%0 Journal Article %T 高磷环境下胆固醇敏感器SCAP功能失调促进巨噬细胞内胆固醇蓄积 %A 刘巧 %A 欧阳南 %A 何泉 %A 周超 %J 第二军医大学学报 %D 2017 %R 10.16781/j.0258-879x.2017.12.1514 %X 目的 观察高磷环境对巨噬细胞内胆固醇蓄积的影响及其分子机制。方法 将人单核细胞株（THP-1）源性巨噬细胞分为对照组（磷1.0 mmol/L）、高磷处理组（磷3.0 mmol/L）、磷甲酸钠（PFA）处理组（磷1.0 mmol/L加PFA 1.0 mmol/L）以及高磷联合PFA处理组（磷3.0 mmol/L加PFA 1.0 mmol/L），按不同要求分别处理各组细胞。培养24 h后，油红O染色观察细胞内中性脂质的分布，酶催化比色法定量测定细胞内的胆固醇含量，qPCR检测细胞内羟甲基戊二酸单酰辅酶A还原酶（HMGCoAR）、低密度脂蛋白受体（LDLR）、固醇调节元件结合蛋白裂解激活蛋白（SCAP） mRNA的表达，蛋白质印迹法测定细胞内SCAP、LDLR、HMGCoAR及核内固醇调节元件结合蛋白2（N-SREBP2）的蛋白水平，激光共聚焦法检测SCAP从内质网向高尔基体的移位情况。结果 与对照组相比，高磷处理组巨噬细胞内中性脂质明显聚集，细胞内总胆固醇与胆固醇酯的含量增加（P<0.05），LDLR、HMGCoAR mRNA与蛋白的表达增高（P<0.05，P<0.01），细胞核内N-SREBP2的蛋白表达水平增加（P<0.05），SCAP从内质网向高尔基体的移位增加；PFA处理（高磷联合PFA处理组）可阻断高磷引起的上述作用（P<0.05，P<0.01）。高磷处理组巨噬细胞内SCAP的蛋白水平相比对照组增高（P<0.05），PFA处理能抑制高磷导致的SCAP蛋白水平增高，但SCAP mRNA的表达在各组间差异均无统计学意义（P>0.05）。结论 高磷环境下，钠磷转运体介导磷离子进入巨噬细胞内，通过基因转录后机制增加SCAP的蛋白水平，诱导其功能失调并促使其异常转运SREBP2至高尔基体裂解释放N-SREBP2，后者转位入核促进HMGCoAR和LDLR的表达，促使细胞内源性胆固醇合成和外源性LDL经LDLR摄入的增加，最终导致细胞内胆固醇异常蓄积。</br>Objective To explore the effects of high phosphate (Pi) condition on the cholesterol accumulation in macrophage and its underlying mechanisms. Methods The human monocytic cell line THP-1 derived macrophages were divided into control group (concentration of Pi being 1.0 mmol/L), high Pi exposure group (concentration of Pi being 3.0 mmol/L), phosphonoformic acid (PFA) treatment group (concentration of Pi and PFA being 1.0 mmol/L) and high phosphate plus PFA treatment group (concentration of Pi and PFA being 3.0 mmol/L and 1.0 mmol/L, respectively). Intracellular neutral lipids were observed by Oil Red O staining. Cholesterol contents were quantified by enzyme catalyzed colorimetry. qPCR was used to dectect the relative mRNA expressions of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMGCoAR), low density lipoprotein receptor (LDLR) and sterol regulatory element binding protein (SREBP) cleavage activating protein (SCAP), and their proteins and nuclear SREBP 2 (N-SREBP2) protein expressions were assessed by Western blotting. The translocation of SCAP from endoplasmic reticulum (ER) to Golgi body was observed by laser confocal microscope. Results Compared with the control group, the macrophages in the high Pi exposure group showed an obvious aggregation of neutral lipids and significantly increased contents intracellular cholesterol ester and total cholesterol (P<0.05). The mRNA and protein expressions of LDLR and HMGCoAR, and protein level of N-SREBP2 in macrophage nucleus in the high Pi exposure group were increased significantly in comparison to the control group (P<0.05, P<0.01). The SCAP translocated obviously from ER to Golgi body. However, above-mentioned changes were significantly %K 巨噬细胞 高磷酸盐血症 胆固醇贮积病 动脉粥样硬化< %K /br> %K macrophage hyperphosphatemia cholesterosis atherosclerosis %U http://www.ajsmmu.cn/ajsmmu/ch/reader/view_abstract.aspx?file_no=20170231&flag=1