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UM E-Theses Collection (澳門大學電子學位論文庫)

Title

Calycosin promotes angiogenesis involving estrogen receptor and mitogen-activated protein kinase (MAPK) signaling pathway

English Abstract

Angiogenesis, the formation of new blood vessels, is essential for normal growth and homeostasis in human body. However, certain diseases can be exacerbated by the loss of balance in angiogenesis, which results in either excessive or insufficient blood vessel formation. Diseases such as cancer, diabetic retinopathy and rheumatoid arthritis are characterized by excessive blood vessel formation while peripheral and coronary ischemia and infarction, chronic wound healing failure and ulcers are characterized by insufficient blood vessel formation. Radix Astragali, named huangqi in Chinese, is commonly used in the prescriptions of traditional Chinese medicine. In traditional systems, it is used to replenish the vital energy for the treatment of lacking strength, anorexia and loose stools, prolapse of uterus and anus, spontaneous sweating, and chronic nephritis with edema and proteinuria, and to dispel pus and accelerate the healing of chronic ulcers. These suggest that Radix Astragli is a potential candidate for treating diseases associated with dysregulation of angiogenesis. The angigogenic effect of Radix Astragali Extract (RAE) and its underlying mechanisms of action are yet to be fully elucidated by real -time imaging-based screening in live cell and zebrafish bioassays. In this study, calycosin, a major constituents isolated from RAE, was identified to have potent angiogenic activity using multiple bioassays. Calycosin specifically induced human endothelial cells (HUVECS) proliferation and the number of branching points during endothelial cell capillary formation on Matrigel but inhibited breast cancer cel 1 proliferation. Calycosin-induced angiogenesis involved activati on of ERK1/2 and p38 MAPK, and specific blockers of ERK1/2 and p38 MAPK inhibited the calycosin-induced HUVEC proliferation. Meanwhile, calycosin competitively bound with estrogen receptor (ER) in a cell free competitive binding assay, and modulated ERa and ERẞ transcriptional reporters in cell based assays. The agonistic activity of ERa and ERẞ induced by calycosin was slight, while the antagonistic activity was significant. Since calycosin selectively modulated ER transcriptional activation activities, the effect of ER inhibitor on calycosin-induced HUVEC proliferation and the expression of phospho-ERK1/2 were examined. These data suggest that calycosin acts as a novel selective estrogen receptor modulator (SERM), and promotes angiogenesis through activation of ER and MAPK signaling pathway. The results provide rationales for further development of Radix Astragali and its active constituent, calycosin as therapeutic agent on treatment of estr ogen deficient problems such as cardiovascular diseases in post-menopausal women.

Chinese Abstract

血管新生(ang ogenesis),是指形成新血管的過程,具有維持人體正常的生 長和平衡的功能。在許多疾病的發生、發展及轉歸、預後中扮演著重要的角色。 近年來,調節血管新生的研究已成為臨床治療中的熱點。研究主要集中在兩個方 面:1許多疾病的發生、發展與血管生成有著密切的關係,如惡性腫瘤在其生長 過程中,腫瘤組織中不斷有新生血管生成,這些新生血管為腫瘤的生長提供了豐 富的供血和營養。同時,由於新生血管管壁較薄,腫瘤細胞易於穿過,從而進入 血循環並發生遠處轉移。因此,如能抑制新生血管的生成,也就可以阻斷腫瘤供 血,達到“餓死”癌細胞的目的。其他一些良性疾病,如類風濕關節炎、眼底病、 銀屑病等也與局部血管過度增生有關。2.促進血管生成對許多疾病的治療是有益 的,對於骨折、創傷患者,如能促進創傷局部的血管生成,就可以改善局部供血, 加快癒合。又如心肌梗塞患者,梗塞部位新生血管的生成對恢復受損心肌組織供 血有重要意義。 黃芪作為一種常用中藥,主要有補氣固表的功效。用於氣虛乏力,自汗,慢性腎 炎蛋白尿等。近來,黃芪及其有效成分黃芪甲苷已被證明具有治療由不規則血管 新生所引起的病症的能力,我們之前的研究也在多種模型上證明黃芪促進血管 新生功能並對其機理進行了闡明。從膜莢黃芪中分離到的毛蕊異黃酮是黃芪中一 種重要的異黃酮類活性化合物,而它也被證實有許多和血管相關的藥理作用。但 是黃芪中的毛蕊異黃酮是否具有血管新生作用及其作用機制卻沒有報導。因此, 本文將以人臍靜脈內皮細胞(HUVEC)作主要體外模型描述毛蕊異黃酮是如何影 響血管新生。毛瑞異黃酮能特異性引起內皮細胞增殖,在管形成實驗中能增加管 型交叉點數量,同時它又能抑制乳腺癌細胞的增殖。此外,免疫印記雜交的結 果指示毛蕊異黃酮能夠快速啟動 ERK1/2, p38 MAPK 和Akt 三個與血管新生關 係密切的蛋白激酶。ERK1/2,p38 MAPK 和Akt 的特異性抑制劑能分別抑制毛 瑞異黃酮介導的內皮細胞增殖。這些都說明毛瑞異黃酮引起的血管新生與 ERK1/2, p38 MAPK 和Akt 有關。同時,螢光偏振的雌激素受體(ER)競爭性結合 實驗顯示毛蕊異黃酮能夠競爭性的與ER 和ERP 結合。而採用 GeneBLAzer beta 報告基因系統進行的雌激素受體轉錄活性測試也表明,毛蕊異黃酮能較弱的啟動 ERa 和ER,並明顯抗雌二醇對ERa和ER 和啟動。 由於毛瑞異黃酮對 ER 的這種選擇性調控,我們考察了ER 抑制劑對毛瑞異黃酮引起的 HUVEC 增殖以 及ERK1/2活化的影響。發現 ER 抑制劑能完全阻斷毛瑞異黃酮產生的這些效應。 所有檢測結果都表明毛蕊異黃酮能夠做為一種選擇性雌激素調節劑發揮作用 (SERM),能通過ER 和MAPK 細胞信號通路促進血管新生。

Issue date

2009.

Author

Tang, Jing Yan

Faculty
Institute of Chinese Medical Sciences
Degree

M.Sc.

Subject

Neovascularization

Materia medica -- Therapeutic use

Supervisor

Lee, Ming-Yuen

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Location
1/F Zone C
Library URL
991005228279706306