Center for Drug Research and Development, Guangdong Pharmaceutical University, Guangzhou, Guangdong, China.
Guangdong Provincial Key Laboratory of Advanced Drug Delivery Systems and Guangdong Provincial Engineering Center of Topical Precise Drug Delivery System, Guangdong Pharmaceutical University, Guangzhou, Guangdong, China.
PLoS One. 2023 Aug 15;18(8):e0290051. doi: 10.1371/journal.pone.0290051. eCollection 2023.
To explore the metabolomics of fatty acids and biological information of related markers in a RAW264.7 cell inflammation model. RAW264.7 macrophage inflammation model was induced by LPS, and RAW264.7 cells were treated with non-steroidal anti-inflammatory drugs (NSAIDs). The fatty acid compositions were identified by GC-MS, combined with standard product spectrum information and NIST (National Institute of Standards and Technology) database. Using chemometrics and Analysis of Variance (ANOVA), the components with VIP > 1 and P < 0.05 were selected as significant difference markers, and combined with biological methods to explore the biological significance of them. GC-MS identified 21 fatty acids in RAW264.7 cells, and screened significant difference biomarkers in each group. Among these biomarkers, C20:5 and C22:6 had significant changes in pairwise comparison among each group. Through ELISA, polymerase chain reaction (PCR) and Western Blot methods, the mRNA and protein expressions of IL-1β, NLRP3, GPR120 and β-Arrestin-2 were up-regulated after RAW264.7 cells induced by LPS and nigericin, and decreased after drug intervention. It indicated that the signal pathway centered on NLRP3 inflammasome was involved in the anti-inflammatory process of ibuprofen. It was the first time to study fatty acid metabolomics in RAW264.7 cell inflammatory model by GC-MS combined with chemometrics. The anti-inflammatory mechanism of ibuprofen was explained from NLRP3 inflammasome perspective without precedent, which enriched the research on the signal pathway of ibuprofen anti-inflammatory mechanism.
探讨 RAW264.7 细胞炎症模型中脂肪酸的代谢组学和相关标志物的生物学信息。采用 LPS 诱导 RAW264.7 巨噬细胞炎症模型，给予非甾体类抗炎药（NSAIDs）处理 RAW264.7 细胞。采用 GC-MS 结合标准品谱图信息和 NIST（美国国家标准与技术研究院）数据库对脂肪酸成分进行鉴定。利用化学计量学和方差分析（ANOVA），选择 VIP>1 和 P<0.05 的成分作为差异显著标志物，并结合生物学方法，探索其生物学意义。GC-MS 鉴定了 RAW264.7 细胞中的 21 种脂肪酸，并筛选了各组中的差异显著标志物。在这些标志物中，C20:5 和 C22:6 在各组间的两两比较中均有显著变化。通过 ELISA、聚合酶链反应（PCR）和 Western blot 方法，发现 LPS 和 Nigericin 诱导 RAW264.7 细胞后，IL-1β、NLRP3、GPR120 和 β-Arrestin-2 的 mRNA 和蛋白表达上调，药物干预后表达降低。表明以 NLRP3 炎性小体为中心的信号通路参与了布洛芬的抗炎过程。这是首次采用 GC-MS 结合化学计量学方法研究 RAW264.7 细胞炎症模型中的脂肪酸代谢组学，从 NLRP3 炎性小体角度解释了布洛芬的抗炎机制，丰富了布洛芬抗炎机制信号通路的研究。
