UNASSIGNED: Cannabidiol (CBD), a non-psychoactive phytocannabinoid of cannabis, is therapeutically used as an analgesic, anti-convulsant, anti-inflammatory, and anti-psychotic drug. There is a growing concern about the adverse side effects posed by CBD usage. Pregnane X receptor (PXR) is a nuclear receptor activated by a variety of dietary steroids, pharmaceutical agents, and environmental chemicals. In addition to the role in xenobiotic metabolism, the atherogenic and dyslipidemic effects of PXR have been revealed in animal models. CBD has a low affinity for cannabinoid receptors, thus it is important to elucidate the molecular mechanisms by which CBD activates cellular signaling and to assess the possible adverse impacts of CBD on pro-atherosclerotic events in cardiovascular system, such as dyslipidemia.
UNASSIGNED: Our study aims to explore the cellular and molecular mechanisms by which exposure to CBD activates human PXR and increases the risk of dyslipidemia.
UNASSIGNED: Both human hepatic and intestinal cells were used to test if CBD was a PXR agonist via cell-based transfection assay. The key residues within PXR\'s ligand-binding pocket that CBD interacted with were investigated using computational docking study together with site-directed mutagenesis assay. The C57BL/6 wildtype mice were orally fed CBD in the presence of PXR antagonist resveratrol (RES) to determine how CBD exposure could change the plasma lipid profiles in a PXR-dependent manner. Human intestinal cells were treated with CBD and/or RES to estimate the functions of CBD in cholesterol uptake.
UNASSIGNED: CBD was a selective agonist of PXR with higher activities on human PXR than rodents PXRs and promoted the dissociation of human PXR from nuclear co-repressors. The key amino acid residues Met246, Ser247, Phe251, Phe288, Trp299, and Tyr306 within PXR\'s ligand binding pocket were identified to be necessary for the agonistic effects of CBD. Exposure to CBD increased the circulating total cholesterol levels in mice which was partially caused by the induced expression levels of the key intestinal PXR-regulated lipogenic genes. Mechanistically, CBD induced the gene expression of key intestinal cholesterol transporters, which led to the increased cholesterol uptake by intestinal cells.
UNASSIGNED: CBD was identified as a selective PXR agonist. Exposure to CBD activated PXR signaling and increased the atherogenic cholesterol levels in plasma, which partially resulted from the ascended cholesterol uptake by intestinal cells. Our study provides potential evidence for the future risk assessment of CBD on cardiovascular disease, such as dyslipidemia.

Platinum (Pt)-based chemotherapy is the main treatment for ovarian cancer (OC); however, most patients develop Pt resistance (Pt-R). This work shows that Pt-R OC cells increase intracellular cholesterol through uptake via the HDL receptor, scavenger receptor type B-1 (SR-B1). SR-B1 blockade using synthetic cholesterol-poor HDL-like nanoparticles (HDL NPs) diminished cholesterol uptake leading to cell death and inhibition of tumor growth. Reduced cholesterol accumulation in cancer cells induces lipid oxidative stress through the reduction of glutathione peroxidase 4 (GPx4) leading to ferroptosis. In turn, GPx4 depletion induces decreased cholesterol uptake through SR-B1 and re-sensitizes OC cells to Pt. Mechanistically, GPx4 knockdown causes lower expression of the histone acetyltransferase EP300, leading to reduced deposition of histone H3 lysine 27 acetylation (H3K27Ac) on the sterol regulatory element binding transcription factor 2 (SREBF2) promoter and suppressing expression of this key transcription factor involved in the regulation of cholesterol metabolism. SREBF2 downregulation leads to decreased SR-B1 expression and diminished cholesterol uptake. Thus, chemoresistance and cancer cell survival under high ROS burden obligates high GPx4 and SR-B1 expression through SREBF2. Targeting SR-B1 to modulate cholesterol uptake inhibits this axis and causes ferroptosis in vitro and in vivo in Pt-R OC.

Aberrantly high dietary cholesterol intake and intestinal cholesterol uptake lead to dyslipidemia, one of the risk factors for cardiovascular diseases (CVDs). Based on previous studies, laminarin, a polysaccharide found in brown algae, has hypolipidemic activity, but its underlying mechanism has not been elucidated. In this study, we investigated the effect of laminarin on intestinal cholesterol uptake in vitro, as well as the lipid and morphological parameters in an in vivo model of high-fat diet (HFD)-fed mice, and addressed the question of whether Niemann-Pick C1-like 1 protein (NPC1L1), a key transporter mediating dietary cholesterol uptake, is involved in the mechanistic action of laminarin. In in vitro studies, BODIPY-cholesterol-labeled Caco-2 cells were examined using confocal microscopy and a fluorescence reader. The results demonstrated that laminarin inhibited cholesterol uptake into Caco-2 cells in a concentration-dependent manner (EC50 = 20.69 μM). In HFD-fed C57BL/6J mice, laminarin significantly reduced the serum levels of total cholesterol (TC), total triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C). It also decreased hepatic levels of TC, TG, and total bile acids (TBA) while promoting the excretion of fecal cholesterol. Furthermore, laminarin significantly reduced local villous damage in the jejunum of HFD mice. Mechanistic studies revealed that laminarin significantly downregulated NPC1L1 protein expression in the jejunum of HFD-fed mice. The siRNA-mediated knockdown of NPC1L1 attenuated the laminarin-mediated inhibition of cholesterol uptake in Caco-2 cells. This study suggests that laminarin significantly improves dyslipidemia in HFD-fed mice, likely by reducing cholesterol uptake through a mechanism that involves the downregulation of NPC1L1 expression.

Yamanashi et al., conducted a study on the absorption of cholesterol and β-sitosterol, as well as the inhibitory effect of ezetimibe (EZE). They used CaCo-2 cells to simulate the intestines and investigated how different mixed micelles, acting as carriers, were absorbed into these cells through the Niemann-Pick C1-like 1 (NPC1L1) protein. The study focused on the impact of micelle shape, size, and zeta potential on absorption and the inhibitory effect of EZE. I utilized small-angle X-ray scattering and a zeta potential measuring device to measure these characteristics. The findings revealed a two-step mechanism: NPC1L1 selectively bound micelles based on their shape and size, and once bound, the absorption was regulated by the molecular structure of the micelle components. EZE\'s inhibitory effect changed with micelle composition, influencing micelle size and shape. EZE initially acted on the micelle\'s shape and size, and then NPC1L1 selectively bound micelles based on their shape and size, allowing EZE to directly inhibit absorption by interacting with NPC1L1. This groundbreaking discovery challenges existing concepts and holds significant implications for researchers in drug development, as well as physicians and pharmacists.

The impact of JAK/STAT inhibitors, which are used in various inflammatory diseases, on cardiovascular risk is controversial and has recently raised safety concerns. Our study investigates the direct effects of tofacitinib on macrophage cholesterol metabolism, which is crucial for atherosclerosis plaque development and stability. Cultured human macrophages THP-1 were used to assess the impact of tofacitinib on cell cholesterol efflux and synthesis via radioisotopic methods, and on cholesterol uptake by measuring the cell cholesterol content with a fluorometric assay. The cholesterol acceptors and donors were either standard lipoproteins or sera from patients with juvenile idiopathic arthritis (JIA) and from control subjects. Tofacitinib significantly increased the macrophage cholesterol efflux to all acceptors; it reduced cholesterol uptake from both the normal and hypercholesterolemic sera; and it reduced cholesterol synthesis. The treatment of macrophages with tofacitinib was able to increase the cholesterol efflux and decrease cholesterol uptake when using sera from untreated JIA patients with active disease as cholesterol acceptors and donors, respectively. In conclusion, our in vitro data support the concept that tofacitinib has a favorable impact on macrophage cholesterol metabolism, even in the presence of sera from rheumatologic patients, and suggest that other mechanisms may be responsible for the cardiovascular risk associated with tofacitinib use in selected patient populations.

In this study, we evaluated the effects of Lactobacillus reuteri NCIMB (LRC™) supplementation on hypercholesterolemia by researching its effects on cellular cholesterol metabolism in hypercholesterolemic rats (KHGASP-22-170) and HepG2 cell line. Rats were separated into six groups after adaptation and were then fed a normal control (NC), a high-cholesterol diet (HC), or a HC supplemented with simvastatin 15 mg/kg body weight (positive control [PC]), LRC 1 × 109 colony-forming units (CFU)/rat/day, LRC 4 × 109 CFU/rat/day, or LRC 1 × 1010 CFU/rat/day (1 × 109, 4 × 109, or 1 × 1010). The rats were dissected to study the effects of LRC on cholesterol metabolism and intestinal excretion at the end of experimental period. We discovered that LRC mainly participated in the restraint of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, the uptake of low-density lipoprotein (LDL) cholesterol into tissues, partially in the transport of cholesteryl esters into high density lipoprotein for maturation, and intestinal excretion of cholesterol. These results are supported by the expression of transcription factors and enzymes such as HMG-CoA reductase, SREBP2, CYP7A1, CETP, and LCAT in both messenger RNA (mRNA) and protein levels in serum and hepatic tissue. Furthermore, the LRC treatment in HepG2 significantly reduced the mRNA expression of HMG-CoA reductase, SREBP2, and CEPT and significantly increased the mRNA expression of LDL-receptor, LCAT, and CYP7A1 at all doses. Hence, we suggest that LRC supplementation could alleviate the serum cholesterol level by inhibiting the intracellular cholesterol synthesis, and augmenting excretion of intestinal cholesterol.

Disordered lipid accumulation in the arterial wall is a hallmark of atherosclerosis. Previous studies found that the expression of triggering receptor expressed on myeloid cells 2 (TREM2), a transmembrane receptor of the immunoglobulin family, is increased in mouse atherosclerotic aortic plaques. However, it remains unknown whether TREM2 plays a role in atherosclerosis. Here we investigated the role of TREM2 in atherosclerosis using ApoE knockout (ApoE-/-) mouse models, primary vascular smooth muscle cells (SMCs), and bone marrow-derived macrophages (BMDMs). In ApoE-/- mice, the density of TREM2-positive foam cells in aortic plaques increased in a time-dependent manner after the mice were fed a high-fat diet (HFD). Compared with ApoE-/- mice, the Trem2-/-/ApoE-/- double-knockout mice showed significantly reduced atherosclerotic lesion size, foam cell number, and lipid burden degree in plaques after HFD feeding. Overexpression of TREM2 in cultured vascular SMCs and macrophages exacerbates lipid influx and foam cell formation by upregulating the expression of the scavenger receptor CD36. Mechanistically, TREM2 inhibits the phosphorylation of p38 mitogen-activated protein kinase and peroxisome proliferator activated-receptor gamma (PPARγ), thereby increasing PPARγ nuclear transcriptional activity and subsequently promoting the transcription of CD36. Our results indicate that TREM2 exacerbates atherosclerosis development by promoting SMC- and macrophage-derived foam cell formation by regulating scavenger receptor CD36 expression. Thus, TREM2 may act as a novel therapeutic target for the treatment of atherosclerosis.

BACKGROUND: Cholesterol plays a vital role in multiple physiological processes. Cellular uptake of cholesterol is mediated primarily through endocytosis of low-density lipoprotein (LDL) receptor. New modifiers of this process remain to be characterized. Particularly, the role of fasting- and CREB-H-induced (FACI) protein in cholesterol homeostasis merits further investigation.
METHODS: Interactome profiling by proximity labeling and affinity purification - mass spectrometry was performed. Total internal reflection fluorescence microscopy and confocal immunofluorescence microscopy were used to analyze protein co-localization and interaction. Mutational analysis was carried out to define the domain and residues required for FACI localization and function. Endocytosis was traced by fluorescent cargos. LDL uptake in cultured cells and diet-induced hypercholesterolemia in mice were assessed.
RESULTS: FACI interacted with proteins critically involved in clathrin-mediated endocytosis, vesicle trafficking, and membrane cytoskeleton. FACI localized to clathrin-coated pits (CCP) on plasma membranes. FACI contains a conserved DxxxLI motif, which mediates its binding with the adaptor protein 2 (AP2) complex. Disruption of this motif of FACI abolished its CCP localization but didn\'t affect its association with plasma membrane. Cholesterol was found to facilitate FACI transport from plasma membrane to endocytic recycling compartment in a clathrin- and cytoskeleton-dependent manner. LDL endocytosis was enhanced in FACI-overexpressed AML12 cells but impaired in FACI-depleted HeLa cells. In vivo study indicated that hepatic FACI overexpression alleviated diet-induced hypercholesterolemia in mice.
CONCLUSIONS: FACI facilitates LDL endocytosis through its interaction with the AP2 complex.

Atherosclerosis is a significant risk factor for coronary heart disease (CHD) and myocardial infarction (MI). Atherosclerosis develops during foam cell generation, which is caused by an imbalance in cholesterol uptake, esterification, and efflux. LOX-1, SR-A1, and CD36 all increased cholesterol uptake. ACAT1 and ACAT2 promote free cholesterol (FC) esterification to cholesteryl esters (CE). The hydrolysis of CE to FC was aided by nCEH. FC efflux was promoted by ABCA1, ABCG1, ADAM10, and apoA-I. SR-BI promotes not only cholesterol uptake but also FC efflux. Circular RNAs (circRNAs), which are single-stranded RNAs with a closed covalent circular structure, have emerged as promising biomarkers and therapeutic targets for atherosclerosis due to their highly tissue, cell, and disease state-specific expression profiles. Numerous studies have shown that circRNAs regulate foam cell formation, acting as miRNA sponges to influence atherosclerosis development by regulating the expression of SR-A1, CD36, ACAT2, ABCA1, ABCG1, ADAM10, apoA-I, SR-B1. Several circRNAs, including circ-Wdr91, circ 0004104, circRNA0044073, circRNA_0001805, circDENND1B, circRSF1, circ 0001445, and circRNA 102682, are potential biomarkers for atherosclerosis to better evaluate cardiovascular risk. It is difficult to deliver synthetic therapeutic circRNAs to the desired target tissues. Nanotechnology, such as GA-RM/GZ/PL, may be an important solution to this problem. In this review, we focus on the potential role and mechanism of circRNA/miRNA axis in foam cell formation in the hopes of discovering new targets for the diagnosis, prevention, and treatment of atherosclerosis.

Isoliquiritigenin (ISL) is a flavonoid with a chalcone structure extracted from the natural herb Glycyrrhiza glabra. Its anti-inflammatory, antibacterial, antioxidant, and anticancer activities have been extensively studied. Moreover, ISL also possess hypolipidemic and atherosclerosis-reducing effects. However, its cholesterol-lowering mechanisms have not been reported yet. Niemann Pick C1 Like 1 (NPC1L1) is a specific transporter of cholesterol uptake. In this study, we found for the first time that ISL downregulates NPC1L1 expression and competitively inhibits cellular cholesterol uptake by binding to NPC1L1 in a concentration-dependent manner in vitro. This study provides a theoretical basis for further investigation of the molecular mechanisms of its cholesterol-lowering effect in vivo and inspired emerging drug research for cholesterol-lowering purposes through NPC1L1 inhibition.