Plexin D1 negatively regulates macrophage-derived foam cell migration via the focal adhesion kinase/Paxillin pathway.

BACKGROUND: Macrophage-derived foam cell formation is a hallmark of atherosclerosis and is retained during plaque formation. Strategies to inhibit the accumulation of these cells hold promise as viable options for treating atherosclerosis. Plexin D1 (PLXND1), a member of the Plexin family, has elevated expression in atherosclerotic plaques and correlates with cell migration; however, its role in macrophages remains unclear. We hypothesize that the guidance receptor PLXND1 negatively regulating macrophage mobility to promote the progression of atherosclerosis. METHODS: We utilized a mouse model of atherosclerosis based on a high-fat diet and an ox-LDL- induced foam cell model to assess PLXND1 levels and their impact on cell migration. Through western blotting, Transwell assays, and immunofluorescence staining, we explored the potential mechanism by which PLXND1 mediates foam cell motility in atherosclerosis. RESULTS: Our study identifies a critical role for PLXND1 in atherosclerosis plaques and in a low-migration capacity foam cell model induced by ox-LDL. In the aortic sinus plaques of ApoE-/- mice, immunofluorescence staining revealed significant upregulation of PLXND1 and Sema3E, with colocalization in macrophages. In macrophages treated with ox-LDL, increased expression of PLXND1 led to reduced pseudopodia formation and decreased migratory capacity. PLXND1 is involved in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK. Additionally, FAK inhibitors counteract the ox-LDL-induced migration suppression by modulating the phosphorylation states of FAK, Paxillin and their downstream effectors CDC42 and PAK. CONCLUSION: Our findings indicate that PLXND1 plays a role in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK to promoting atherosclerosis.

Emodin relieves morphine-stimulated BV2 microglial activation and inflammation through the TLR4/NF-κB/NLRP3 pathway.

The objective of this study is to disclose the role of emodin, a natural anthraquinone derivative that has been proposed to suppress microglial activation and inflammation, in morphine tolerance. Here, cell counting kit-8 method assayed the viability of BV2 microglial cells treated by ascending concentrations of emodin. In emodin-pretreated BV2 microglial cells challenged with morphine with or without transfection of toll-like receptor 4 (TLR4) overexpression plasmids, transwell assay measured cell migration. Immunofluorescence staining and western blot detected the expression of microglial markers. Inflammatory levels were subjected to ELISA and western blot. BODIPY 581/591 C11 assay estimated lipid reactive oxygen species activity. Iron assay kit examined total iron content. Western blot tested the expression of ferroptosis- and TLR4/nuclear factor-kappaB (NF-κB)/NOD-like receptor 3 (NLRP3) pathway-associated proteins. Molecular docking predicted the binding affinity of emodin to TLR4. Emodin was noted to obstruct the migration, activation, inflammatory response, and ferroptosis of BV2 microglial cells induced by morphine. In addition, emodin had a high binding affinity with TLR4 and inactivated TLR4/NF-κB/NLRP3 pathway in morphine-challenged BV2 microglial cells. Upregulation of TLR4 partially countervailed the protective role of emodin against morphine-elicited BV2 microglial cell migration, activation, inflammation, and ferroptosis. Accordingly, emodin might target TLR4 and act as an inactivator of TLR4/NF-κB/NLRP3 pathway, thus inhibiting BV2 microglial activation and inflammation to mitigate morphine tolerance.