Trindade Michael C D, Shida Jun-ichi, Ikenoue Takashi, Lee Mel S, Lin Eric Y, Yaszay Burt, Yerby Scott, Goodman Stuart B, Schurman David J, Smith R Lane
Orthopaedic Research Laboratory, Stanford University School of Medicine, Stanford, CA 94305-5341, USA.
Osteoarthritis Cartilage. 2004 Sep;12(9):729-35. doi: 10.1016/j.joca.2004.05.008.
This study tested the hypothesis that intermittent hydrostatic pressure applied to human osteoarthritic chondrocytes modulates matrix metalloproteinase and pro-inflammatory mediator release in vitro.
Human osteoarthritic articular chondrocytes were isolated and cultured as primary high-density monolayers. For testing, chondrocyte cultures were transferred to serum-free medium and maintained without loading or with exposure to intermittent hydrostatic pressure (IHP) at 10 MPa at a frequency of 1 Hz for periods of 6, 12 and 24 h. Levels of matrix metalloproteinase-2, -9 (MMP-2, -9), tissue inhibitor of metalloproteinase-1 (TIMP-1), and the pro-inflammatory mediators, interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1), released into the culture medium were assessed by ELISA. Matrix metalloproteinase activity was confirmed by zymographic analysis.
In the absence of IHP, levels of MMP-2, TIMP-1, IL-6, and MCP-1 in the chondrocyte culture medium increased in a time-dependent manner. Application of IHP decreased MMP-2 levels at all time periods tested, relative to unloaded control cultures maintained for the same time periods. Although 84/82 kDa bands were faintly detectable by zymography, MMP-9 levels were not quantifiable in medium from loaded or unloaded cultures by ELISA. TIMP-1 levels were not altered in response to IHP at any time period tested. IL-6 and MCP-1 levels decreased in cultures exposed to IHP at 12 and 24 h, relative to unloaded control cultures maintained for the same time periods.
IHP decreased release of MMP-2, IL-6 and MCP-1 by osteoarthritic chondrocytes in vitro suggesting that pressure influences cartilage stability by modulating chondrocyte expression of these degradative and pro-inflammatory proteins in vivo.
本研究验证了这样一个假设，即施加于人类骨关节炎软骨细胞的间歇性静水压力在体外可调节基质金属蛋白酶和促炎介质的释放。
分离人类骨关节炎关节软骨细胞，并将其培养为原代高密度单层细胞。为进行测试，将软骨细胞培养物转移至无血清培养基中，在不加载或暴露于10兆帕、1赫兹频率的间歇性静水压力（IHP）下分别维持6、12和24小时。通过酶联免疫吸附测定（ELISA）评估释放到培养基中的基质金属蛋白酶-2、-9（MMP-2、-9）、金属蛋白酶组织抑制剂-1（TIMP-1）以及促炎介质白细胞介素-6（IL-6）和单核细胞趋化蛋白-1（MCP-1）的水平。通过酶谱分析确认基质金属蛋白酶活性。
在无IHP情况下，软骨细胞培养基中MMP-2、TIMP-1、IL-6和MCP-1的水平呈时间依赖性增加。与在相同时间段维持的未加载对照培养物相比，施加IHP在所有测试时间段均降低了MMP-2水平。尽管通过酶谱法可微弱检测到84/82 kDa条带，但通过ELISA无法对加载或未加载培养物培养基中的MMP-9水平进行定量。在任何测试时间段，TIMP-1水平均未因IHP而改变。与在相同时间段维持的未加载对照培养物相比，在暴露于IHP 12和24小时的培养物中，IL-6和MCP-1水平降低。
IHP在体外降低了骨关节炎软骨细胞中MMP-2、IL-6和MCP-1的释放，这表明压力通过在体内调节这些降解性和促炎蛋白的软骨细胞表达来影响软骨稳定性。
