β极低密度脂蛋白经细胞外信号调节激酶信号途径诱导其受体表达上调

Very Low Density Lipoprotein Receptor Transcription by β-Very Low Density Lipoprotem is Mediated by the Extracellular Signal-Regulated Kinase Signaling Pathway

为探讨β极低密度脂蛋白诱导极低密度脂蛋白受体表达上调的信号转导途径及其对巨噬细胞胞内脂质堆积的影响,采用Western方法检测β极低密度脂蛋白温育后巨噬细胞内的细胞外信号调节激酶活性,在体外培养的巨噬细胞中加入不同信号途径关键激酶的抑制剂,观察各信号途径对受体调控的阻断效应,测定胞内胆固醇和甘油三酯的含量,观察泡沫细胞的形成。结果发现,β极低密度脂蛋白以蛋白激酶C依赖的方式激活巨噬细胞中的细胞外信号调节激酶1/2活性。细胞外信号调节激酶1/2抑制剂和蛋白激酶C抑制剂可显著阻断β极低密度脂蛋白诱导巨噬细胞极低密度脂蛋白受体转录的效应,而P38抑制剂和蛋白激酶C激动剂对β极低密度脂蛋白诱导极低密度脂蛋白受体表达上调无明显影响。细胞外信号调节激酶1/2抑制剂可抑制β极低密度脂蛋白诱导的巨噬细胞内胆固醇和甘油三酯含量升高。上述结果表明,蛋白激酶C依赖的细胞外信号调节激酶1/2信号级联可能是介导β极低密度脂蛋白诱导极低密度脂蛋白受体表达上调的主要信号转导途径,特异地抑制此信号途径可抑制β极低密度脂蛋白诱导的巨噬细胞胞内脂质堆积。

Aim To elucidate the intracellular signaling pathways for β-very low density lipoprotein (VLDL) -induced VLDLR transcription and their effects on lipid accumulation in macrophages. Methods Phosphorylated extracelcular signal regucated kinase (ERK1/2) protein was detected with Western blot. mRNA level of VLDL receptor was assayed by RT-PCR. Results We found that β-VLDL induced an increase in ERK1/2 activity in a protein kinase C (PKC)-dependent manner in murine RAW264.7 macrophages. Studies using different protein kinases inhibitors or activators showed that the effect of β-VLDL induced VLDL receptor transcription, which was monitored by RT-PCR analysis of VLDL receptor mRNA, was not affected by the inhibitor of P38 kinase and cAMP analog, but extremely abolished by pretreating cells with an inhibitor of ERK and an inhibitor of PKC. Studies on effects of PD98059 (the inhibitor of ERK) on β-VLDL-induced cellular lipid accumulation, which was assessed by cholesterol and triglycerid assay kit showed that ERK inhibitor decreased cellular cholestorol and triglycerid increase exposed to β-VLDL in a dose-dependent manner. Conclusions These results demonstrated that the PKC-ERK1/2 cascade was the essential signaling pathway by which β-VLDL activated VLDL receptor mRNA expression and inhibition of the ERK1/2 signaling cascade resulted in suppression of the cellular lipid accumulation induced by β-VLDL in macrophages.