College of Veterinary Medicine, Northwest A&F University, Yangling, Xianyang 712100, China.
Int J Mol Sci. 2022 Nov 24;23(23):14659. doi: 10.3390/ijms232314659.
The purpose of this article is to study the effects and mechanism of miR-4796 in the process of ophiopogon polysaccharide liposome (OPL) regulation of the immune activity of Kupffer cells (KCs). In this study, KCs were used as cell models, and were treated with OPL in different concentrations after being transfected with miR-4796 mimic or miR-4796 inhibitor. Firstly, the secretion of NO and iNOS, phagocytic activity, the expression of surface molecules CD14 and MHC II, apoptosis and ROS secretion were measured by Griess, flow cytometry, fluorescence staining and ELISA. Then, real-time PCR and Western blot were used to measure the expression of TLR4, IKKβ, MyD88 and NF-κB in the TLR4-NF-κB signaling pathway. The results showed that after transfection with miR-4796 mimic, the secretion of NO and iNOS, cell migration, cell phagocytosis and expression levels of CD14 and MHC II in the OPL group were significantly higher than those in the miR-4796 mimic control group (p < 0.05; p < 0.01). In addition, the mRNA and protein expression levels of TLR4, MyD88 and NF-κB were significantly higher than those in miR-4796 mimic control group (p < 0.05; p < 0.01). After transfection with miR-4796 inhibitor, the secretion of NO and iNOS, cell migration, cell phagocytosis, expression of CD14 and MHCII in OPL group were significantly higher than those in the miR-4796 inhibitor control group (p < 0.05; p < 0.01). These results indicated that OPL could regulate the immune activity of KCs by regulating miR-4796 and activating the TLR4-NF-κB signaling pathway.
本文旨在研究 miR-4796 在麦冬多糖脂质体（OPL）调节枯否细胞（KCs）免疫活性过程中的作用及机制。本研究以 KCs 为细胞模型，转染 miR-4796 模拟物或 miR-4796 抑制剂后，用不同浓度的 OPL 处理细胞。首先，通过 Griess 法、流式细胞术、荧光染色和 ELISA 法检测 NO 和 iNOS 的分泌、吞噬活性、表面分子 CD14 和 MHC II 的表达、细胞凋亡和 ROS 的分泌。然后，采用实时 PCR 和 Western blot 法检测 TLR4-NF-κB 信号通路中 TLR4、IKKβ、MyD88 和 NF-κB 的表达。结果表明，转染 miR-4796 模拟物后，OPL 组 NO 和 iNOS 的分泌、细胞迁移、细胞吞噬和 CD14、MHC II 的表达水平均显著高于 miR-4796 模拟物对照组（p＜0.05；p＜0.01）。此外，TLR4、MyD88 和 NF-κB 的 mRNA 和蛋白表达水平也明显高于 miR-4796 模拟物对照组（p＜0.05；p＜0.01）。转染 miR-4796 抑制剂后，OPL 组 NO 和 iNOS 的分泌、细胞迁移、细胞吞噬、CD14 和 MHC II 的表达水平均显著高于 miR-4796 抑制剂对照组（p＜0.05；p＜0.01）。这些结果表明，OPL 可通过调节 miR-4796 激活 TLR4-NF-κB 信号通路来调节 KCs 的免疫活性。
