DFMG attenuates the activation of macrophages induced by co‑culture with LPC‑injured HUVE‑12 cells via the TLR4/MyD88/NF‑κB signaling pathway

Cong,Li; Yang,Shuting; Zhang,Yong; Cao,Jianguo; Fu,Xiaohua

doi:10.3892/ijmm.2018.3511

DFMG attenuates the activation of macrophages induced by co‑culture with LPC‑injured HUVE‑12 cells via the TLR4/MyD88/NF‑κB signaling pathway

Authors:
- Li Cong
- Shuting Yang
- Yong Zhang
- Jianguo Cao
- Xiaohua Fu
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Affiliations: College of Medicine, Hunan Normal University, Changsha, Hunan 410013, P.R. China, Center for Molecular Medicine, Xiangya Hospital, Collaborative Innovation Center for Cancer Medicine, Central South University, Changsha, Hunan 410078, P.R. China
Published online on: February 23, 2018 https://doi.org/10.3892/ijmm.2018.3511
Pages: 2619-2628
Copyright: © Cong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

7‑difluoromethoxy‑5,4'‑dimethoxy‑genistein (DFMG) is a novel active chemical entity, which modulates the function and signal transduction of endothelial cells and macrophages (MPs), and is essential in the prevention of atherosclerosis. In the present study, the activity and molecular mechanism of DFMG on MPs was investigated using a Transwell assay to construct a non‑contact co‑culture model. Human umbilical vein endothelial cells (HUVE‑12), which were incubated with lysophosphatidylcholine (LPC), were seeded in the upper chambers, whereas PMA‑induced MPs were grown in the lower chambers. The generation of reactive oxygen species (ROS) and the release of lactate dehydrogenase (LDH) were measured using the corresponding assay kits. The proliferation and migration were assessed using 3‑(4,5‑dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazolium bromide and wound healing assays, respectively. Foam cell formation was examined using oil red O staining and a total cholesterol assay. The protein expression levels of Toll‑like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear factor (NF)‑κB p65 were detected by western immunoblotting. The secretion of interleukin (IL)‑1β was examined using an enzyme‑linked immunosorbent assay. It was found that LPC significantly increased the generation of ROS and the release of LDH in HUVE‑12 cells. The LPC‑injured HUVE‑12 cells activated MPs under co‑culture conditions and this process was inhibited by DFMG treatment. LPC upregulated the expression levels of TLR4, MyD88 and NF‑κB p65, and the secretion of IL‑1β in the supernatant of the co‑cultured HUVE‑12 cells and MPs. These effects were reversed by the application of DFMG. Furthermore, CLI‑095 and IL‑1Ra suppressed the activation of MPs that was induced by co‑culture with injured HUVE‑12 cells. These effects were further enhanced by co‑treatment with DFMG, and DFMG exhibited synergistic effects with a TLR4‑specific inhibitor. Take together, these findings revealed that DFMG attenuated the activation of MP induced by co‑culture with LPC‑injured HUVE‑12 cells. This process was mediated via inhibition of the TLR4/MyD88/NF‑κB signaling pathway in HUVE‑12 cells.

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