陳錦澤2006-07-262018-07-112006-07-262018-07-112004http://ntur.lib.ntu.edu.tw//handle/246246/23658心臟纖維化為由高血壓及心肌梗塞等疾 患所引發的基本反應，導致心臟舒張功能失 常及衰竭，臨床上由心臟舒張功能不良所引 起之心臟衰竭約佔30 到50%，心臟纖維化的 過程為心臟纖維細胞的增生及胞外基質的堆 積。血管收縮素會造成心臟纖維細胞的增 生，然而其分子機制目前尚不清楚；本計劃 的部份研究結果發現：血管收縮素會增加內 皮素基因的表現，並於纖維細胞增生過程中 扮演重要的角色。近期，糖尿病用藥 rosiglitazone 經由活化細胞內過氧小體增生 活化受體(peroxisome proliferator-activated receptor,PPAR) 能產生許多有益於心血管系統的作用，引起 醫界的高度重視。最新的文獻報告在鼠的實 驗觀察發現：rosiglitazone 有改善心臟肥大及 心肌梗塞所造成的損傷等作用，然 rosiglitazone 是否會影響血管收縮素誘發心 臟纖維細胞增生以及內皮素基因表現，進而 改善心臟纖維化的現象，目前仍然未知。所 以本研究同時深入探討內皮素在血管收縮素 誘發心臟纖維細胞增生所扮演的角色及血管 收縮素誘發內皮素基因表現的調控機制，探 究rosiglitazone 對於血管收縮素誘發心臟纖 維細胞增生以及內皮素基因表現的影響。於 培養的初生鼠心臟纖維細胞給予血管收縮素 的處理後，分析去氧核醣核酸合成的增加以 及內皮素的基因表現，同時觀察rosiglitazone 對血管收縮素誘發心臟纖維細胞增生及內皮 素基因表現的作用。實驗結果顯示:血管收縮 素增加細胞增生的作用可為其拮抗劑 losartan 及內皮素A 型受體拮抗劑BQ485 所 阻斷。以北方式點墨法搭配啟動子活性分析 法確定血管收縮素會誘發內皮素基因表現， 胞外訊號調節激酶（ERK）抑制劑 PD98059 可完全抑制血管收縮素誘發內皮素基因的表 現，進一步共轉染Ras 、 Raf 以及MEK1 的負 向突變可減弱血管收縮素之增加內皮素基因 啟動子活性，顯示血管收縮素的誘發內皮素 基因須要經由Ras-Raf-ERK 路徑，去除部份 啟動子區域搭配突變分析法進一步發現在血 管收縮素誘發內皮素基因表現時，內皮素基 因啟動子區域內的活化蛋白1（activator protein-1;AP-1）的結合部位扮演重要的角 色；rosiglitazone 則有抑制血管收縮素增加細 胞增生以及內皮素基因表現的作用。Fibroblasts play an important role in maintaining cardiac function by providing structural support for the cardiomyocytes and serving as a source for autocrine/paracrine growth factors. After myocardial infarction, reactive fibrosis results in excessive scar formation as proliferating fibroblasts invade the necrotic area. This remodeling leads to an increase of the ventricular stiffness and ultimately compromises the function of the heart. Recent studies in humans and animal models have shown that the expression of myocardial endothelin-1 (ET-1) is increased during cardiac fibrosis. It is suggested that ET-1 might contribute to cardiac fibroblast proliferation resulting in cardiac fibrosis. ET-1 works as a paracrine as well as an autocrine. In the first part of this project, our data revealed that angiotensin II (Ang II)-stimulated fibroblast proliferation is mediated by release of ET-1 from fibroblasts rather than by direct action. Rosiglitazone, approved by the FDA during the spring of 1999, was the second thiazolidinedione to be marketed in the United States. Rosiglitazone improves insulin sensitivity in patients with type 2 diabetes by activating peroxisome proliferator-activated receptor-γ (PPAR摯瑬敳獩 ) receptors in adipose tissues, skeletal muscles, and the liver. Using Northern blot analysis and reverse transcriptase-polymerase chain reaction in samples of rat heart, Wayman et al. (2002) documented the expression of the mRNA for PPAR摯瑬敳獩 (isoform 1 but not isoform 2) in freshly isolated cardiac myocytes and cardiac fibroblasts and in the left and right ventricles of the heart. Using a rat model of regional myocardial ischemia and reperfusion (in vivo), Wayman et al. (2002) discovered that rosiglitazone causes a substantial reduction of myocardial infarct size in the rat. Treatment of normal and diabetic rat hearts with rosiglitazone, also improves postischemic functional recovery. Using Western immunoblotting, it was demonstrated that the acute cardioprotective effect of rosiglitazone is associated with an inhibition of Jun NH(2)-terminal kinase phosphorylation in both normal and diabetic rat hearts. Furthermore, rosiglitazone also inhibited activating protein-1(AP-1) DNA-binding activity. Our data indicate that rosiglitazone inhibits Ang II–induced proliferation in cardiac fibroblasts. Ang II increased DNA synthesis which was inhibited with rosiglitazone. Rosiglitazone inhibited the Ang II-induced ET-1 gene expression as revealed by Nothern blotting and promoter activity assay. Rosiglitazone also inhibited Ang II-increased intracellular ROS generation as measured by a redox sensitive fluorescent dye, 2' 7'-dichlorofluorescin diacetate, and ERK phosphorylation. Furthermore, rosiglitazone and antioxidants such as N-acetyl-cysteine and diphenylene iodonium decreased Ang II-induced cell proliferation, ET-1 promoter activity, ET-1 mRNA, ERK phosphorylation, and activator protein-1-mediated reporter activity. In summary, our results suggest that rosiglitazone inhibits Ang II-induced cell proliferation and ET-1 gene expression, partially by interfering with the ERK pathway via attenuation of ROS generation.application/pdf268729 bytesapplication/pdfzh-TW國立臺灣大學醫學院內科血管收縮素內皮素rosiglitazone細胞增生訊息傳遞心臟纖維細胞過氧 小體增生活化受體angiotensin IIendothelin-1proliferationsignal transductioncardiac fibroblastsperoxisome proliferator-activated receptor[SDGs]SDG3reporthttp://ntur.lib.ntu.edu.tw/bitstream/246246/23658/1/922314B002309.pdf