Objective: This study aims to explore the effect of lipid-induced macrophage M1/M2 polarization on lipid metabolism in hepatocytes. Methods: RAW264.7 macrophages were incubated with different kinds of fatty acids including saturated fatty acids-palmitic acid (PA), monounsaturated fatty acids-oleic acid (OA) and polyunsaturated fatty acids-docosahexaenoic acid (DHA), and cell culture supernatants were collected to prepare conditioned medium (CM). Hepatocytes were isolated by in situ perfusion of the liver with collagenase in mice, and a macrophage-hepatocyte CM co-culture system was established. Macrophage M1/M2 phenotype markers were detected by Real-time PCR. Lipid synthesis and decomposition related mRNA and protein expressions in hepatocytes were detected by Real-time PCR and Western Blot. Lipid depositions in hepatocytes were detected by oil red O staining. An analysis of variance was used for comparison of means between multiple groups. Results: Compared with control groups, PA polarized macrophages to a M1 phenotype (expression of TNF-α and IL-6 significantly increased, F≥22.68, P < 0.01), OA polarized macrophages to a M1/M2 mixed phenotype (expression of IL-6, Mrc2 and IL-10 increased F≥4.94, P < 0.05) and DHA polarized macrophages to a M2 phenotype (expression of Mrc2 and IL-10 significantly increased, F≥4.94, P < 0.01). CM-PA significantly increased lipid synthesis related genes, including SREBP1C, ACC1 mRNA expression (F≥5.66, P < 0.01) and FASN, ACC1 protein expression (F≥38.34, P < 0.05) in hepatocytes, and decreased lipid decomposition gene ACOX1 protein expression (F=154.48, P < 0.01). CM-OA affected several lipid metabolism genes expression. CM-DHA significantly increased CPT1A mRNA expression (F = 10.30, P < 0.01) and ACOX1, CPT1A protein expression (F≥47.06, P < 0.05), and decreased SREBP1C, ACC1 protein expression (F≥65.84, P < 0.05) in hepatocytes. Massive lipid droplets were deposited in hepatocytes in CM-PA treated hepatocytes, and a few amount of lipid droplets were deposited in CM-DHA treated hepatocytes. Conclusion: Different fatty acids affect the balance of lipid metabolism in hepatocytes and liver by inducing macrophage M1 / M2 polarization, thus promoting or delaying the progression of non-alcoholic fatty liver disease.
目的： 探讨脂质诱导的巨噬细胞M1/M2型极化对肝细胞脂质代谢的影响。 方法： 分别以饱和脂肪酸——棕榈酸（PA）、单不饱和脂肪酸——油酸（OA）和多不饱和脂肪酸——二十二碳六烯酸（DHA）孵育巨噬细胞RAW264.7，并制备条件培养液（CM）。胶原酶原位灌注法分离C57BL/6小鼠肝细胞，体外巨噬细胞CM培养原代肝细胞。Real-time PCR检测巨噬细胞M1/M2型极化基因表达，以Real-time PCR和Western blot检测肝细胞脂质合成和分解代谢相关基因的mRNA和蛋白表达，油红O染色检测肝细胞内脂质沉积情况。多组样本数据间比较采用单因素方差分析，两两比较采用t检验。 结果： 与对照组相比，PA诱导巨噬细胞M1型极化[肿瘤坏死因子α、白细胞介素（IL）-6 mRNA表达水平显著升高，F≥22.68，P值均< 0.01]，OA诱导巨噬细胞M1/M2型混合极化[IL-6、甘露糖受体c2（Mrc2）和IL-10 mRNA表达水平升高，F≥4.94，P值均< 0.05]，而DHA则诱导巨噬细胞M2型极化（Mrc2、IL-10 mRNA表达水平增加，F≥4.94，P值均< 0.01）。CM-PA显著增加肝细胞脂质合成基因固醇调节元件结合蛋白1C（SREBP1C）、乙酰辅酶A羧化酶1（ACC1）mRNA表达（F≥5.66, P值均< 0.01）和脂肪酸合酶、ACC1蛋白表达（F≥38.34, P值均< 0.05），并降低脂质分解蛋白脂酰辅酶A氧化酶1（ACOX1）表达（F = 154.48，P < 0.01），促进肝细胞内大量脂滴沉积；CM-OA影响肝细胞部分脂质代谢基因表达；而CM-DHA显著增加肝细胞脂质分解基因肉碱棕榈酰转移酶1A（CPT1A）mRNA表达（F = 10.30，P < 0.01）和ACOX1、CPT1A蛋白表达（P值均< 0.05），并降低脂质合成蛋白SREBP1C、ACC1的表达（F≥65.84，P值均< 0.05），CM-DHA孵育的肝细胞内见少量脂滴沉积。 结论： 不同脂肪酸通过诱导巨噬细胞M1/M2型极化改变影响肝细胞和肝脏脂质代谢平衡，从而促进或延缓非酒精性脂肪性肝病的进展。.
Keywords: Fatty acid; Fatty liver; Lipid metabolism; Macrophage polarization.