The Effects of the oxLDL/β2GPI/anti‐β2GPI Complex on Macrophage Autophagy and its Mechanism

ABSTRACT Background Previous research has established that the oxidized low‐density lipoprotein/β2‐glycoprotein I/anti‐β2‐glycoprotein I antibody (oxLDL/β2GPI/anti‐β2GPI) complex can stimulate macrophages to secrete molecules associated with atherosclerosis (AS), such as monocyte chemotactic protein 1 (MCP‐1), tissue factor (TF), and tumor necrosis factor‐α (TNF‐α). This complex also enhances the uptake of oxLDL, thereby accelerating foam cell formation through the Toll‐like receptor‐4/nuclear factor kappa B (TLR4/NF‐κB) pathway. Given the critical role of macrophage autophagy in the instability of vulnerable atherosclerotic plaques, it is imperative to investigate whether the oxLDL/β2GPI/anti‐β2GPI complex influences macrophage autophagy in AS. This study aims to elucidate the effects and underlying mechanisms of the oxLDL/β2GPI/anti‐β2GPI complex on macrophage autophagy in AS. Methods Experiments were conducted using murine macrophage RAW264.7 cells and the human monocytic cell line THP‐1. Western blot analysis was employed to determine the expressions of autophagy‐associated markers and signaling pathway proteins. Autophagosomes were detected through mRFP‐GFP‐LC3 adenoviral transfection and transmission electron microscopy (TEM). Results Treatment of macrophages with the oxLDL/β2GPI/anti‐β2GPI complex resulted in decreased expressions of Beclin1 and LC3 proteins, alongside an upregulation of SQSTM1/P62 protein expression. Additionally, there was a reduction in the number of autophagosomes and autolysosomes. An increase in the phosphorylation levels of phosphoinositide‐3‐kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) was also observed. Notably, the expressions of autophagy‐associated markers were partially restored when the TLR4/NF‐κB and PI3K/AKT/mTOR pathways were inhibited by their respective inhibitors. Conclusions Our findings indicate that the oxLDL/β2GPI/anti‐β2GPI complex inhibits macrophage autophagy in AS via the TLR4/NF‐κB and PI3K/AKT/mTOR signaling pathways.