脂多糖通过LDLr诱导巨噬细胞泡沫化:炎症应激下巨噬细胞泡沫化的另一条通路

发布时间：2018-05-21 03:26

本文选题：THP-巨噬细胞 + 脂多糖　；参考：《中国药理学通报》2013年08期

【摘要】：目的研究炎症介质-脂多糖(LPS)诱导的炎症应激是否通过扰乱固醇调节元件结合蛋白裂解激活蛋白-固醇调节元件结合蛋白2(SCAP-SREBP2)复合物介导的低密度脂蛋白受体(LDLr)负反馈调控,增加THP-1巨噬细胞对非修饰低密度脂蛋白胆固醇(LDL)摄取,导致泡沫细胞形成。方法诱导分化成功的THP-1巨噬细胞在无血清培养基中培养4 h后,分为对照组(继续无血清培养),高脂组(加入LDL负荷,终浓度为25 mg·L-1),高脂加炎症刺激组(加入LDL负荷,终浓度为25 mg·L-1,同时加入炎症介质LPS,终浓度200μg·L-1),单纯炎症刺激组(加入炎症介质LPS,终浓度200μg·L-1),以上各组细胞培养24 h后收获。ELISA法检测细胞培养基中TNF-α浓度,酶化学法检测细胞内胆固醇浓度,琼脂糖电泳检测LDL氧化程度,Real time PCR法检测LDLr、SREBP2和SCAP mRNA表达水平,Western blot法检测LDLr、SREBP2和SCAP蛋白表达,激光共聚焦检测SCAP在内质网与高尔基体间的转位情况。结果细胞内胆固醇测定发现,炎症介质LPS通过增加THP-1巨噬细胞对非修饰LDL摄取,导致巨噬细胞转变为泡沫细胞。在无炎症介质LPS时,25 mg·L-1LDL减少LDLr mRNA和蛋白表达(P0.05)。然而,LPS增加LDLr mRNA和蛋白表达,逆转25 mg·L-1LDL对LDLr的抑制效应,不恰当的增加了巨噬细胞对LDL摄取(P0.05),LPS刺激也导致了SCAP和SREBP2的mRNA和蛋白过表达(P0.05),同时促进了SCAP从内质网转移到高尔基体。这些结果提示LPS干扰了胆固醇介导的LDLr负反馈调控,使非修饰LDL在巨噬细胞内堆积,导致泡沫细胞形成。结论本研究提示,在LPS诱导的炎症应激状态条件下,天然LDL可以通过LDLr被巨噬细胞大量摄取入细胞内,导致泡沫细胞形成,这可能是巨噬细胞泡沫化的另一条通路。
[Abstract]:Objective to investigate whether the inflammatory stress induced by lipopolysaccharide (LPS) disrupts the negative feedback regulation of LDLrs mediated by the steroid regulatory element binding proteolytic activator-steroid regulatory element binding protein (2SCAP-SREBP2) complex. Increased uptake of unmodified low density lipoprotein cholesterol (LDL) by THP-1 macrophages resulted in foam formation. Methods the differentiated THP-1 macrophages were cultured in serum-free medium for 4 h and were divided into two groups: control group (continuous serum-free culture), hyperlipidemia group (adding LDL load, final concentration 25 mg / L), hyperlipidemia plus inflammatory stimulation group (adding LDL load). The final concentration was 25 mg / L, and the final concentration was 200 渭 g / L ~ (-1). The TNF- 伪 concentration in the cell culture medium was measured by Elisa after 24 hours of culture, and the final concentration was 200 渭 g / L ~ (-1) in the simple inflammatory stimulation group, and the final concentration was 200 渭 g / L ~ (-1). The intracellular cholesterol concentration was detected by enzyme method, the degree of oxidation of LDL by agarose electrophoresis and the expression level of SREBP2 and SCAP mRNA by real time PCR method. The expression of SREBP2 and SCAP protein was detected by Western blot method. The translocation of SCAP between endoplasmic reticulum and Golgi apparatus was detected by confocal laser. Results Intra-cellular cholesterol assay showed that the inflammatory mediator LPS increased the uptake of unmodified LDL by THP-1 macrophages and resulted in the transformation of macrophages into foam cells. In the absence of LPS, 25 mg L-1LDL decreased the expression of LDLr mRNA and protein (P 0.05). However, LPs increased the expression of LDLr mRNA and protein, reversed the inhibitory effect of 25 mg L-1LDL on LDLr, and improperly increased the uptake of LDL by macrophages. LPs also resulted in the overexpression of mRNA and protein in SCAP and SREBP2, and promoted the transfer of SCAP from endoplasmic reticulum to Golgi body. These results suggest that LPS interferes with the cholesterol-mediated negative feedback regulation of LDLr, resulting in the accumulation of unmodified LDL in macrophages and the formation of foam cells. Conclusion under the condition of inflammatory stress induced by LPS, natural LDL can be absorbed into the cells by macrophages through LDLr, leading to the formation of foam cells, which may be another pathway of macrophage foaming.
【作者单位】：泸州医学院附属医院心血管内科;重庆医科大学脂质研究中心重庆市糖脂代谢重点实验室;重庆医科大学附属第一院心血管内科;
【基金】：国家自然科学基金资助项目(No 30670869) 四川省卫生厅基金资助项目(No 2011-110335) 泸州市科技局基金资助项目[No 2011-I-S37(6/7)] 泸州医学院基金资助项目(No 2010-108) 泸州医学院附属医院基金资助项目(No2011-43)
【分类号】：R363

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