- 學位論文
探討心肌梗塞後心肌纖維化的致病因子與組蛋白去乙醯酶抑制劑對心臟缺血再灌流損傷的影響
Investigation of the pathogenic factors for cardiac fibrosis after myocardial infarction and potential effects of histone deacetylase inhibitors on cardiac ischemia-reperfusion injury
摘要
心肌梗塞 (myocardial infarction; MI) 是全世界人口主要的致死原因之一，其病因是由於冠狀動脈阻塞所引起。雖然經皮冠狀動脈介入手術 (percutaneous coronary intervention; PCI) 進行"再灌流"治療 (reperfusion) 能有效重建冠狀動脈血流，但缺血性細胞死亡以及再灌流治療會引發心臟缺血再灌流損傷(ischemia-reperfusion injury; IR injury)。心臟缺血再灌流損傷代表著梗塞面積 (infarct size) 的擴大和進行性的心肌梗塞後心肌纖維化 (post-MI cardiac fibrosis)，最後將導致心室功能不全，心臟衰竭和不良預後。儘管已有研究嘗試多種藥物治療以預防心臟缺血再灌流損傷，但大多數的研究結果顯示沒有明確療效。因此在本研究中，我們藉由分析動物模型和心肌梗塞病人的血液樣本，探討心肌梗塞後心肌纖維化的潛在致病因子，以及評估選擇性組蛋白去乙醯酶 (histone deacetylase; HDAC) 6抑制劑ACY1215對梗塞面積的影響。 在本論文的第一部分，我們研究了特定趨化因子CXCL9和CCL20對心肌梗塞後心肌纖維化的影響。與健康受試者相比，心肌梗塞病人血液中轉化生長因子-β (transforming growth factor-β; TGF-β) 與CXCL9的表現明顯增強。在異丙腎上腺素 (isoproterenol; ISO) 誘發大鼠心肌梗塞後心肌纖維化的動物模型中，接受ISO注射的大鼠相較於對照組大鼠具有明顯增加的血清CXCL9濃度和顯著的心肌纖維化。血清CCL20濃度升高的趨勢在心肌梗塞病人和ISO注射的大鼠中並不顯著。而細胞實驗發現CXCL9會促進心臟纖維母細胞 (cardiac fibroblast) 的增生和遷移能力。上述研究結果顯示CXCL9在心肌梗塞後的表現明顯增加，且藉由活化心臟纖維母細胞以促進心肌梗塞後心肌纖維化的形成。 在本論文的第二部分，我們檢測選擇性HDAC6抑制劑與缺氧誘導因子-1α (hypoxia inducible factor-1α; HIF-1α) 基因表現的關係。高通量基因表達測定(high-throughput gene expression assay) 顯示HDAC6抑制劑的使用與HIF-1α的基因表現降低有關，由此推測HDAC6抑制劑應在組織缺氧中扮演一定角色。因此我們接著研究選擇性HDAC6抑制劑ACY1215對於梗塞面積的影響。在大鼠心臟缺血再灌流的動物模型中，ACY1215處理的大鼠相較於對照組大鼠具有較小的梗塞面積，較高的血液C-反應蛋白 (C-reactive protein; CRP) 濃度，且血液中TGF-β濃度有下降趨勢。上述結果顯示ACY1215可能藉由調節HIF-1α的基因表現而進一步減少大鼠在心臟缺血再灌流過程中的梗塞面積。 我們的研究結果說明了CXCL9可視為心肌梗塞後心肌纖維化的潛在致病因子與治療目標，HDAC6抑制劑ACY1215可以減少心臟缺血再灌流損傷之後的梗塞面積，血液中TGF-β和CRP的濃度以及HIF-1α的基因表現可以協助我們監控ACY1215的安全性和功效，未來也需要更深入的研究以釐清ACY1215對於CXCL9與心臟纖維母細胞的影響。綜合而言，我們的研究結果對於預防心臟缺血再灌流損傷的藥物研發領域提供了有價值的資訊與研究線索。
並列摘要
Myocardial infarction (MI), mainly caused by coronary artery occlusion, is one of the leading causes of death in the world. Despite restoration of coronary blood flow by percutaneous coronary intervention with reperfusion, ischemic cell death followed by reperfusion eventually leads to ischemia-reperfusion (IR) injury. Cardiac IR injury represents expansion of infarct size and progressive post-MI cardiac fibrosis that result in ventricular dysfunction, heart failure, and poor prognosis. Despite a variety of pharmacological treatment has been tried to prevent cardiac IR injury, the results of most studies showed undetermined conclusions. In our present research, we aimed to investigate the potential pathogenic factors of post-MI cardiac fibrosis and examine the effects of selective histone deacetylase (HDAC) 6 inhibitor ACY1215 on infarct size by analyzing experimental animal models and blood samples from patients with MI. In the first part of this dissertation, we examined the effects of specific chemokines, CXCL9 and CCL20, on post-MI cardiac fibrosis. Patients with MI had significantly enhanced levels of serum transforming growth factor-β (TGF-β) and CXCL9 compared to healthy subjects. Isoproterenol (ISO)-treated rats had increased serum CXCL9 levels and marked cardiac fibrosis compared to control rats. The trend of increased serum CCL20 in patients with MI and ISO-treated rats was not significant. CXCL9-incubated cardiac fibroblasts showed enhanced proliferation and migration. The findings of our study suggested that an enhanced expression of CXCL9 following MI might play a role in post-MI cardiac fibrosis by activating cardiac fibroblasts. In the second part of this dissertation, we initially examined the association between HDAC6 inhibitors and the gene expression of hypoxia inducible factor-1α (HIF-1α), a key regulator of cellular responses to hypoxia. The high-throughput gene expression assay showed that treatment of HDAC6 inhibitors was associated with a decreased gene expression of HIF-1α, implicating that HDAC6 inhibitors might play a role in tissue hypoxia. We subsequently investigated the effects of HDAC6 inhibitor ACY1215 on infarct size in rats with cardiac IR injury. ACY1215-treated rats had a smaller infarct size, a higher serum C-reactive protein (CRP) level, and a trend in decreased serum TGF-β levels compared to control rats. These findings indicated that ACY1215 might reduce infarct size in rats with cardiac IR injury by possibly modulating HIF-1α expression. Our data not only showed that CXCL9 might be a potential pathogenic factor and a treatment target of post-MI cardiac fibrosis, but also revealed that HDAC6 inhibitor ACY1215 might be a promising pharmacological treatment to limit infarct size. Additionally, the serum TGF-β and CRP levels, and expression of HIF-1α are useful biomarkers to monitor the safety and efficacy of ACY1215. Further in-depth studies are needed to clarify the effects of ACY1215 on the expression of CXCL9 and the activities of cardiac fibroblasts. Collectively, the findings of our research provided a valuable implication in the field of drug discovery regarding prevention of cardiac IR injury.
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