姚树桐, 赵莉, 苗成, 田华, 杨娜娜, 郭守东, 翟雷, 陈军, 王义围, 秦树存
泰山医学院1动脉粥样硬化研究所，山东省高校动脉粥样硬化重点实验室；基础医学院，泰安 271000；承德医学院附属医院，承德 067000
本文旨在研究内质网应激(endoplasmic reticulum stress, ERS)是否介导氧化低密度脂蛋白(oxidized low density lipoprotein, ox-LDL)所诱导的巨噬细胞清道夫受体A1 (scavenger receptor A1, SR-A1)上调。体外培养RAW264.7巨噬细胞，给予20 mmol/L ERS抑制剂4-苯丁酸(4-phenylbutyric acid, PBA)处理30 min后，再加入ox-LDL (50 mg/L)继续培养12 h或2 mg/L ERS诱导剂衣霉素(tunicamycin, TM)或2 μmol/L毒胡萝卜素(thapsigagin, TG)继续培养4 h。另外培养巨噬细胞分别给予0.5、1和2 mg/L TM处理4 h或给予2 mg/L TM 处理1、2和4 h。采用试剂盒检测细胞内总胆固醇(total cholesterol, TC)含量；分别采用免疫印迹法和实时定量聚合酶链反应(real-time PCR)技术检测SR-A1和ERS标志分子糖调节蛋白78 (glucose-regulated protein 78, GRP78)蛋白和mRNA表达变化；采用多功能酶标仪检测Dil-ox-LDL摄取情况。结果显示，PBA显著抑制ox-LDL所诱导的巨噬细胞内胆固醇蓄积。ox-LDL可显著上调SR-A1和GRP78表达，而PBA可明显抑制ox-LDL所诱导的SR-A1上调(P < 0.05)，并使GRP78降低39.3% (P = 0.057)。TM明显上调SR-A1蛋白表达，并促进巨噬细胞对ox-LDL的摄取，且呈浓度和时间依赖性，但对SR-A1转录水平没有明显影响，且ERS另一诱导剂TG也可明显上调SR-A1表达；而PBA则明显抑制TM和TG所诱导的上述变化。以上结果表明，ERS在ox-LDL所诱导的SR-A1上调中具有重要作用，进而促使巨噬细胞摄取更多的ox-LDL，导致泡沫细胞形成。
[Endoplasmic reticulum stress mediates oxidized low density lipoprotein-induced scavenger receptor A1 upregulation in macrophages.] [Article in Chinese]
YAO Shu-Tong, ZHAO Li, MIAO Cheng, TIAN Hua, YANG Na-Na , GUO Shou-Dong, ZHAI Lei, Chen Jun, WANG Yi-Wei, QIN Shu-Cun
Institute of Atherosclerosis, Key Laboratory of Atherosclerosis in Universities of Shandong； College of Basic Medical Sciences, Taishan Medical University, Tai'an 271000, China； Affiliated Hospital of Chengde Medical University, Chengde 067000, China
The present study was to investigate whether endoplasmic reticulum stress (ERS) was involved in oxidized low density
lipoprotein (ox-LDL)-induced scavenger receptor A1 (SR-A1) upregulation in macrophages. RAW264.7 cells were pretreated with 20 mmol/L of 4-phenylbutyric acid (PBA) for 30 min and then treated with ox-LDL (50 mg/L) for 12 h or stimulated with 2 mg/L tunicamycin (TM) or 2 μmol/L thapsigagin (TG) for 4 h. In addition, RAW264.7 cells were incubated with 0.5, 1 and 2 mg/L TM for 4 h or treated with 2 mg/L TM for 1, 2 and 4 h, respectively. The intracellular total cholesterol (TC) content was measured using a tissue/cell total cholesterol assay kit. The protein and mRNA expressions of SR-A1 and glucose-regulated protein 78 (GRP78) were analyzed by Western blot and real-time PCR, respectively. Dil-ox-LDL uptake was detected using a microplate reader. The results showed that ox-LDL-induced cholesterol accumulation in macrophages was attenuated by PBA, an ERS inhibitor. Ox-LDL caused significant SR-A1 upregulation with concomitant activation of ERS as assessed by upregulation of GRP78, whereas PBA significantly inhibited the ox-LDL-induced SR-A1 upregulation (P < 0.05) and slightly decreased GRP78 expression by 39.3% (P = 0.057). TM, an ERS inducer, upregulated SR-A1 protein expression and ox-LDL uptake in dose- and time-dependent manner, but had no significant effect on SR-A1 mRNA level. However, the TM- or TG-induced SR-A1 upregulation and ox-LDL uptake were significantly mitigated by PBA. These data indicate that ERS plays a critical role in ox-LDL-induced SR-A1 upregulation, which in turn enhances the foam cell formation by uptaking more ox-LDL.
通讯作者：王义围,秦树存 E-mail: email@example.com,firstname.lastname@example.org
姚树桐, 赵莉, 苗成, 田华, 杨娜娜, 郭守东, 翟雷, 陈军, 王义围, 秦树存. 内质网应激介导氧化低密度脂蛋白所诱导的巨噬细胞清道夫受体A1上调[J]. 生理学报 2014; 66 (5): 612-618.
YAO Shu-Tong, ZHAO Li, MIAO Cheng, TIAN Hua, YANG Na-Na , GUO Shou-Dong, ZHAI Lei, Chen Jun, WANG Yi-Wei, QIN Shu-Cun. [Endoplasmic reticulum stress mediates oxidized low density lipoprotein-induced scavenger receptor A1 upregulation in macrophages.] [Article in Chinese]. Acta Physiol Sin 2014; 66 (5): 612-618 (in Chinese with English abstract).