TRPA1在巨噬细胞表型极化及动脉粥样硬化中的作用及机制

Recruited monocytes can be differentiated to either pro-inflammatory M1 or anti-inflammatory M2 macrophages in the vascular wall. Moreover, the macrophage polarization plays a vital role in the development of atherosclerotic plaque. Transient receptor potential channel A1 (TRPA1) is involved in the regulation of macrophage function and inflammation; however, the role of TRPA1 in macrophage polarization remains unclear. In our preliminary experiments, the results showed that TRPA1 was functionally expressed in monocytes and was down-regulated during the macrophage differentiation. Moreover, the TRPA1 agonist inhibited the proliferation and migration of monocytes. Thus, we hypothesized that TRPA1 may mediate the calcium influx, activate protein kinases, and subsequently regulate macrophage polarization. In our present studies, using ApoE-/- and Trpa1-/-/ApoE-/- mice, we will induce the vulnerable plaque with atherogenic diet and subcutaneously injection of angiotensin II, polarize the bone marrow cells into M1 and M2 macrophages to explore the role of TRPA1 in macrophage polarization and atherosclerosis. The bone marrow transplantation will be performed to confirm the critical role of TRPA1. We will measure the intracellular calcium and inflammatory cytokines. Cells will also be treated with protein kinase inhibitors to reveal the molecular mechanism in TRPA1-indued macrophage polarization. The aim of our study is to study the role and mechanisms of TRPA1 in macrophage polarization and atherosclerosis, and to provide a new idea for the prevention and treatment of atherosclerosis.

单核细胞向巨噬细胞(Mφ)分化过程中可被极化为促炎的M1型和抗炎的M2型，该过程对动脉粥样硬化过程产生重要影响，尤其蛋白激酶，其活性与钙离子内流有关。瞬时受体电位通道A1(TRPA1) 通过介导Ca2+内流调控Mφ功能及炎症平衡，但在其极化中的作用尚不清楚。我们发现TRPA1在单核细胞功能性表达且在向Mφ分化中显著降低、其激动剂可抑制单核细胞增殖和迁移，故提出假设：TRPA1介导Ca2+内流激活蛋白激酶进而调控Mφ极化。为此，拟①用ApoE-/-及Trpa1-/-/ApoE-/-小鼠，以高脂饮食联合Ang-II诱导易损斑块，分离骨髓细胞诱导极化为M1和M2型Mφ，探讨TRPA1在Mφ极化及动脉粥样硬化中的作用，并通过骨髓细胞移植做验证；②检测胞内钙信号和炎症因子表达、蛋白激酶抑制剂干预，揭示TRPA1调控Mφ极化的分子机制，进而阐明其对动脉粥样硬化的作用及机制，为临床防治提供新思路。

研究背景：TRPA1是一种钙离子通道，在动脉粥样硬化斑块中表达增加，但其在巨噬细胞中的调控作用尚不清楚，本研究拟探讨TRPA1在巨噬细胞极化和炎症激活中的作用及机制。结果：经高脂饮食喂养后，与Apo E-/-小鼠相比较，TRPA1-/-/Apo E-/-小鼠动脉粥样硬化斑块面积明显增加，TRPA1激动剂桂皮醛延缓小鼠动脉粥样硬化进展。TRPA1敲除可导致小鼠血管中动脉粥样硬化炎症的相关基因信号表达上调。与Apo E-/-小鼠相比，TRPA1-/-/Apo E-/-小鼠的巨噬细胞M1型标志物表达显著增高、M2型标志物表达明显降低，而TRPA1激动剂可以导致Apo E-/-小鼠的巨噬细胞M1型标志物表达降低、M2型标志物表达增加。进一步实验证实，TRPA1防止EZH2被蛋白酶体降解，催化组蛋白H3K27三甲基化（H3K27me3），H3K27me3与M1型标志物基因启动子结合，抑制M1型标志物的表达，促进巨噬细胞从M1型向M2型转化。结论：TRPA1在动脉粥样硬化血管中表达增加，TRPA1调控巨噬细胞M1/M2型表型转化、炎症反应和动脉粥样硬化进展，因此，TRPA1可能是治疗动脉粥样硬化的新靶点。