Background Clinical observation suggests the atheroprotective effect of chloroquine and its derivatives, while its mechanism remains unclear. This study aimed to observe the protective effect of chloroquine against atherosclerosis and explore the underlying mechanism.

Methods Ataxia telangiectasia mutated (ATM) wild-type or haploinsufficient apolipoprotein-E-knockout (ATM^+/+ApoE^−/− or ATM^+/−ApoE^−/−) mice were treated with different dosages of chloroquine. Anti-CD25 antibody was used to deplete natural Tregs in ATM^+/+ApoE^−/− mice. The atherosclerotic burden in different groups of mice was comprehensively evaluated by H&E staining and Masson staining. The effect of chloroquine on the regulatory T cells (Tregs) was assessed in vivo and in vitro by flow cytometry and immunohistochemical staining. The expression of related proteins was detected by real-time polymerase chain reaction and western blotting.

Results In ATM^+/+ApoE^−/− mice, chloroquine alleviated atherosclerotic lesions, stabilized the plaque, and increased Treg counts in the atherosclerotic lesions and spleens. However, in ATM haploinsufficient mice (ATM^+/−ApoE^−/−), chloroquine no longer prevented atherosclerosis or impacted Treg counts. Abolishing Treg cells using an anti-CD25 antibody in vivo abrogated the atheroprotective effect of chloroquine. In vitro, chloroquine promoted the differentiation of Tregs from naïve T cells, which was accompanied by enhanced ATM/AMP-activated protein kinase (AMPK) activity and reduced downstream mammalian target of rapamycin (mTOR) activity.

Discussion These findings suggest that chloroquine ameliorates atherosclerosis and stabilizes plaque by modulating Tregs differentiation through the regulation of the ATM/AMPK/mTOR pathway.

背景临床观察表明氯喹及其衍生物具有动脉粥样硬化保护作用，但其机制尚不清楚。本研究旨在观察氯喹对动脉粥样硬化的保护作用并探讨其潜在机制。

方法用不同剂量的氯喹治疗共济失调毛细血管扩张突变 (ATM) 野生型或单倍剂量不足的载脂蛋白 E 敲除小鼠（ATM ^+/+ ApoE ^−/−或 ATM ^+/- ApoE ^{−/− ）。}抗 CD25 抗体用于消除 ATM ^+/+ ApoE ^−/−小鼠中的天然 Tregs。通过H&E染色和Masson染色综合评估不同组小鼠的动脉粥样硬化负荷。通过流式细胞术和免疫组织化学染色在体内和体外评估氯喹对调节性 T 细胞 (Treg) 的影响。通过实时聚合酶链反应和蛋白质印迹检测相关蛋白的表达。

结果在ATM ^+/+ ApoE ^−/−小鼠中，氯喹减轻了动脉粥样硬化病变，稳定了斑块，并增加了动脉粥样硬化病变和脾脏中的Treg 计数。然而，在 ATM 单倍体不足的小鼠 (ATM ^+/- ApoE ^−/− ) 中，氯喹不再预防动脉粥样硬化或影响 Treg 计数。在体内使用抗 CD25 抗体消除 Treg 细胞可以消除氯喹的动脉粥样硬化保护作用。在体外，氯喹促进Treg细胞从初始T细胞分化，同时增强ATM/AMP激活蛋白激酶（AMPK）活性并降低下游哺乳动物雷帕霉素靶点（mTOR）活性。

讨论这些发现表明，氯喹通过调节 ATM/AMPK/mTOR 通路来调节 Tregs 分化，从而改善动脉粥样硬化并稳定斑块。