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標題: | 探討機械力刺激骨細胞之基質金屬蛋白酵素-3
調控機制 The upregulation of MMP-3 upon mechanical stimulation in bone cells |
作者: | Chih-Ching Liao 廖志清 |
指導教授: | 姚宗珍 |
關鍵字: | 基質金屬蛋白酵素-3,機械力, MMP-3,mechanical stimulation, |
出版年 : | 2009 |
學位: | 碩士 |
摘要: | 矯正治療帶來骨質重塑的現象,代表接受刺激的牙週組織環境有著明顯的改變,在受壓力區的骨質吸收與MMPs酵素分解細胞外基質有密切相關,而MMPs家族中的MMP-3在此現象的角色尚未明朗,本研究利用人類類成骨細胞株MG-63作為實驗對象,培養於3D膠原蛋白凝集體中,模擬生理環境,施予細胞5%週期性壓力刺激24小時後,以基因微陣列分析,發現有四個基因與本實驗室先前所做的結果重複,分別是Ornithine decarboxylase 1 (ODC1),Human methyl sterol oxidase (ERG25) mRNA (SC4MOL),3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR)及本實驗中的MMP-3皆有向上調控的現象。此外,針對MG-63細胞施予1%及5%週期性壓力刺激1小時,3小時及24小時後,以即時定量聚合酶鏈鎖反應(Real-time PCR)分析,發現MMP-3皆有上升趨勢,且皆於24小時後到達高峰,而5%的壓力刺激讓MG-63表現更多的MMP-3,而利用自人類齒槽骨培養之成骨細胞施予5%壓力24小時後,卻發現MMP-3表現有向下減少的趨勢。為了更近一步探討成骨細胞表現MMP-3的可能訊息傳導路徑,本實驗於MG-63自細胞受壓1小時前加入3種MAPK inhibitor,分別為MEK1/2 inhibitor (U0126),JNK inhibitor (SB600125),p38 inhibitor (SB202190),以Real-time PCR分析,發現加入MEK1/2 inhibitor及p38 inhibitor後,原本受壓24小時應上升之MMP-3,有向下調控的現象,因此我們的結果顯示週期性壓力刺激具有促進MG-63細胞的MMP-3向上調控,而可能的機制是經由MEK1/2及p38這兩條路徑來調控。 The bone remodeling process resulted from orthodontic treatment implies the micro-environment of the periodontium undergoing obvious changes. The matrix metalloproteinases (MMPs) in cleaving extra-cellular matrix play important roles for bone resorption during orthodontic tooth movement. MMP-3, one of the members of MMPs which also called stromelysin 1,likely would contribute this process. To verify how the mechanical compression may affect the MMP-3 gene expression, we gave 5% cyclic compression to human osteoblast-like MG-63 cells for 24 hours and performed a microarray analysis. We also tested the effects of 1% and 5% cyclic compression to MG-63 cells for 1, 3 and 24 hours, and of 5% compression to human primary bone cells for 24 hours. Real-time PCR was performed to evaluate the amount of its mRNA expression. Besides, to identify the possible pathways of signal transduction in the regulation of MMP-3, 3 different MAPK inhibitors ,including MEK1/2 inhibitor (U0126), JNK inhibitor (SB600125), and p38 inhibitor (SB202190), were added to the MG-63 cells prior to mechanical compression and their effect were followed by real-time PCR assays. From the microarray data, 4 genes including ODC1, SC4MOL,HMGCR and MMP-3 were again found to be upregulated when compared to our previous study. In real-time PCR analysis of MG-63 cells, MMP-3 was upregulated after 1 hour and peaked at 24 hours either undergoing 1% or 5% compression. The response of MMP-3 was more obvious when 5% stimulation applied than the one when 1% stimulation applied, indicating a force-dependent effect for MMP-3 in MG-63 cells. For the human primary bone cells, the result showed the down regulation for MMP-3 gene instead. For identifying possible pathways of MMP-3 regulation, the real-time PCR results showed that MEK1/2 and p38 inhibitors both had reduced the expression of MMP-3 gene, but not the JNK inhibitor, which on the contrary increased the expression of MMP-3 gene. We concluded that MG-63 cells were responsive to the cyclic compression in a force dependent way and further study may be needed to verify if MEK1/2 and p38 pathways are involved for the mechanical stimulated expresssion of MMP-3. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/42340 |
全文授權: | 有償授權 |
顯示於系所單位: | 臨床牙醫學研究所 |
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