Isoeugenodilol抗血管平滑肌細胞增生機制之研究

Study on Antiproliferative Mechamism of Isoeugenodilol in Vascular Smooth Muscle Cells

張玉佩(Yu-Pei Chang)

指導教授 ： 葉竹來

高雄醫學大學/醫學院/醫學研究所/碩士(2004年)

摘要

血管平滑肌細胞不正常的增生是心血管疾病的主要致病原因之一，而β-blockers在治療缺血性心臟疾病方面扮演著一個相當重要的角色。Isoeugenodilol是一種新型甲/乙型阻斷劑（α/β-adrenoceptor blocker），且具有抗氧化的活性。本論文的目的是在比較傳統乙型阻斷劑（如propranolol）與新型乙型阻斷劑（如labetalol、isoeugenolol、isoeugenodilol）對於老鼠主動脈血管平滑肌細胞生長之影響，並更深入探討isoeugenodilol可能的抗增生分子機轉。
實驗結果發現，在10 μM的劑量之下isoeugenodilol和isoeugenolol可以顯著的抑制PDGF-BB所誘發的血管平滑肌細胞遷徙作用，而propranolol和labetalol則沒有這項作用。利用XTT和BrdU插入實驗發現，isoeugenodilol、propranolol、labetalol和isoeugenolol都會劑量相關的抑制10% FBS或20 ng/ml PDGF-BB所誘發的血管平滑肌細胞增生作用。利用流體細胞分析方法分析DNA的含量也發現，血清所引起的細胞生長週期進行會被isoeugenodilol所阻斷，不過卻不會被isoeugenolol所阻斷。利用西方點墨法實驗發現，由FBS或PDGF-BB所引起的ERK 1/2、MEK1/2和Pyk2蛋白質的磷酸化會被isoeugenodilol所抑制。Isoeugenodilol還會抑制PDGF-BB所誘發的細胞內鈣離子濃度增加現象。除此之外，isoeugenodilol還會抑制PCNA蛋白質的表現。
這些實驗結果顯示，isoeugenodilol抑制血管平滑肌細胞增生的作用，可能是經由干擾細胞生長週期的進行以及抑制Pyk2和鈣離子相關的ERK 1/2活化路徑而產生。因為細胞增生現象是粥狀動脈硬塊發展與穩固一項重要的決定因素，所以認為isoeugenodilol的這項性質具有可以抑制粥狀動脈硬化惡化的作用。

關鍵字

抗血管平滑肌細胞增生 ； 甲/乙型阻斷劑 ； 粥狀動脈硬化

並列摘要

Abnormal vascular smooth muscle cell proliferation has a fundamental role in the pathogenesis of vascular diseases. β-blockers play an important role in the treatment of ischemic heart diseases. Isoeugenodilol is a new α/β-adrenoceptor blocker with antioxidant activity. The purposes of this study are to comparison the effects of the traditional β-blockers (i.e. propranolol) and new type β-blockers (i.e. labetalol, isoeugenolol and isoeugenodilol) on cell proliferation of rat aortic vascular smooth muscle cells (VSMCs), and further objective was to elucidate the possible antiproliferative mechanism of isoeugenodilol.
Whereas propranolol and labetalol did not affect VSMCs migration at the concentration of 10 μM, isoeugenodilol and isoeugenolol significantly inhibited platelet-derived growth factor (PDGF)-BB-induced VSMCs migration. Isoeugenodilol, propranolol, labetalol and isoeugenolol significantly inhibited both 10% fetal bovine serum (FBS)- and 20 ng/ml PDGF-BB-induced VSMCs proliferation in a concentration-dependent manner, as demonstrated by XTT and bromodeoxyuridine (BrdU) incorporation assays. Flow cytometry analysis of DNA content revealed blocking of the 10% FBS-inducible cell-cycle progression by isoeugenodilol but not by isoeugenolol. Isoeugenodilol could suppress the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, mitogen-activated protein kinase kinase (MEK) 1/2 and proline-rich tyrosine kinase 2 (Pyk2) activated by 10% FBS or 20 ng/ml PDGF-BB. In addition, isoeugenodilol decreased cytosolic free calcium stimulated by 20 ng/ml PDGF-BB. Furthermore, it also suppressed the levels of proliferative cell nuclear antigen (PCNA) activated by 10% FBS.
These results indicate that isoeugenodilol may inhibit cell proliferation by perturbing cell cycle progression and attenuating activation of the ERK 1/2 pathway coupling with Pyk2 and calcium. Isoeugenodilol thus shows antiatherogenic potential by inhibiting growth factors-stimulated VSMCs proliferation, a critical element in the development of atherosclerosis.

並列關鍵字

atherosclerosis ； antiproliferation ； adrenoceptor blocker

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