Research of bacteria related to periapical diseases activate MMP and inactivate TIMP

基质金属蛋白酶(MMPs)在细胞外基质(ECM)的动态平衡中起着重要作用。金属蛋白酶组织抑制因子(TIMPs)与基质金属蛋白酶及其抑制物(TIMPs)的失衡在组织破坏中起重要作用。据报道，基质金属蛋白酶-1和基质金属蛋白酶-9被与根尖周病发生有关的细菌激活。然而，有关原-基质金属蛋白酶-2的激活和TIMP的失活的报道还很少。本研究的目的是评价培养细胞中基质金属蛋白酶-2原的活化和基质金属蛋白酶-2的失活。1)基质金属蛋白酶-2活性的测定。用明胶酶谱方法进行分析。2)TIMP-1和TIMP-2蛋白的测定用KoDama等人的EIA法测定。在P.G.的培养上清液中未检测到TIMP-1。SBE处理的PL细胞。经提取物处理的HT1080细胞培养上清液中TIMP-1和TIMP-2的含量显著降低。3)TIMP-1活性的测定纯化的TIMP-1溶液与SBE反应后，明胶酶谱检测TIMP-1对基质金属蛋白酶-1或基质金属蛋白酶-2的抑制活性。P.G.SeE可显著降低TIMP-1对基质金属蛋白酶-2的抑制活性。

Matrix metalloproteinases (MMPs) play a critical role in the homeostasis of the extracellular matrix (ECM). The imbalance between production of MMPs and that of their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs), plays an important role in tissue destruction. It has been reported that MMP-1 and MMP-9 are activated by the bacteria associated with the production of periapical diseases. However few reports regarding the activation of pro-MMP-2 and inactivation of TIMP have been published. The purpose of the present study was to evaluate the activation of pro-MMP-2 and inactivation of TIMP in cultured cells.l )Measurement of MMP-2 activityThe supernatant obtained from medium containing PL cells or HT1080 cells and a sonicated bacterial extract (SEE) from P. gingivalis (P.g.) was analyzed by the gelatin zymography method. When PL cells or HT1080 cells were treated with the SBE, visible bands that corresponded to active MMP-2 were observed on the gelatin zymograms.2) Measurement of TIMP-1 and TIMP-2 proteinsThe amount of TIMP-1 was measured by the EIA method of Kodama et al. TIMP-1 was not detected in the supernatant obtained from the medium of cultures of P.g. SBE-treated PL cells. The amount of TIMP-1 and TIMP-2 in the supernatant obtained from HT1080 cell cultures treated with the extract decreased in a P.g. SBE concentration-dependent manner.3) Measurement of TIMP-1 activityAfter a solution of purified TIMP-1 had been reacted with SBE, TIMP-1 retained in the solution was evaluated by gelatin zymography in terms of its inhibitory activity toward MMP-1 or MMP-2. P.g. SEE remarkably decreased the inhibitory activity of TIMP-1 toward MMP-2.