組織蛋白去乙醯酶抑制劑誘發CD1d表現之研究

Investigation of HDAC Inhibitors-induced CD1d Expression

林沛潔(Pei-Jie Lin)

指導教授 ： 陳青周

國立臺灣大學/醫學院/藥理學研究所/碩士(2008年)

https://doi.org/10.6342/NTU.2008.02230

摘要

本論文探討新穎抗癌藥物，HDAC抑制劑引發CD1d表現與其免疫調節之功能。在A549和TC-1細胞，TSA和SAHA引發dose-及time-dependent之CD1d表現，進一步探討TSA誘導CD1d mRNA表現之機轉，A549及TC-1細胞預先處理PKC抑制劑Ro318220、ROCK抑制劑Y27632、CaM kinase II抑制劑KN-62、p38抑制劑SB203580、MEK抑制劑PD98059、JNK抑制劑SP600125、PI3K抑制劑LY294002、mTOR抑制劑rapamycin、Akt抑制劑SH-5或PI3K抑制劑wortmannin，皆不影響TSA所誘導之CD1d mRNA表現。Sp1抑制劑MTM可阻斷SAHA所誘導的CD1d mRNA表現及Sp1 luciferase和RARE3-tk-luciferase reporter活性。將GAL4-Sp1及Fc-luciferase reporter同時transfect至A549細胞，發現HDAC抑制劑是經由增加Sp1 transactivation活性促進Sp1之reporter活性。ChIP實驗證明，HDAC抑制劑會增加Sp1與乙醯化histone-H3結合至CD1d啟動子。將TC-1細胞與C57BL/6小鼠之脾臟細胞共同培養，HDAC抑制劑所誘導表現之CD1d蛋白，可呈現glycolipid給NKT細胞，使之活化增加IFN-γ釋放，但降低IL-4之釋放。DNMT抑制劑會促進HDAC抑制劑增加CD1d之mRNA與蛋白表現；HDAC6抑制劑tubacin不影響CD1d之mRNA，但增加膜上蛋白之表現。

關鍵字

組織蛋白去乙醯酶CD1d

並列摘要

We investigate the effect of novel anticancer drugs, HDAC inhibitors on CD1d expression in tumor cells, and the related immune regulation. TSA and SAHA were found to induce CD1d expression in a dose- and time-dependent manner in A549 ( human lung adenocarcinoma cell line) and TC-1 ( mouse lung cancer cell line) cells. TSA-induced CD1d was not blocked by the pre-treatment with PKC inhibitor (Ro318220)、ROCK inhibitor (Y27632)、CaM kinase II inhibitor (KN-62)、p38 inhibitor (SB203580)、MEK inhibitor PD98059、JNK inhibitor (SP600125)、PI3K inhibitor (LY294002)、mTOR inhibitor (rapamycin)、Akt inhibitor (SH-5) or PI3K inhibitor (wortmannin). However, Sp1 inhibitor MTM blocked the CD1d mRNA expression, Sp1 luciferase and RARE3-tk-luciferase reporter activity induced by SAHA. Co-transfection of GAL4-Sp1 and Fc-luciferase reporter demonstrated that HDAC inhibitor induced Sp1 luciferase reporter activity by enhancing Sp1 transactivation activity. The binding of Sp1 and acetylated histone-H3 to CD1d promoter was increased by HDAC inhibitors. Co-culture of C57BL/6 splenocytes with SAHA-treated TC-1 cells showed the presentation of glycolipid to the splenocytes, resulting in the increased secretion of IFN-γ and decreased secretion of IL-4. DNMT inhibitor, Zebularine promoted the CD1d induction by HDAC inhibitor. HDAC6 inhibitor, tubacin induced CD1d protein but not mRNA expression. In summary, these results indicate that HDAC inhibitors could up-regulate CD1d expression in tumor cells, and tumor/glycolipid are effective for IFN-γ secretion by splenocytes to exert possible anti-tumor activity.

並列關鍵字

HDAC inhibitors ； CD1d

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