Atherosclerosis (AS) is the most common cardiovascular disease, and reverse cholesterol transport (RCT) plays an important role in maintaining cholesterol homeostasis. Both endoplasmic reticulum (ER) stress and LXRα can affect the metabolism of cholesterol. However, whether ER stress can modulate cholesterol metabolism by LXRα in hepatocytes and macrophages remains unclear. Therefore, in this study, we aimed to explore the relationship between ER stress induced by tunicamycin and LXRα in hepatocytes and macrophages and clarify their possible mechanisms and roles in AS. C57BL/6 mice and Huh-7 and THP-1 cells were treated with tunicamycin and LXR-623 (an agonist of LXRα) alone or in combination. Tunicamycin-induced ER stress caused liver injury; promoted the accumulation of cholesterol and triglycerides; inhibited the expression of LXRα, ABCA1 and ABCG1 in the livers of mice, thus reducing serum high-density lipoprotein (HDL)-C, low-density lipoprotein (LDL)-C, total cholesterol and triglyceride levels; however, LXR-623 could attenuate ER stress and reverse these changes. We also obtained the same results in Huh-7 and THP-1 cells. ER stress induced by tunicamycin could clearly be reversed by activating LXRα because it promoted cholesterol efflux by enhancing the expression of ABCA1 and ABCG1 in hepatocytes and macrophages, contributing to attenuation of the development of AS.

Cholesterol metabolism affects endoplasmic reticulum (ER) stress and modulates epithelial-mesenchymal transition (EMT). Our previous study demonstrated that 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) attenuated EMT by blocking the transforming growth factor (TGF)-β/Smad signaling pathway and activating ER stress in MDA-MB-231 cells. To further assess the detailed mechanisms between cholesterol metabolism, ER stress, and EMT, LXR-623 (an agonist of LXRα) and simvastatin were used to increase and decrease cholesterol efflux and synthesis, respectively. Here, we found that high HP-β-CD concentrations could locally increase cholesterol levels in the ER by decreasing LXRα expression and increasing Hydroxymethylglutaryl-Coenzyme A reductase (HMGCR) expression in MDA-MB-231 and BT-549 cells, which triggered ER stress and inhibited EMT. Meanwhile, tunicamycin-induced ER stress blocked the TGF-β/Smad signaling pathway. However, low HP-β-CD concentrations can decrease the level of membrane cholesterol, enhance the TGF-β receptor I levels in lipid rafts, which helped to activate TGF-β/Smad signaling pathway, inhibit ER stress and elevate EMT. Based on our findings, the use of high HP-β-CD concentration can lead to cholesterol accumulation in the ER, thereby inducing ER stress, which directly suppresses TGF-β pathway-induced EMT. However, HP-β-CD is proposed to deplete membrane cholesterol at low concentrations and concurrently inhibit ER stress and induce EMT by promoting the TGF-β signaling pathways.

Neutrophil-mediated inflammation plays a crucial role in the pathogenesis of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). Dysregulated neutrophil death has been implicated in promoting neutrophil priming and activation. PANoptosis, an inflammatory programmed cell death, participates in the development of various autoimmune diseases. The current study aims to identify the key PANoptosis-related marker and investigate its association with disease activity of AAV. Gene microarray data of patients with ANCA-associated glomerulonephritis (ANCA-GN) was downloaded from the Gene Expression Omnibus (GEO). PANoptosis-associated genes were sourced from GeneCards. The hub-marker of PANoptosis-related genes was identified by integrated bioinformatical analysis. The single-cell sequencing data from ANCA-GN mice were used to determine the location of hub genes. Then levels of activated CD11b on neutrophils from peripheral blood, as well as soluble CD11b in urine and plasma were measured in AAV patients. Their associations with clinicopathological parameters of AAV patients were subsequently analyzed. Co-localization of CD11b and neutrophils in renal specimens of AAV patients was evaluated by immunofluorescence staining. Changes of intracellular metabolic pathways of neutrophils from AAV patients were further investigated. A small molecule inhibitor was used to explore the effect of intracellular fatty acid metabolism (FAO) on neutrophil CD11b activation and PANoptosis. CD11B was identified as the pivotal PANoptosis-related gene in AAV by integrated bioinformatics analysis. The level of activated CD11b on neutrophils was significantly higher in active AAV patients compared with healthy controls, and elevated neutrophil CD11b levels positively correlated with disease activity, as evidenced by higher levels of hypersensitive C-reactive protein (hCRP), increased Birmingham Vasculitis Activity Score (BVAS), elevated serum creatinine levels at sampling and a higher proportion of cellular crescents in renal specimens of AAV patients. We also demonstrated the presence of neutrophil CD11b in renal specimen of AAV patients. Moreover, along with elevated Cd11b expression, neutrophils from ANCA-GN mice exhibited an aberrant FAO phenotype and a down-regulated peroxisome proliferators-activated receptor (PPAR)-α pathway, as compared with healthy controls. GW7647, a PPARα agonist, could alleviate AAV serum-induced neutrophil CD11b activation and PANoptosis. CD11B was a pivotal PANoptosis-related gene in AAV. The level of activated CD11b on neutrophils was associated with disease activity in AAV patients.