探討新型組織蛋白去乙醯酶抑制劑影響人類單核球細胞基質金屬蛋白酵素活性之作用及機制

Investigation of Inhibitory Mechanisms of the New Histone Deacetylase Inhibitors on Matrix Metalloproteinase Expression in Human Monocytic Cells

張宛儒(Wan-Ru Chang)

指導教授 ： 蕭哲志

臺北醫學大學/醫學院/醫學科學研究所/碩士(2013年)

摘要

臨床研究指出，在急性骨髓性白血症及敗血症病人的體內，基質金屬蛋白酵素 (Matrix metalloproteinases, MMPs) 會持續被表現並與病人的存活率密切相關，其中MMP-9的過度生成與釋放，會影響發炎疾病及急性骨髓性白血病的病程進展。革蘭氏陰性菌之致病成分脂多醣體 (Lipopolysaccharide, LPS) 會活化白血球，進而產生MMPs並引發組織重組與發炎反應。組織蛋白去乙醯酶抑制劑 (Histone deacetylase inhibitor, HDACi)目前已被應用於臨床前癌症治療，也被證實在發炎性疾病的動物模式中可降低發炎反應。
本研究探討新型組織蛋白去乙醯酶抑制劑AN及其前驅物PBHA，對於LPS誘導THP-1細胞表現及活化MMP-9 之抑制能力及其機轉。由實驗結果顯示，PBHA對HDAC1及HDAC3之活性皆有抑制作用，但不會影響HDAC3及HDAC4之蛋白表現。酵素電泳分析實驗得知AN及PBHA對LPS所誘導的MMP-9表現及活化具有抑制的作用。LPS活化訊息包含NF-κB, Akt及mitogen-activated protein kinase (MAPKs)。在NF-?羠活化之相關實驗中，AN不影響IκBα降解及p65轉位進核，且AN及PBHA對NF-?羠 reporter基因的活化不具抑制作用。另一方面，Akt 及p38活性不受AN及PBHA影響，但AN及PBHA對於ERKs磷酸化上具有濃度抑制。將microarray結果利用IPA資料庫分析，得知ERK及MMP-9表現具有相關性。綜合以上，本研究顯示AN及PBHA可藉由調控ERK訊息路徑抑制MMP-9的表現，期望以此發現做為未來抗發炎藥物發展及癌症治療藥物開發的依據。

關鍵字

組織蛋白去乙醯酶抑制劑 ； 脂多醣體

並列摘要

In clinical study, patients with AML or sepsis showed constitutive release of several matrix metalloproteinases relative to survival rate. Especially the excessive production and release of MMP-9 have been implicated in clinical progression of inflammatory disease and AMLs. Lipopolysaccharide (LPS) is a major component of Gram-negative bacteria and it’s a critical factor to induce pro-inflammatory mediators and degrading enzymes from the leukocytes. Histone deacetylase inhibitors (HDACi) now are used on cancer therapy and shown to suppress inflammatory responses via inhibit inflammatory mediator expression. In thist study, we investigate the inhibitory mechanism of new HDAC inhibitor, PBHA and AN, on the LPS-induced MMP-9 expression in human monocytic (THP-1) cells. First, we found PBHA and AN showed the inhibitory effect on HDAC1 and HDAC3 activity, but the protein expression of HDAC3 and HDAC4 were not affected. PBHA and AN had the inhibition on activation of MMP-9 in THP-1 cell. According to LPS stimulation, the signaling pathways of NF-κB, Akt and mitogen-activated protein kinase have been evaluated. It found that there were no effects on NF-κB signaling as detected by the degradation of IκBα, the p65 translocation and the NF-κB reporter gene assay. On the other hand, Akt or p38 was not affected by PBHA and AN. However, PBHA and AN had the concentration-dependent inhibition on activation of ERK. Consistently, the ERK signaling and IPA-related gene network of MMP-9 were apparently interacted by microarray analyses. In conclusion, we demonstrate that PBHA and AN might involve the ERK signal pathway to attenuate MMP-9 expression, and provide new opportunities for the development of new anti-inflammatory drugs.

並列關鍵字

HDACi ； LPS

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