Journal of Lipid Research最新文献

High-density lipoprotein (HDL)-free cholesterol (FC) transfers to other lipoproteins and cells, the former by a spontaneous mechanism and the latter by both spontaneous and receptor-mediated mechanisms. Macrophages are an important cell type in all stages of atherosclerotic cardiovascular disease (ASCVD), and the magnitude of FC efflux from macrophages to HDL, a metric of HDL function, inversely associated with several metrics of ASCVD. Very high plasma HDL concentrations are associated with increased all-cause and ASCVD mortality, suggesting that the reverse process, FC influx from HDL into macrophages, is atherogenic. We hypothesize that HDL-FC is a metric of dysfunctional HDL, and when combined with HDL particle number (HDL-P), is an ASCVD risk factor. The magnitude of FC influx from HDL to macrophages is expected to be a function of HDL-P and HDL-FC content. Here we show that plasma HDL-FC content varies 2-fold among normolipidemic human subjects and linearly correlates with low-density lipoprotein (LDL)-FC content. The influx of HDL-FC into macrophages and transfer to LDL increase linearly with HDL-FC. As expected, the influx of HDL-FC into macrophages and the transfer to LDL are positively correlated. These data support the hypothesis that high HDL FC content is a marker for dysfunctional HDL, resulting in greater influx into macrophages and greater HDL-FC transfer to LDL. HDL-FC transfer to LDL is a valid surrogate for influx into macrophages. This study of HDL composition and function of normolipidemic subjects provides the basis for further investigation and establishment of HDL-FC content as an ASCVD risk factor.

Ceramides are key components of the skin's permeability barrier. In atopic dermatitis, pathological hydrolysis of ceramide precursors - glucosylceramides and sphingomyelin - into lysosphingolipids, specifically glucosylsphingosine (GS) and sphingosine-phosphorylcholine (SPC), and free fatty acids (FFAs) has been proposed to contribute to impaired skin barrier function. This study investigated whether replacing ceramides with lysosphingolipids and FFAs in skin lipid barrier models would exacerbate barrier dysfunction. When applied topically to human stratum corneum sheets, SPC and GS increased water loss, decreased electrical impedance, and slightly disordered lipid chains. In lipid models containing isolated human stratum corneum ceramides, reducing ceramides by ≥ 30% significantly increased permeability to four markers, likely due to loss of long-periodicity phase (LPP) lamellae and phase separation within the lipid matrix, as revealed by X-ray diffraction and infrared spectroscopy. However, when the missing ceramides were replaced by lysosphingolipids and FFAs, no further increase in permeability was observed. Conversely, these molecules partially mitigated the negative effects of ceramide deficiency, particularly with 5%-10% SPC, which reduced permeability even compared to control with "healthy" lipid composition. These findings suggest that while ceramide deficiency is a key factor in skin barrier dysfunction, the presence of lysosphingolipids and FFAs does not aggravate lipid structural or functional damage, but may provide partial compensation, raising further questions about the behavior of lyso(sphingo)lipids in rigid multilamellar lipid environments, such as the stratum corneum, that warrant further investigation.

Microsomal triglyceride transfer protein (MTP) plays crucial roles in the assembly and secretion of apolipoprotein B-containing lipoproteins and loss of function MTP variants are associated with abetalipoproteinemia, a disease characterized by the absence of these lipoproteins. MTP is a heterodimeric protein of two subunits, MTP and protein disulfide isomerase (PDI). In this study, we report a proband with abetalipoproteinemia who was monitored annually for 10 years in her third decade and had very low plasma lipids and undetectable apoB-containing lipoproteins. Genetic testing revealed biallelic variants in the MTTP gene. She has a well-documented nonsense mutation Gly865∗ that does not interact with the PDI subunit. She also has a novel missense MTP mutation, Ile344Asn. We show that this mutation abrogates lipid transfer activity in MTP and does not support apolipoprotein B secretion. This residue is present in the central α-helical domain of MTP and the substitution of Ile with Asn at this position disrupts interactions between MTP and PDI subunits. Ile344 is away from the known MTP:PDI interacting sites identified in the crystal structure of MTP suggesting that MTP:PDI interactions are more dynamic than previously envisioned. Identification of more missense mutations will enhance our understanding of the structure-function of MTP and the role of critical residues in these interactions between the two subunits. This knowledge may guide us in developing novel treatment modalities to reduce plasma lipids and atherosclerosis.

Enzymatically oxygenated phospholipids (eoxPL) from lipoxygenases (LOX) or cyclooxygenase (COX) are prothrombotic. Their generation in arterial disease, and their modulation by cardiovascular therapies is unknown. Furthermore, the Lands cycle acyl-transferases that catalyze their formation are unidentified. eoxPL were measured in platelets and leukocytes from an atherosclerotic cardiovascular disease (ASCVD) cohort and retrieved human arterial thrombi from three anatomical sites. The impact of age, gender, and aspirin was characterized in platelets from healthy subjects administered low-dose aspirin. The role of lysophosphatidylcholine acyltransferase 3 (LPCAT3) in eoxPL biosynthesis was tested using an inhibitor and a cell-free assay. Platelets from ASCVD patients generated lower levels of COX-derived eoxPL but elevated 12-LOX-diacyl forms, than platelets from healthy controls. This associated with aspirin and was recapitulated in healthy subjects by aspirin supplementation. P2Y12 inhibition had no impact on eoxPL. LPCAT3 inhibition selectively prevented 12-LOX-derived diacyl-eoxPL generation. LPCAT3 activity was not directly altered by aspirin. P2Y12 inhibition or aspirin had little impact on eoxPL in leukocytes. Complex aspirin-dependent gender and seasonal effects on platelet eoxPL generation were seen in healthy subjects. Limb or coronary (ST-elevation myocardial infarction, STEMI) thrombi displayed a platelet eoxPL signature while carotid thrombi had a white cell profile. EoxPL are altered in ASCVD by a commonly used cardiovascular therapy, and LPCAT3 was identified as the acyltransferase generating aspirin-sensitive 12-LOX diacyl forms. These changes to the phospholipid composition of blood cells in humans at risk of thrombosis may be clinically significant where the procoagulant membrane plays a central role in driving elevated thrombotic risk.

Lipid droplets (LDs) are transient lipid storage organelles that can be readily tapped to resupply cells with energy or lipid building blocks, and therefore play a central role in cellular metabolism. Double FYVE Domain Containing Protein 1 (DFCP1/ZFYVE1) has emerged as a key regulator of LD metabolism, where the nucleotide-dependent accumulation of DFCP1 on LDs influences their size, number, and dynamics. Here we show that DFCP1 regulates lipid metabolism by directly modulating the activity of Adipose Triglyceride Lipase (ATGL/PNPLA2), the rate-limiting lipase driving the catabolism of LDs. We show through pharmacological inhibition of key enzymes associated with LD metabolism that DFCP1 specifically regulates lipolysis and, to a lesser extent, lipophagy. Consistent with this observation, DFCP1 interacts with and recruits ATGL to LDs in starved cells, irrespective of other known regulatory factors of ATGL. We further establish that this interaction prevents dynamic disassociation of ATGL from LDs and thereby impedes the rate of LD lipolysis. Collectively, our findings indicate that DFCP1 is a nutrient-sensitive regulator of LD catabolism.

Both age and diet can contribute to alterations in triglyceride metabolism and subsequent metabolic disease. In humans, plasma triglyceride levels increase with age. Diets high in saturated fats can increase triglyceride levels while diets high in omega-3 fatty acids decrease triglyceride levels. Here we asked how age and long-term diet altered triglyceride metabolism in mice. We fed male and female C57Bl/6 mice a low-fat diet, a western diet (WD), or a diet high in polyunsaturated and omega-3 fatty acids (n3D) for up to 2 years. We measured survival, body composition, plasma triglyceride levels, chylomicron clearance, and oral fat, glucose, and insulin tolerance. Triglyceride levels in mice did not increase with age, regardless of diet. Oral fat tolerance increased with age, while chylomicron clearance remained unchanged. Decreased survival was observed in WD-fed mice. Interestingly, n3D-fed mice gained more lean mass and had lower insulin levels than WD-fed or LFD-fed mice. Moreover, triglyceride uptake into the hearts of n3D-fed mice was strikingly higher than in other groups. Our data indicate that in C57Bl/6 mice, age-induced changes in triglyceride metabolism differ from those observed in humans. Mice, like humans, appeared to have decreased fat absorption with age, but in mice plasma triglyceride clearance did not decrease with age, resulting in lower plasma triglyceride levels and improved fat tolerance with age. Although a chronic diet high in omega-3 fatty acids increased insulin sensitivity and triglyceride uptake specifically into the heart, how these observations are connected is unclear.

Diastolic dysfunction in aging mice is linked to mitochondrial abnormalities, including mitochondrial morphology disorders and decreases in membrane potential. Studies also show that aberrant mitochondrial lipid metabolism impairs mitochondrial function in aging cardiomyocytes. Our lipidomic analysis revealed that phosphatidylethanolamine (PE) levels were significantly decreased in aging myocardial mitochondria. Here, we investigated whether a reduction in PE levels in myocardial mitochondria contributes to mitochondrial injury as well as HFpEF pathogenesis and whether modulation of PE levels could ameliorate aging-induced HFpEF. Echocardiography was used to assess cardiac diastolic function in adult and aging mice treated with lysophosphatidylethanolamine (LPE) or saline. Mitochondrial morphologies from tissue samples were evaluated by transmission electron microscopy (TEM), while mitochondrial membrane potential and reactive oxygen species (ROS) levels were assessed using JC-1, MitoSOX, and DCFH-DA detection assays. We performed GO enrichment analysis between adult and aging mice and discovered significant enrichment in transcriptional programs associated with mitochondria and lipid metabolism. Also, mitochondrial PE levels were significantly decreased in aging cardiomyocytes. Treatment with LPE (200 μg/kg) significantly enhanced PE content in aging mice and improved the structure of mitochondria in cardiac cells. Also, LPE treatment protects against aging-induced deterioration of mitochondrial injury, as evidenced by increased mitochondrial membrane potential and decreased mitochondrial ROS. Furthermore, treatment with LPE alleviated severe diastolic dysfunction in aging mice. Taken together, our results suggest that LPE treatment enhances PE levels in mitochondria and ameliorates aging-induced diastolic dysfunction in mice through a mechanism involving improved mitochondrial structure and function.

During the last 30 years, an increasing number of cellular functions have been reported to be regulated by the lipid ceramide. The diversity in the ceramide structure, leading to tens of ceramide species and the discrete distribution based on subcellular topology, could explain the wide variety of functions attributed to this bioactive lipid. One of these pools of ceramide resides in the plasma membrane, and several works have suggested that an increase in plasma membrane ceramide (PMCer) in response to stimulation leads to cell death and modulates cell adhesion and migration. However, there is a limitation in studying PMCer content in this location primarily due to the inability to quantify its mass. Our group recently developed a method to specifically quantitate PMCer. In this work, we interrogate what sphingolipid metabolizing enzymes are responsible for modulating the basal levels of plasma membrane ceramide. An in-silico prediction and experimental confirmation found an almost perfect correlation between the endogenous expression levels of neutral sphingomyelinase (nSMase2) and the amount of plasma membrane ceramide in unstimulated cells. Manipulating the expression levels of nSMase2, but not other candidate enzymes of ceramide metabolism, profoundly affected PMCer. Moreover, a physiologic induction of nSMase2 during cell confluence resulted in a nSMase2-dependent dramatic increase in PMCer. Together, these results identify nSMase2 as the primary enzyme to regulate plasma membrane ceramide.

Oxysterols and bile acids are interconnected bioactive lipids playing pivotal roles in diverse physiological and pathological processes. For this reason, they are increasingly studied together for their implications in various diseases. However, due to analytical challenges inherent to the nature of these analytes, very few methods have been developed for the simultaneous analysis of these lipids. We here report the development of a sensitive LC-MS/MS method for the combined quantification of 18 oxysterols, 11 unconjugated, 15 conjugated bile acids, and 1 bile acid precursor, using 8 isotope-labeled internal standards, addressing the need for a more comprehensive analysis of these interesting lipid families. During the method development, we investigated different extraction protocols, set up a purification step, and achieved chromatographic separation for these lipids, overcoming challenges such as the large number of analytes, isomers, and wide range of polarity across the analytes. Finally, the method was successfully applied to the analysis of preclinical and clinical samples, quantifying 12 oxysterols and 14 bile acids in human plasma, 10 oxysterols and 18 bile acids in mouse plasma from the vena cava, and 10 oxysterols and 20 bile acids in mouse plasma from the portal vein within a single chromatographic run.