原儿茶酸增加动脉粥样硬化易损斑块稳定性及其抑制miRNA-10b靶点的机制研究

Although it is generally accepted that the rupture of vulnerable atherosclerotic lesions is the major cause of acute cardiovascular and cerebrovascular events in humans, there are lacking effective targets and measures. Impaired engulfment of apoptotic cells (efferocytosis) by macrophages is recently revealed to be a critical mechanism for the formation and development of vulnerable atherosclerotic lesions. Among the different phenotypical and functional specialization (polarization) of macrophages, macrophage polarization toward M2c has the highest capacity of efferocytosis. Our previous studies have shown that protocatechuic acid, one specific photochemical, is able to inhibit the formation of atherosclerosis and miRNA-10b expression in macrophages (Cir Res. 2012). Furthermore, our preliminary data have provided a clue that antagomiR-10b treatment promotes peritoneal macrophage polarization toward M2c and increases the stability of vulnerable atherosclerotic lesions in ApoE-/- mice. In the current study, using ApoE-/- mice as mouse model, we are planning to further investigate whether and how antagomiR-10b treatment affects the stability of vulnerable atherosclerotic lesions, especially through focusing on regulating macrophage efferocytosis by promoting macrophage polarization toward M2c. Next, we would turn to unravel whether protocatechuic acid can enhance the stability of vulnerable atherosclerotic lesions in ApoE-/- mice by inhibition of miRNA-10b. Finally, we would investigate how protocatechuic acid inhibits miRNA-10b in macrophages. These studies would provide evidence whether miRNA-10b, and protocatechuic acid or antagomiR-10b serve as a new candidate target and measure for the prevention and treatment of the vulnerable atherosclerotic lesions, respectively.

动脉粥样硬化易损斑块的破裂是急性心脑血管事件发生的主要原因，目前缺乏有效的防治靶点和措施。巨噬细胞吞噬凋亡细胞（即胞葬）效率下降促进易损斑块的发生发展，而巨噬细胞M2c型极化促进胞葬。我们前期研究发现植物化学物原儿茶酸可抑制动脉粥样硬化斑块形成和巨噬细胞miRNA-10b表达（Cir Res. 2012）。近来我们初步研究提示miRNA-10b拮抗剂可促进ApoE-/-小鼠腹腔巨噬细胞向M2c型极化并增加易损斑块稳定性。本项目拟以M2c型极化促进巨噬细胞胞葬为切入点，进一步明确miRNA-10b拮抗剂对ApoE-/-小鼠易损斑块稳定性的影响及机制；继而研究抑制miRNA-10b在原儿茶酸调控易损斑块稳定性和巨噬细胞胞葬中的作用，并探讨原儿茶酸抑制巨噬细胞miRNA-10b表达的分子机制。该研究将为易损斑块的防治提供新靶点（miRNA-10b）和新措施（原儿茶酸或miRNA-10b拮抗剂）。

动脉粥样硬化易损斑块的破裂是急性心脑血管事件发生的主要原因，目前缺乏有效的防治靶点和措施。营养膳食是影响动脉粥样硬化发生和发展的重要环境因素。膳食多酚可抑制早期动脉粥样硬化的形成，但其对晚期动脉粥样硬化发生发展的影响尚不清楚。我们前期系列研究发现，膳食多酚原儿茶酸不仅可抑制早期动脉粥样硬化的形成，还可促进早期动脉粥样硬化的消退。在此基础上，结合巨噬细胞、ApoE-/-小鼠以及健康成人志愿者为研究对象，运用膳食补充原儿茶酸或富含原儿茶酸的蔬菜菊苣、遗传学和分子生物学等方法，深入研究原儿茶酸或菊苣对晚期动脉粥样硬化即动脉粥样硬化易损斑块发展的影响及抑制巨噬细胞miR-10b表达的机制。本项目研究进一步发现：（1）抑制miR-10b表达虽不能改变ApoE-/-小鼠血浆脂质水平，但可提高巨噬细胞胆固醇逆向转运效率、降低动脉粥样硬化易损斑块面积、增加动脉粥样硬化易损斑块纤维帽厚度和胶原含量、降低动脉粥样硬化易损斑块内钙化面积、降低动脉粥样硬化斑块内凋亡细胞数目；（2）膳食补充原儿茶酸或富含原儿茶酸的蔬菜菊苣可通过提高四氢生物蝶（BH4）水平抑制eNOS脱偶联途径增加一氧化氮含量，进而改善血管舒张功能、降低动脉粥样硬化易损斑块面积、增加动脉粥样硬化易损斑块纤维帽厚度和胶原含量、降低动脉粥样硬化斑块内凋亡细胞和巨噬细胞数目、增加动脉粥样硬化易损斑块内平滑肌细胞数目，并降低动脉粥样硬化易损斑块内的炎症水平；（3）健康成人志愿者食用菊苣（地中海常见蔬菜，并被我国认定为药食两用）后，约24%的菊苣原儿茶酸进入血液。此外，与中山大学凌文化教授合作研究发现，膳食多酚槲皮素可抑制树突状细胞活化抑制动脉粥样硬化。这些原创性成果不仅提供了预防动脉粥样硬化易损斑块破裂的新靶点即miR-10b，还提供了靶向这个新靶点的营养干预措施即膳食补充原儿茶酸,菊苣和槲皮素。