目的 考察在热休克蛋白（HSP）65攻击下养阴方对载脂蛋白E敲除（ApoE^-/-）小鼠动脉粥样硬化（AS）的病变及辅助性T细胞17（Th17）、调节性T细胞（Treg）的影响及可能机制。方法 将ApoE^-/-小鼠分别饲喂普通和高脂饲料，并分别sc无菌PBS或HSP65，以辛伐他汀为阳性药，研究养阴方对AS模型小鼠血脂、斑块、Th17细胞与Treg细胞比例（Th17/Treg）及相关炎症指标的影响。给药方式皆为ig给药，并以等体积生理盐水做对照。采用酯酶法检测血清总胆固醇（TC）、三酰甘油（TG）、低密度脂蛋白（LDL）及高密度脂蛋白（HDL）水平；油红O染色法测定主动脉斑块面积；流式细胞术测定抗凝血中IL-17A⁺CD4⁺ Th17及Foxp3⁺CD25⁺CD4⁺ Treg细胞数量；ELISA法测定血清白细胞介素-6（IL-6）、IL-17、IL-2、转化生长因子-β（TGF-β）、IL-10水平；实时荧光定量PCR（qRT-PCR）法测定脾脏中Stat3、Stat5a、Foxp3和主动脉中Stat3、Stat5a mRNA表达；Western blotting法测定脾脏中磷酸化STAT3（pSTAT3）、磷酸化STAT5（pSTAT5）、FOXP3和主动脉中pSTAT3蛋白表达。结果 养阴方使小鼠血清TC、TG、LDL水平显著降低（P<0.01），HDL显著升高（P<0.05）；主动脉斑块面积明显减小（P<0.01）；外周血IL-17A⁺CD4⁺ Th17细胞显著减少（P<0.05），Foxp3⁺CD25⁺CD4⁺ Treg显著增加（P<0.05），Th17/Treg值大幅下降（P<0.01）；血清IL-6水平显著下降（P<0.05），IL-2（P<0.01）、TGF-β（P<0.05）、IL-10（P<0.05）水平显著升高；脾脏pSTAT3蛋白水平下降（P<0.01），pSTAT5（P<0.01）和FOXP3（P<0.05）蛋白水平上升；主动脉Stat3 mRNA（P<0.01）和pSTAT3蛋白（P<0.05）水平下降。结论 养阴方能够改善HSP65攻击下ApoE^-/-小鼠的高脂血症及AS，同时调节外周血中Th17/Treg的平衡和炎症因子的分泌从而改善炎症。养阴方对Th17、Treg的效应可能与减弱的IL-6/pSTAT3和增强的IL-2/pSTAT5信号有关。

Objective To investigate the effects of Yangyin Recipe on atherosclerosis (AS) lesions, helper T cells 17 (Th17), and regulatory T cells (Treg) in apolipoprotein E knockout (ApoE^-/-) mice under the heat shock protein (HSP) 65 attack, and explore the potential mechanisms. Methods ApoE^-/- mice were fed with normal and high-fat diets respectively, and injected with sterile PBS or HSP65. Simvastatin was used as the positive drug to study the effects of Yangyin Recipe on blood lipids, plaques, Th17/Treg cells and related inflammation indicators in AS model mice. Drugs were all intragastric administrated with equal volume of normal saline as a control. Serum total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL) and high-density lipoprotein (HDL) were detected by esterase method; The area of aortic plaque was determined by oil red O staining; The amounts of IL-17A⁺CD4⁺ Th17 and Foxp3⁺CD25⁺CD4⁺ Treg cells in anticoagulated blood were determined by flow cytometry; Serum IL-6, IL-17, IL-2, TGF-β, and IL-10 were determined by ELISA; mRNA expression of Stat3, Stat5a, Foxp3 in spleens and Stat3, Stat5a in aortas were detected by qPCR; Protein expression of phosphorylated STAT3 (pSTAT3), phosphorylated STAT5 (pSTAT5), FOXP3 in spleen, and pSTAT3 in aortas were determined by Western blotting. Results Under the influence of Yangyin Recipe, serum TC, TG, LDL levels decreased (P < 0.01), and HDL increased (P < 0.05); Aortic plaque area decreased remarkably (P < 0.01); IL-17A⁺CD4⁺Th17 cells in peripheral blood decreased (P < 0.05), Foxp3⁺CD25⁺CD4⁺ Treg increased (P < 0.05), and Th17/Treg ratio decreased drastically (P < 0.01); Serum IL-6 decreased (P < 0.05), IL-2 (P < 0.01), TGF-β and IL-10 (P < 0.05) increased; Protein levels of pSTAT3 in spleens decreased (P < 0.01), but pSTAT5 (P < 0.01) and FOXP3 (P < 0.05) increased; Aortic Stat3 mRNA (P < 0.01) and pSTAT3 (P < 0.05) protein decreased.Conclusion Yangyin Recipe can improve hyperlipidemia and AS in ApoE^-/- mice during HSP65 attack, and regulate the balance of Th17/Treg in peripheral blood and the secretion of inflammatory factors to improve inflammation, which may be related to attenuate IL-6/pSTAT3 and enhance IL-2/pSTAT5 signaling.

国家自然科学基金资助项目（81573929）；国家自然科学基金资助项目（81373232）；江苏省自然科学基金资助项目（BK2012458）