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绿原酸抑制巨噬细胞激活的机制研究
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| 篇名： | 绿原酸抑制巨噬细胞激活的机制研究 |
| TITLE： | Mechanism study of chlorogenic acid alleviating macrophage activation |
| 摘要： | 目的 观察绿原酸对脂多糖（LPS）致巨噬细胞激活的影响，并探讨骨髓细胞2表达的触发受体（TREM2）蛋白在其中的作用。方法为筛选LPS造模浓度，分别以1、10、100ng/mL的LPS培养细胞24h，检测细胞培养上清液中白细胞介素6（IL-6）水平和细胞中诱导型一氧化氮合酶（iNOS）蛋白表达水平。为筛选绿原酸给药浓度，将细胞分为Control组、LPS处理组和3个不同浓度绿原酸（0.01、0.1、1μmol/L）干预组，检测细胞培养上清液中肿瘤坏死因子α（TNF-α）、IL-1β水平和细胞中iNOS、TREM2蛋白表达水平以及细胞活力。为观察TREM2在绿原酸抑制巨噬细胞激活中的作用，采用TREM2小干扰RNA（TREM2-siRNA）干扰细胞TREM2蛋白表达，将细胞分为Control组、LPS处理组、绿原酸+LPS组、TREM2-siRNA+绿原酸+LPS组和乱序-siRNA（SC-siRNA）+绿原酸+LPS组，在含有上述药物的培养基/空白培养基中孵育24h后，检测细胞培养上清液中TNF-α和IL-1β水平以及细胞中TREM2、iNOS和核因子κBp65（NF-κBp65）的蛋白表达水平。结果10ng/mLLPS可显著促进细胞释放IL-6，并增加细胞中iNOS蛋白表达水平，故选择该质量浓度LPS进行后续实验。与LPS处理组相比，0.1μmol/L绿原酸干预组细胞培养上清液中TNF-α、IL-1β水平和细胞中iNOS蛋白表达水平均显著降低（P＜0.05），细胞中TREM2蛋白表达水平显著升高（P＜0.05），细胞活力未受影响，故选择该浓度的绿原酸进行后续实验。在观察TREM2蛋白在绿原酸抑制巨噬细胞激活中的作用中发现，与Control组比较，LPS处理组细胞中iNOS和NF-κBp65蛋白表达水平均显著升高（P＜0.05）；与LPS处理组比较，绿原酸+LPS组细胞中iNOS和NF-κBp65蛋白表达水平均显著降低、TREM2蛋白表达水平显著升高（P＜0.05）；与绿原酸+LPS组比较，TREM2-siRNA+绿原酸+LPS组细胞中iNOS和NF-κBp65蛋白表达水平均显著升高、TREM2蛋白表达水平显著降低（P＜0.05），TREM2-siRNA明显逆转了绿原酸的上述作用；而SC-siRNA未对绿原酸产生的上述作用造成影响。结论绿原酸可显著抑制LPS对巨噬细胞的激活，其抗炎作用可能是通过TREM2蛋白介导的。 |
| ABSTRACT： | OBJECTIVE To observe the effects of chlorogenic acid on the activation of macrophage induced by lipopolysaccharide （LPS）， and to explore the role of triggering receptors expressed on myeloid cells-2 （TREM2） in the action. METHODS To find a suitable LPS concentration， the cells were cultured with 1， 10 and 100 ng/mL LPS for 24 h. The level of interleukin 6 （IL-6） in the cell culture supernatant and protein expression of inducible nitric oxide synthase （iNOS） in the cells were detected. To search for a suitable chlorogenic acid concentration， the cells were divided into control group， LPS group and three chlorogenic acid （0.01， 0.1 and 1 μmol/L）+LPS groups. The levels of tumor necrosis factor α （TNF-α） and IL-1β in the cell culture supernatant， the protein expressions of iNOS and TREM2 in the cells and cell viability were detected. To observe the effects of TREM2 in chlorogenic acid alleviating macrophage activation， TREM2-small interfering RNA （TREM2-siRNA） was taken to intervene in TREM2 protein expression. The cells were divided into control group， LPS group， chlorogenic acid+LPS group， TREM2-siRNA+chlorogenic acid+LPS group and SC-siRNA+chlorogenic acid+LPS group. After 24 h incubation， the levels of TNF- α and IL-1β in the cell culture supernatant and protein expressions of TREM2， iNOS and nuclear factor κB p65 （NF-κB p65） in the cells were detected. RESULTS 10 ng/mL LPS promoted IL-6 release and increased iNOS protein expression， and 10 ng/mL LPS was taken in the next experiments. Compared with the LPS group， 0.1 μmol/L chlorogenic acid decreased TNF-α jiaji1981@126.com and IL-1β levels， and down-regulated iNOS expression，meanwhile increased TREM2 expression without effect on cell viability， and 0.1 μmol/L chlorogenic acid was taken in the next experiments. Compared with the control group， the protein expressions of iNOS and NF- κB p65 in the LPS group were significantly increased （P＜0.05）； compared with the LPS group， the protein expressions of iNOS and NF- κB p65 in the chlorogenic acid+LPS group were significantly decreased， the protein expressions of TREM2 was significantly increased （P＜ 0.05）； compared with the chlorogenic acid+LPS group， the protein expressions of iNOS and NF-κB p65 of TREM2-siRNA+ chlorogenic acid+LPS group were significantly increased， the protein expressions of TREM2 was significantly decreased （P＜0.05）. TREM2-siRNA could significantly reverse the above effects of chlorogenic acid， while SC-siRNA did not significantly affect the above anti-inflammatory effects of chlorogenic acid. CONCLUSIONS Chlorogenic acid can inhibit the LPS-induced macrophage activation， and its anti-inflammatory may be mediated by TREM2 protein. |
| 期刊： | 2023年第34卷第21期 |
| 作者： | 郑薇；郎静；黄西凤；肖锐；白荷；贾济 |
| AUTHORS： | ZHENG Wei，LANG Jing，HUANG Xifeng，XIAO Rui，BAI He，JIA Ji |
| 关键字： | 绿原酸；巨噬细胞；脂多糖；炎症；骨髓细胞2表达的触发受体 |
| KEYWORDS： | chlorogenic acid； macrophage； lipopolysaccharide； inflammation； triggering receptors expressed on myeloid cells-2 |
| 阅读数： | 70 次 |
| 本月下载数： | 7 次 |
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