IL-35下调CD36表达减少巨噬细胞内脂质积累缓解动脉粥样硬化的机制研究

IL-35 alleviates atherosclerosis by reducing intramacrophage lipid accumulation via CD36 downregulation

摘要:

目的 分析冠状动脉粥样硬化性心脏病（CHD）患者血清白细胞介素-35（IL-35）水平与血脂四项的相关性，探究IL-35抑制人髓系白血病单核细胞（THP-1）泡沫化机制。方法 选取2022年6月—2023年6月在泰州人民医院就诊的疑似CHD患者154例，分为对照组和CHD组，空腹采血后用酶联免疫吸附试验检测两组血清IL-35水平，分析与血脂四项的相关性。采用油红染色和胆固醇检测试剂盒对比IL-35刺激THP-1细胞前后细胞内胆固醇含量变化；Western blotting检测CD36、SR-A、Lox-1、p38、p-p38蛋白的表达；实时荧光定量聚合酶链反应检测CD36、SR-A和Lox-1 mRNA的表达。结果 CHD组血清IL-35水平低于对照组（P <0.05）。CHD患者血清IL-35水平与TC、LDL-C、TG呈负相关（r_s =-0.321、-0.218和-0.215，P =0.003、0.044和0.047），与HDL-C呈正相关（r_s =0.322，P =0.003）。对照组与实验组THP-1细胞2、4、6、8和10 h的荧光强度（FI）比较，结果 ①不同时间点FI比较，差异有统计学意义（F =726.726，P =0.000）；②实验组与对照组FI比较，差异有统计学意义（F =8.102，P =0.012），实验组FI较低；③两组FI变化趋势比较，差异有统计学意义（F =111.061，P =0.000）。oxLDL组和实验组CD36、SR-A、Lox-1蛋白和mRNA相对表达量均高于对照组和IL-35组（P <0.05），实验组CD36蛋白和mRNA相对表达量低于oxLDL组（P <0.05）。oxLDL组和实验组p-p38/p38蛋白相对表达量均高于对照组和IL-35组（P <0.05），实验组p-p38/p38蛋白相对表达量低于oxLDL组（P <0.05）。oxLDL组、实验组、P79350组CD36蛋白相对表达量均高于对照组（P <0.05），oxLDL组和P79350组CD36蛋白相对表达量均高于实验组（P <0.05）。结论 CHD患者血清IL-35与血脂四项有相关性，IL-35可能通过抑制p38 MAPK信号通路介导巨噬细胞CD36表达下调，减少细胞内脂质。

Abstract:

Objective To analyze the correlations between the serum level of interleukin-35 (IL-35) and levels of blood lipids in patients with coronary heart disease (CHD), and to investigate the mechanism underlying the inhibition of transformation from THP-1-derived macrophages into foam cells via IL-35.Methods The 154 patients with suspected CHD in Taizhou People's Hospital from June 2022 to June 2023 were enrolled and divided into the control group and the CHD group. The serum level of IL-35 in each group was determined by enzyme-linked immunosorbent assay (ELISA) in the condition of fasting, and its correlations with blood lipid profiles were analyzed. The change of the cholesterol content in THP-1 cells before and after IL-35 stimulation was determined by oil red O staining and the cholesterol detection kit. The protein expressions of CD36, SR-A, Lox-1, p38 and p-p38 were measured by Western blotting, and mRNA expressions of CD36, SR-A and Lox-1 were detected by quantitative real-time polymerase chain reaction.Results Compared with the control group, the serum level of IL-35 in the CHD group was lower (P < 0.05). The serum level of IL-35 in CHD patients was negatively correlated with levels of total cholesterol (r_s = -0.321, P = 0.003), low-density lipoprotein cholesterol (r_s = -0.218, P = 0.044) and triglyceride (r_s = -0.215, P = 0.047), and positively correlated with the level of high-density lipoprotein cholesterol (r_s = 0.322, P = 0.003). The FI of THP-1 cells in the control group and the experimental group at 2 h, 4 h, 6 h, 8 h, and 10 h was compared, and the results indicated that FI was different among the time points (F = 726.726, P = 0.000) and between the groups (F = 8.102, P = 0.012), where FI in the experimental group was lower than that in the control group. Besides, the change trends of FI were different between the two groups (F = 111.061, P = 0.000). The protein and mRNA expressions of CD36, SR-A and Lox-1 in the oxLDL group and the experimental group were higher than those in the control group and the IL-35 group (P < 0.05). The protein and mRNA expressions of CD36 in the experimental group were lower than those in the oxLDL group (P < 0.05). The relative protein expression of p-p38 to that of p38 (p-p38/p38) in the oxLDL group and the experimental group was higher than that in the control group and the IL-35 group (P < 0.05), while p-p38/p38 in the experimental group was lower than that in the oxLDL group (P < 0.05). The relative protein expression of CD36 in the oxLDL group, experimental group and P79350 group was higher than that in the control group (P < 0.05), and that in the oxLDL group and the P79350 group was higher than that in the experimental group (P < 0.05).Conclusions Serum IL-35 is correlated with the blood lipid profiles in CHD patients. Besides, IL-35 may mediate the downregulation of CD36 on macrophages and the decrease in intramacrophage lipid accumulation by inhibiting the p38 MAPK signaling pathway.