Targeting Filamin A Reduces Macrophage Activity and Atherosclerosis
Targeting Filamin A Reduces Macrophage Activity and Atherosclerosis
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DOI：
10.1161/circulationaha.119.039697
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发表时间：
2019-07-02
期刊：
影响因子：
37.8
通讯作者：
Akyiirek, Levent M.
中科院分区：
文献类型：
--
作者：
Bandaru, Sashidar;Ala, Chandu;Akyiirek, Levent M.
背景：肌动蛋白结合蛋白Flna(微丝A)调节对细胞运动至关重要的信号转导，但巨噬细胞特异的Flna在动脉粥样硬化形成中的作用尚不清楚。方法：采用免疫荧光法检测人颈动脉粥样硬化斑块中Flna的表达。我们还用表达巨噬细胞特异性溶酶体M启动子(Lc)的Cre的小鼠培育了条件FLNA基因敲除小鼠(FLNA(o/fl))，从而产生了Flna缺乏的巨噬细胞。通过将雄性FLNA(o/fl)/LC小鼠的骨髓移植到致动脉粥样硬化的低密度脂蛋白受体缺陷(Ldlr(-/-))小鼠，以及用AdPCSK9(腺病毒载体高表达枯草杆菌枯草杆菌/可信9型)感染FLNA(o/fl)和FLNA(o/fl)/LC小鼠来研究体内的动脉粥样硬化。此外，将AdPCSK9感染C57BL/6小鼠，然后用钙蛋白酶抑制剂calPeptin来抑制Fla的切割。结果：我们发现巨噬细胞Flna在晚期斑块中的表达高于在中级斑块中的表达。与对照组相比，Flna(o/fl)/LC巨噬细胞的增殖和迁移减少；表达低水平的磷酸化AKT和ERK1/2；减少泡沫细胞的形成和脂质摄取；分泌更多的脂质。巨噬细胞中Flna的缺失使Ldlr(-/-bmt：flnao/fl/lc)和AdPCSK9感染的flna(o/fl)/lc小鼠的动脉粥样硬化斑块明显缩小。动脉粥样硬化斑块内膜/中膜比值和CD68阳性巨噬细胞数均低于对照组。此外，我们还发现，STAT3与Flna的钙蛋白裂解的羧基末端片段相互作用。在巨噬细胞中，用钙肽蛋白抑制钙蛋白酶介导的Flna裂解可降低核内磷酸化STAT3、白介素6的分泌、泡沫细胞的形成和脂质摄取。最后，钙肽治疗减少了感染AdPCSK9的C57BL/6小鼠的动脉粥样硬化斑块的大小。结论：FLNA基因失活和化学抑制依赖于钙蛋白酶的裂解作用可损害巨噬细胞的信号和功能，减轻小鼠的动脉粥样硬化，提示靶向Flna的药物可能在动脉粥样硬化的治疗中发挥作用。
Background: The actin-binding protein FLNA (filamin A) regulates signal transduction important for cell locomotion, but the role of macrophage-specific FLNA during atherogenesis has not been explored. Methods: We analyzed FLNA expression in human carotid atherosclerotic plaques by immunofluorescence. We also produced mice with Flna-deficient macrophages by breeding conditional Flna-knockout mice (Flna(o/fl)) with mice expressing Cre from the macrophage-specific lysosome M promoter (LC). Atherosclerosis in vivo was studied by transplanting bone marrow from male Flna(o/fl)/LC mice to atherogenic low-density lipoprotein receptor-deficient (Ldlr(-/-)) mice; and by infecting Flna(o/fl) and Flna(o/fl)/LC mice with AdPCSK9 (adenoviral vector overexpressing proprotein convertase subtilisin/kexin type 9). Furthermore, C57BL/6 mice were infected with AdPCSK9 and then treated with the calpain inhibitor calpeptin to inhibit FLNA cleavage. Results: We found that macrophage FLNA expression was higher in advanced than in intermediate human atherosclerotic plaques. Flna(o/fl)/LC macrophages proliferated and migrated less than controls; expressed lower levels of phosphorylated AKT and ERK1/2; exhibited reduced foam cell formation and lipid uptake; and excreted more lipids. The deficiency of Flna in macrophages markedly reduced the size of aortic atherosclerotic plaques in both Ldlr(-/-BMT:Flnao/fl/LC) and AdPCSK9-infected Flna(o/fl)/LC mice. Intima/media ratios and numbers of CD68-positive macrophages in atherosclerotic plaques were lower in Flna-deficient mice than in control mice. Moreover, we found that STAT3 interacts with a calpain-cleaved carboxyl-terminal fragment of FLNA. Inhibiting calpain-mediated FLNA cleavage with calpeptin in macrophages reduced nuclear levels of phosphorylated STAT3, interleukin 6 secretion, foam cell formation, and lipid uptake. Finally, calpeptin treatment reduced the size of atherosclerotic plaques in C57BL/6 mice infected with AdPCSK9. Conclusions: Genetic inactivation of Flna and chemical inhibition of calpain-dependent cleavage of FLNA impaired macrophage signaling and function, and reduced atherosclerosis in mice, suggesting that drugs targeting FLNA may be useful in the treatment of atherosclerosis.