Distribution and relative activity of matrix metalloproteinase-2 in human coronal dentin.
Distribution and relative activity of matrix metalloproteinase-2 in human coronal dentin.
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DOI：
10.4248/ijos11070
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发表时间：
2011-10
影响因子：
14.9
通讯作者：
Yamauchi M
中科院分区：
文献类型：
--
作者：
Boushell LW;Kaku M;Mochida Y;Yamauchi M
基质金属蛋白酶-2(MMP2)在牙本质中的表达已有报道，但其在成熟人冠牙本质中的分布和活性水平尚不清楚。本研究的目的是确定脱矿牙本质中基质金属蛋白酶-2的分布和相对活性。将28颗磨牙的牙冠分为内(ID)、中(MD)和外牙本质(OD)区，进行脱矿处理。用0.33m ol·L−-1EDTA/2m ol·L-−-1盐酸胍提取基质金属蛋白酶-2，用双抗体夹心酶联免疫吸附测定法测定基质金属蛋白酶-2的浓度。进一步的鉴定通过Western blotting分析和明胶酶谱完成。牙本质区每毫克牙本质蛋白中MMP2的平均浓度差异有统计学意义(P=0.043)，分别为0.9 ng(ID)、0.4 ng(MD)和2.2 ng(OD)。基质金属蛋白酶-2的浓度模式为OD、ID、MD。免疫印迹分析检测到∼66和∼72 kDa的免疫阳性蛋白分别对应于前和成熟的MMP2，而∼66 kDa的蛋白在OD中表达。所有区域均检测到与基质金属蛋白酶-2一致的明胶溶解活性。有趣的是，Western印迹免疫检测水平和明胶溶解活性的模式是MD>；ID>；OD。人冠状牙本质中基质金属蛋白酶-2的浓度在牙本质中最高，在牙本质中最低，在牙本质中最低。然而，蛋白质印迹免疫检测的水平和明胶溶解活性与估计的基质金属蛋白酶-2的区域浓度并不相关，这可能表明区域特异性的蛋白质相互作用。
The presence of matrix metalloproteinase-2 (MMP-2) in dentin has been reported, but its distribution and activity level in mature human coronal dentin are not well understood. The purpose of this study was to determine the MMP-2 distribution and relative activity in demineralized dentin. Crowns of twenty eight human molars were sectioned into inner (ID), middle (MD), and outer dentin (OD) regions and demineralized. MMP-2 was extracted with 0.33 mol·L−1 EDTA/2 mol·L−1 guanidine-HCl, pH 7.4, and MMP-2 concentration was estimated with enzyme-linked immunoabsorbant assay (ELISA). Further characterization was accomplished by Western blotting analysis and gelatin zymography. The mean concentrations of MMP-2 per mg dentin protein in the dentin regions were significantly different (P=0.043): 0.9 ng (ID), 0.4 ng (MD), and 2.2 ng (OD), respectively. The pattern of MMP-2 concentration was OD>ID>MD. Western blotting analysis detected ∼66 and ∼72 kDa immunopositive proteins corresponding to pro- and mature MMP-2, respectively, in the ID and MD, and a ∼66 kDa protein in the OD. Gelatinolytic activity consistent with MMP-2 was detected in all regions. Interestingly, the pattern of levels of Western blot immunodetection and gelatinolytic activity was MD>ID>OD. The concentration of MMP-2 in human coronal dentin was highest in the region of dentin that contains the dentinoenamel junction and least in the middle region of dentin. However, levels of Western blot immunodetection and gelatinolytic activity did not correlate with the estimated regional concentrations of MMP-2, potentially indicating region specific protein interactions.