Alterations in mRNA and protein levels of metalloproteinases-2,-9, and-14 and tissue inhibitor of metalloproteinase-2 responses to traumatic skeletal muscle injury
Alterations in mRNA and protein levels of metalloproteinases-2,-9, and-14 and tissue inhibitor of metalloproteinase-2 responses to traumatic skeletal muscle injury
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DOI：
10.1152/ajpcell.00217.2009
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发表时间：
2009-12-01
影响因子：
5.5
通讯作者：
Urso, Maria L.
中科院分区：
文献类型：
--
作者：
Barnes, Brian R.;Szelenyi, Eric R.;Urso, Maria L.
首页--期刊主要分类--期刊细介绍--期刊题录与文摘内容创伤性骨骼肌损伤后金属蛋白酶-2、-9和-14及其组织抑制因子-2基因和蛋白水平的变化。Am J Physiol Cell Physiol 297：C1501-C1508,2009。2009年9月30日首次出版；DOI：10.1152/ajpcell.00217.2009。-这项研究描述了损伤后骨骼肌中膜类型1基质金属蛋白酶(MT1-MMPs)和金属蛋白酶组织抑制因子-2(TIMP2)表达的时间关系。用冷钢探头(-79℃)暴露60只小鼠的胫前肌，致伤10只S，分别于伤后3、10、24、48、72h采集未损伤和损伤小腿的胫前肌，用定量RT-PCR、免疫印迹、酶谱和免疫荧光等方法分析其局部MT1-MMP1、TIMP-2、基质金属蛋白酶-2和基质金属蛋白酶-9(MMP2、MMP9)的表达。所有数据均以受伤小腿与未受伤小腿的折叠变化表示。伤后48h和72h，MT1-MMPmRNA表达水平分别下降9倍和21倍(P<；0.01)。伤后10~72h，伤侧小腿TIMP-2和基质金属蛋白酶-2mRNA的表达均显著下降，降幅接近4~10倍(P<；0.01)。损伤后10、24、48h，6-(P<；0.05)、25-和12倍(P<；0.01)的基质金属蛋白酶-9的mRNA表达显著增加。损伤后48h和72h，潜伏期(63 KDa)的MT1-基质金属蛋白酶的蛋白含量下降了近2倍(P<；0.01)。MT1-MMP50 kDa的可溶性片段含量在伤后24、48和72h分别增加17、25和67倍(P<；0.01)。TIMP-2蛋白水平在伤后3-48小时下降1.5倍至1.8倍(P<；0.01)，伤后72小时恢复到基线水平。凝胶电泳图显示明胶酶活性在对应于基质金属蛋白酶-9和基质金属蛋白酶-2的分子量区明显增加。综上所述，骨骼肌损伤启动了MT1-MMP蛋白水解级联反应的一系列事件，导致MT1-MMP50 kDa的可溶性片段水平升高，这可能促进细胞外基质的重塑。
Barnes BR, Szelenyi ER, Warren GL, Urso ML. Alterations in mRNA and protein levels of metalloproteinases-2, -9, and -14 and tissue inhibitor of metalloproteinase-2 responses to traumatic skeletal muscle injury. Am J Physiol Cell Physiol 297: C1501-C1508, 2009. First published September 30, 2009; doi:10.1152/ajpcell.00217.2009.-This study characterizes the temporal relationship of membrane type-1 matrix metalloproteinase (MT1-MMP) and tissue inhibitor of metalloproteinase-2 (TIMP-2) expression in skeletal muscle following injury. Tibialis anterior (TA) muscles from 60 mice were exposed and injured by applying a cold steel probe (-79 degrees C) to the muscle for 10 s. Thereafter, TA muscles from uninjured and injured legs were collected at 3, 10, 24, 48, and 72 h postinjury for analysis of local MT1-MMP, TIMP-2, and matrix metalloproteinases-2 and -9 (MMP-2 and MMP-9) mRNA and protein content via quantitative RT-PCR, immunoblotting, zymography, and immunofluorescence. All data are expressed as fold change of injured leg vs. uninjured leg. MT1-MMP mRNA levels were decreased significantly at 48 and 72 h postinjury by similar to 9- and 21-fold, respectively (P < 0.01). Both TIMP-2 and MMP-2 mRNA expression significantly decreased in the injured leg by similar to 4- to 10-fold at 10-72 h postinjury (P < 0.01). MMP-9 mRNA expression was significantly increased at 10, 24, and 48 h postinjury by 6- (P < 0.05), 25-, and 12-fold (P < 0.01), respectively. Protein content of latent (63 kDa) MT1-MMP was decreased at 48 and 72 h postinjury by similar to 2-fold (P < 0.01). Content of the soluble (50 kDa) fragment of MT1-MMP was significantly increased by similar to 17-, 25-, and 67-fold at 24 (P < 0.05), 48, and 72 h (P < 0.01) postinjury, respectively. TIMP-2 protein levels diminished from 3 to 48 h postinjury by 1.5-fold to 1.8-fold (P < 0.01), before returning to baseline levels at 72 h postinjury. Zymography revealed visual increases in gelatinase activity in molecular weight regions corresponding to MMP-9 and MMP-2. In conclusion, skeletal muscle injury initiates a sequence of events in the MT1-MMP proteolytic cascade resulting in elevated levels of the soluble (50 kDa) fragment of MT1-MMP, which could enhance pericellular extracellular matrix remodeling.