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目的:探讨THP-1巨噬细胞泡沫化过程中NO/PKG的变化,以及NO/PKG对THP-1巨噬细胞ATP结合盒转运体A1(ABCA1)基因mRNA表达和胆固醇含量的影响。方法:THP-1细胞诱导分化为巨噬细胞,用氧化低密度脂蛋白(ox-LDL)处理,油红“O”染色观察巨噬细胞脂滴,酶标仪测定一氧化氮(NO)的释放;用一氧化氮供体L-精氨酸和硝普钠(SNP)、乙二醇二乙醚二胺四乙酸(EGTA)、PKG激动剂处理巨噬细胞,RT-PCR法测定细胞ABCA1基因mRNA的表达,高效液相色谱法(HPLC)测定细胞内胆固醇的含量变化。结果:ox-LDL可导致THP-1巨噬细胞脂质的蓄积和NO释放的增加,实验组NO释放量较对照组显著升高(P<0.05),实验组中50mg/L组、100mg/L组、200mg/L组与25mg/L组相比显著升高(P<0.05)。用PKG激动剂处理巨噬细胞后促进ABCA1基因mRNA表达上调(P<0.05);用PKG激动剂、L-精氨酸、SNP处理后的巨噬细胞内胆固醇较对照组显著减少(P<0.05)。EGTA能显著增加细胞内胆固醇含量(P<0.05)。结论:NO/PKG能降低细胞内胆固醇,其机制可能是PKG激活后通过上调ABCA1的表达,减少细胞内胆固醇的蓄积。
AIM: To investigate the changes of NO / PKG and the effect of NO / PKG on THP-1 macrophage ATP-Binding Cassette Transporter A1 (ABCA1) gene mRNA expression and cholesterol content in THP-1 macrophages. Methods: THP-1 cells were induced to differentiate into macrophages. Lipoproteins were detected by ox-LDL staining and lipid red staining, and nitric oxide (NO) was measured by microplate reader ); L-arginine and sodium nitroprusside (SNP), ethylene glycol diethylenediaminetetraacetic acid (EGTA) and PKG agonist were used to treat macrophages, and the cells were determined by RT-PCR ABCA1 gene mRNA expression, high performance liquid chromatography (HPLC) determination of changes in intracellular cholesterol content. Results: Ox-LDL could induce the accumulation of lipid and NO release in THP-1 macrophages, and the NO release in experimental group was significantly higher than that in control group (P <0.05) L group, 200mg / L group and 25mg / L group was significantly higher (P <0.05). After treated with PKG agonist, the mRNA expression of ABCA1 was up-regulated (P <0.05). The levels of cholesterol in macrophages treated with PKG agonist, L-arginine and SNP were significantly lower than those in control group ). EGTA significantly increased intracellular cholesterol (P <0.05). CONCLUSION: NO / PKG can decrease intracellular cholesterol. The mechanism may be that PKC increases the expression of ABCA1 and decreases the accumulation of intracellular cholesterol after PKG activation.