Dysfunction of cholesterol sensor SCAP promotes inflammation activation in THP-1 macrophages
Dysfunction of cholesterol sensor SCAP promotes inflammation activation in THP-1 macrophages
复制标题
胆固醇传感器SCAP功能障碍促进THP-1巨噬细胞炎症激活
DOI：
10.1016/j.yexcr.2018.03.032
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发表时间：
2018-06-15
影响因子：
3.7
通讯作者：
Zhou, Chao
中科院分区：
文献类型：
--
作者：
Ouyang, Nan;Gan, Hua;Zhou, Chao
血脂异常和慢性炎症之间存在交叉，两者都是动脉粥样硬化的促发因素。固醇调节元件结合蛋白裂解激活蛋白（SCAP）在调节胆固醇稳态中起关键作用。本研究探讨了SCAP功能障碍对THP-1巨噬细胞炎性细胞因子表达和脂质代谢的影响。细胞内胆固醇含量通过油红0染色和定量测定来评估。实时荧光定量RT-PCR和Western blotting检测细胞核SCAP、HMGCR、pro-IL-1 β、N-SREBP 2、p65（N）的表达。通过ELISA测定上清液中分泌蛋白IL-1 β、TNF-α和MCP-1的水平。共聚焦显微镜观察到SCAP从内质网（ER）向高尔基体的转位。我们的结果表明，SCAP的过表达显著增加了HMGCR，pro-IL-1 β在细胞质中的表达，以及成熟IL-1 β，TNF-α，MCP-1在上清液中的表达，而敲低SCAP则显著降低了这些分子的表达。桦木醇有效地抑制了SCAP过表达的THP-1巨噬细胞中细胞内胆固醇的积累，但不影响炎性细胞因子的表达，表明SCAP的促炎作用与其调节胆固醇稳态的常规作用无关。此外，我们研究了介导血脂异常和炎症反应之间串扰的分子机制。敲低SCAP可减弱LPS诱导的I κ B磷酸化并降低p65的核水平，而过表达SCAP可增加p65的核水平。敲低p65消除了由SCAP过表达的THP-1巨噬细胞中炎性介质表达升高所代表的促炎作用，表明SCAP功能障碍通过激活NF-κ B信号通路刺激炎症反应。总之，胆固醇传感器SCAP在调节THP-1巨噬细胞中炎症因子如IL-1 β、TNF-α和MCP-1的表达中起作用。SCAP通过激活NF-κ B通路介导炎症反应。SCAP的这种新功能与其在脂质代谢中的作用无关。
Crosstalk occurs between dyslipidemia and chronic inflammation, which are both precipitants of atherosclerosis. Sterol regulatory element binding proteins cleavage-activating protein (SCAP) plays a key role in regulating cholesterol homeostasis. The present study investigated the effects of SCAP dysfunction on the expression of inflammatory cytokines and lipid metabolism in THP-1 macrophages. Intracellular cholesterol content was assessed by Oil Red 0 staining and quantitative assays. The expression of SCAP, HMGCR, pro-IL-1 beta and N-SREBP2, p65(N) in the nucleus were examined by real-time quantitative RT-PCR and Western blotting. The level of secretary proteins IL-1 beta, TNF-alpha and MCP-1 in the supernatants were determined by ELISA. The translocation of SCAP from the endoplasmic reticulum (ER) to the Golgi was detected by confocal microscopy. Our results demonstrated that over-expression of SCAP significantly increased the expression of HMGCR, pro-IL-1 beta in the cytoplasm, and mature IL-1 beta, TNF-alpha, MCP-1 in the supernatants, while knocking down SCAP dramatically decreased the expression of these molecules. Betulin effectively suppressed the accumulation of intracellular cholesterol in the SCAP over-expressed THP-1 macrophages, but did not affect the expression of inflammatory cytokines, indicating that the pro-inflammatory effect of SCAP was independent of its routine role in regulating cholesterol homeostasis. Furthermore, we investigated the molecular mechanisms mediating the crosstalk between dyslipidemia and inflammatory responses. Knocking down SCAP attenuated LPS-induced I kappa B phosphorylation and reduced the nuclear level of p65, while over-expression of SCAP increased the nuclear level of p65. Knocking down p65 abolished the proinflammatory effect represented by elevated expression of the inflammatory mediators in the SCAP over-expressed THP-1 macrophages, suggesting that SCAP dysfunction stimulated inflammatory responses via activating the NF-kappa B signaling pathway. In conclusion, the cholesterol sensor SCAP plays a role in regulating the expression of inflammatory factors such as IL-1 beta, TNF-alpha, and MCP-1 in THP-1 macrophages. SCAP mediates the inflammatory response via activating the NF-kappa B pathway. This new function of SCAP is independent of its role in lipid metabolism.