| 周垂杨,柯乐斌,高仁贤.miR-519d-3p调控TLR4抑制脂多糖所致THP-1源性巨噬细胞炎症的机制研究[J].浙江中西医结合杂志,2022,32(1): |
| miR-519d-3p调控TLR4抑制脂多糖所致THP-1源性巨噬细胞炎症的机制研究 |
| Study on the mechanism of miR-519d-3p regulating TLR4 to inhibit lipopolysaccharide-induced inflammation of THP-1-derived macrophages |
| 投稿时间：2021-06-28 修订日期：2021-12-05 |
| DOI： |
| 中文关键词: 巨噬细胞 炎症 |
| 英文关键词:miR-519d-3p TLR4 |
| 基金项目:温州市基础性医疗卫生科技项目(Y2020521) |
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| 中文摘要: |
| 目的 探究miR-519d-3p在脂多糖（Lipopolysaccharide，LPS）诱导的巨噬细胞炎症中的作用和分子机制。方法 用PMA诱导THP-1细胞分化为巨噬细胞，LPS刺激细胞产生炎症反应作为后续实验的细胞模型。在该细胞模型中，应用荧光定量PCR检测miR-519d-3p的表达；利用antago-miR-519d-3p及antago-NC或者miR-519d-3p mimic及mimic-NC分别转染细胞，构建细胞内miR-519d-3p低表达或者过表达的THP-1巨噬细胞系，siRNA构建细胞内TLR4表达敲低模型；利用Elisa检测细胞上清内炎症因子的分泌水平，western blot检测各细胞系中TLR4、p-p65、T-p65、p-IκBα、T-IκBα蛋白水平。结果 LPS诱导的THP-1源巨噬细胞炎症模型中miR-519d-3p的表达与对照组相比明显升高（P＜0.05）；抑制miR-519d-3p表达显著抑制细胞中炎症因子TNF-α、IL-6、IL-1β分泌（P＜0.05），同时降低p-p65、p-IκBα蛋白水平。进一步研究发现，抑制miR-519d-3p能明显抑制LPS刺激的巨噬细胞中TLR4表达，在TLR4敲低的细胞模型中的研究表明miR-519d-3p通过TLR4调节炎症反应。结论 miR-519d-3p通过调控TLR4表达参与LPS诱导的巨噬细胞炎症反应，进而可能在急性肺损伤中发挥重要作用。 |
| 英文摘要: |
| Objective To explore the role and molecular mechanism of miR-519d-3p in lipopolysaccharide (LPS)-induced macrophage inflammation. Methods Firstly, PMA was used to stimulate THP-1 cells to differentiate into macrophages as a cell model for subsequent experiments. LPS was added to THP-1 derived macrophages to stimulate 12 h to construct a macrophage inflammation model, and the expression of miR-519d-3p was detected by fluorescence quantitative PCR; using antago-miR-519d-3p and antago-NC or miR-519d -3p mimic and mimic-NC were transfected into cells to construct THP-1 macrophages that inhibited miR-519d-3p expression or overexpressed miR-519d-3p, and siRNA constructed intracellular TLR4 expression knockout model; Elisa Detection of miR-519d-3p expression inhibition cell supernatant, western blot detection of TLR4, p-p65, T-p65, p-IκBα, T-IκBα protein levels in each cell line; immunofluorescence detection of TLR4 expression in cells distributed. Results The expression of miR-519d-3p in THP-1 macrophages induced by LPS was significantly higher than that in the control group (P<0.05); inhibition of miR-519d-3p expression significantly inhibited TNF-α, IL-6, IL-1β secretion in the LPS induced macrophages (P<0.05), while reducing p-p65 and p-IκBα protein levels. Further studies have found that inhibiting miR-519d-3p can significantly inhibit the expression of TLR4 in macrophages induced by LPS, furthermore, TLR4 knockdown model proves that miR-519d-3p regulate inflammation via TLR4. Conclusion miR-519d-3p can regulate of LPS-induced inflammatory response in macrophages through TLR4, and may play an important role in acute lung injury. |
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