性增加，TIMP-1和TIMP-2免疫阳性降低。仅在第7天和第28天，聚集酶切割的聚集蛋白新表位免疫阳性细胞百分比增加，TIMP-3免疫阳性降低。在纤维环中，mmp切割的聚集蛋白新表位表现出相同的表达模式。仅在第7天和第28天，随着ADAMTS-4和ADAMTS-5免疫阳性的增加，聚集酶切割的聚集蛋白新表位增加。这个大鼠尾部持续静态压缩模型模拟了人类退行性椎间盘中MMPs、聚集酶和TIMPs的ECM代谢失衡。MMP-3/TIMP-1和TIMP-2相对于ADAMTS-4和ADAMTS-5/TIMP-3的明显失衡表明椎间盘退变处于晚期。
The longitudinal degradation mechanism of extracellular matrix (ECM) in the interbertebral disc remains unclear. Our objective was to elucidate catabolic and anabolic gene expression profiles and their balances in intervertebral disc degeneration using a static compression model. Forty-eight 12-week-old male Sprague-Dawley rat tails were instrumented with an Ilizarov-type device with springs and loaded statically at 1.3 MPa for up to 56 days. Experimental loaded and distal-unloaded control discs were harvested and analyzed by real-time reverse transcription-polymerase chain reaction (PCR) messenger RNA quantification for catabolic genes [matrix metalloproteinase (MMP)-1a, MMP-2, MMP-3, MMP-7, MMP-9, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4, and ADAMTS-5], anti-catabolic genes [tissue inhibitor of metalloproteinases (TIMP)-1, TIMP-2, and TIMP-3], ECM genes [aggrecan-1, collagen type 1-α1, and collagen type 2-α1], and pro-inflammatory cytokine genes [tumor necrosis factor (TNF)-α, interleukin (IL)-1α, IL-1β, and IL-6]. Immunohistochemistry for MMP-3, ADAMTS-4, ADAMTS-5, TIMP-1, TIMP-2, and TIMP-3 was performed to assess their protein expression level and distribution. The presence of MMP- and aggrecanase-cleaved aggrecan neoepitopes was similarly investigated to evaluate aggrecanolytic activity. Quantitative PCR demonstrated up-regulation of all MMPs and ADAMTS-4 but not ADAMTS-5. TIMP-1 and TIMP-2 were almost unchanged while TIMP-3 was down-regulated. Down-regulation of aggrecan-1 and collagen type 2-α1 and up-regulation of collagen type 1-α1 were observed. Despite TNF-α elevation, ILs developed little to no up-regulation. Immunohistochemistry showed, in the nucleus pulposus, the percentage of immunopositive cells of MMP-cleaved aggrecan neoepitope increased from 7 through 56 days with increased MMP-3 and decreased TIMP-1 and TIMP-2 immunopositivity. The percentage of immunopositive cells of aggrecanase-cleaved aggrecan neoepitope increased at 7 and 28 days only with decreased TIMP-3 immunopositivity. In the annulus fibrosus, MMP-cleaved aggrecan neoepitope presented much the same expression pattern. Aggrecanase-cleaved aggrecan neoepitope increased at 7 and 28 days only with increased ADAMTS-4 and ADAMTS-5 immunopositivity. This rat tail sustained static compression model mimics ECM metabolic imbalances of MMPs, aggrecanases, and TIMPs in human degenerative discs. A dominant imbalance of MMP-3/TIMP-1 and TIMP-2 relative to ADAMTS-4 and ADAMTS-5/TIMP-3 signifies an advanced stage of intervertebral disc degeneration.
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影响因子：
2.8
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