| 摘要: | 血管鈣化在患有動脈粥樣硬化、慢性腎病、糖尿病和高血壓的患者中普遍存在，是心血管疾病發病率和死亡率增加的強而有力的獨立預測因子。血管鈣化是一種活躍的細胞介導過程，類似於成骨作用和骨質疏鬆症，形成機制是受到調控骨骼鈣化的基因所影響。近年來關於心血管鈣化調控機制的研究主要集中在成骨細胞去分化、基質囊泡、細胞外基質分解和礦化以及細胞凋亡等方面。黃腐醇是啤酒花和啤酒中具有生物活性的異戊烯化類黃酮在傳統醫學中長期用作鎮靜劑和抗微生物劑。最近的研究更確定黃腐醇可改善部分新陳代謝症候群指標，讓人們開始關注到黃腐醇對心血管和代謝疾病的保護作用。然而，黃腐醇在血管鈣化中的作用尚未有人研究。因此，在本研究中，我們使用β-glycerophosphate誘導的大鼠血管平滑肌細胞鈣化的體外模型來評估黃腐醇對血管鈣化的作用和機制。實驗中使用大鼠血管平滑肌細胞培養於含有10 mM β-glycerophosphate、100 nM insulin、50 μg/ml ascorbic acid與10 mM sodium pyruvate的10% DMEM培養液中培養，經過14天後與對照組相比，β-GP組顯著地增加大鼠血管平滑肌中的ALP活性和鈣含量，並進一步由von Kossa和Alizarin Red S染色測定證實可形成多細胞結節和鈣離子沉積。透過西方墨點法和免疫細胞化學螢光染色確定成骨細胞分化標誌物和信息傳導途徑的蛋白質表現。我們的數據顯示，經β-GP培養14天後血管平滑肌細胞的確有礦物化的現象產生，而黃腐醇卻可以濃度相關性地降低β-GP誘導的成骨細胞分化和VSMC的鈣化，包括ALP活性，鈣含量和成骨細胞分化標記BMP-2、Runx-2、Pit-1與active β-catenin等蛋白質的表現以及礦化結核的形成。此外，黃腐醇顯示出可以抑制β-GP誘導的ROS產生、細胞凋亡和caspase 3和-9的蛋白質表現，這些是已知會造成血管鈣化的因素。因此，本研究顯示黃腐醇可能在預防鈣化相關血管疾病中起作用， 需要進一步的動物研究來證實這些新發現。
Vascular calcification (VC) is prevalent in patients with atherosclerosis, chronic kidney disease, diabetes mellitus, and hypertension, is a strong independent predictor of increased cardiovascular morbidity and mortality. VC is an active, cell-regulated process. Recent studies on the regulatory mechanism of cardiovascular calcification have been focused osteogenic dedifferentiation, matrix vesicles, extracellular matrix degradation & mineralization and cell apoptosis. Xanthohumol (XN), a bioactive prenylated flavonoid in hops (Humulus lupulus L.) and beer, has long been used in traditional medicine as a sedative and antimicrobial agent. More recently, attention has been devoted to the protective effects on the cardiovascular and metabolic diseases. However, the role of xanthohumol in VC is still unclear. In this study, an in vitro model of rat aortic vascular smooth muscle cells (VSMCs) calcification induced by β-glycerophosphate (β-GP) are used to identify the effect and mechanisms of XN on VC. Methods: Incubation of VSMCs with β-GP for 14 days induced an osteoblast-like morphological change. The mineralization was visualized by Von Kossa and Alizarin red staining. Alkaline phosphatase activity (ALP) and calcium content were also detected. The protein expression of osteoblastic differentiation markers and signaling pathways were determined by western blot and immunocytochemistry. Results: Our data showed that xanthohumol concentration-dependently reduced β-GP-induced osteoblastic differentiation and calcification of VSMCs including ALP activity, calcium content and bone morphogenetic protein-2 (BMP-2), Runt-related transcription factor 2 (Runx2), sodium-phosphate cotransporter Pit-1 and β-catenin expression as well as the formation of mineralized nodule. Furthermore, xanthohumol was shown to inhibit the β-GP-induced ROS production, apoptosis and protein expressions of caspase 3 and -9, which are known contributors to vascular calcification. Therefore, this study indicates that xanthohumol may have a role in prevention of calcification-associated vascular diseases. Further animal studies are needed to substantiate these novel findings. |
