Very Low Density Lipoprotein Triglyceride Research Articles

FITM2 deficiency results in ER lipid accumulation, ER stress, and reduced apolipoprotein B lipidation and VLDL triglyceride secretion in vitro and in mouse liver

ObjectivesTriglycerides (TGs) associate with apolipoprotein B100 (apoB100) to form very low density lipoproteins (VLDLs) in the liver. The repertoire of factors that facilitate this association is incompletely understood. FITM2, an integral endoplasmic reticulum (ER) protein, was originally discovered as a factor participating in cytosolic lipid droplet (LD) biogenesis in tissues that do not form VLDL. We hypothesized that in the liver, in addition to promoting cytosolic LD formation, FITM2 would also transfer TG from its site of synthesis in the ER membrane to nascent VLDL particles within the ER lumen. MethodsExperiments were conducted using a rat hepatic cell line (McArdle-RH7777, or McA cells), an established model of mammalian lipoprotein metabolism, and mice. FITM2 expression was reduced using siRNA in cells and by liver specific cre-recombinase mediated deletion of the Fitm2 gene in mice. Effects of FITM2 deficiency on VLDL assembly and secretion in vitro and in vivo were measured by multiple methods, including density gradient ultracentrifugation, chromatography, mass spectrometry, stimulated Raman scattering (SRS) microscopy, sub-cellular fractionation, immunoprecipitation, immunofluorescence, and electron microscopy. Main findings1) FITM2-deficient hepatic cells in vitro and in vivo secrete TG-depleted VLDL particles, but the number of particles is unchanged compared to controls; 2) FITM2 deficiency in mice on a high fat diet (HFD) results in decreased plasma TG levels. The number of apoB100-containing lipoproteins remains similar, but shift from VLDL to low density lipoprotein (LDL) density; 3) Both in vitro and in vivo, when TG synthesis is stimulated and FITM2 is deficient, TG accumulates in the ER, and despite its availability this pool is unable to fully lipidate apoB100 particles; 4) FITM2 deficiency disrupts ER morphology and results in ER stress. Principal conclusionsThe results suggest that FITM2 contributes to VLDL lipidation, especially when newly synthesized hepatic TG is in abundance. In addition to its fundamental importance in VLDL assembly, the results also suggest that under dysmetabolic conditions, FITM2 may be an important factor in the partitioning of TG between cytosolic LDs and VLDL particles.

Importance of Dyslipidemias in Very Premature Coronary Artery Disease in India: A Registry-based Study.

Premature coronary artery disease (CAD) is an important cause of death in India. To identify risk factors in patients with premature CAD, we performed a registry-based study. Consecutive patients admitted for percutaneous coronary intervention (PCI) from October 2020 to June 2021 were recruited. The patients were classified into three groups-group I (very premature CAD < 40 years), group II (premature CAD 40-59 years), and group III (nonpremature CAD > 60 years). Major risk factors were determined, and intergroup comparison was performed. A total of 627 patients were enrolled (men 541, women 86). Group I had 79 (12.4%), group II had 420 (66.9%), and group III had 128 (20.4%) patients. The prevalence of risk factors in groups I, II, and III, respectively, were-CAD family history in 45.1, 41.1, and 26.6% (p = 0.005), current smoking/tobacco use in 29.1, 21.0, and 10.2% (p = 0.002), hypertension in 31.6, 43.6, and 59.4% (p < 0.001), and diabetes in 22.8, 34.3, and 46.1% (p < 0.001). High total cholesterol (>170 mg/dL) was present in 50.0, 38.0, and 29.9% (p = 0.005), nonhigh-density lipoprotein (HDL) cholesterol (>100 mg/dL) in 76.9, 64.4, and 54.5% (p = 0.001), low-density lipoprotein (LDL) cholesterol (>70 mg/dL) in 85.9, 76.8, and 76.4% (p = 0.167), triglycerides (>150 mg/dL) in 56.4, 45.3, and 33.1% (p = 0.001), and very low density lipoprotein (VLDL) cholesterol (>30 mg/dL) in 24.4, 10.4, and 9.4% (p = 0.005). Age- and sex-adjusted odds ratios (OR) and 95% confidence intervals (CI) for smoking/tobacco use in groups I and II compared to group III, respectively, were 3.17 (1.60-6.27) and 2.59 (1.51-4.46); high total cholesterol 2.39 (1.29-4.13) and 1.42 (0.92-2.17); high non-HDL cholesterol 2.70 (1.45-5.03) and 1.48 (0.99-2.20); and high triglycerides 2.57 (1.44-4.58) and 1.64 (1.08-2.49). Important coronary risk factors in very premature and premature CAD in India are a family history of CAD, any tobacco use, and dyslipidemias (raised total, LDL, non-HDL, and VLDL cholesterol and triglycerides).

Metabolites Associated with Polygenic Risk of Breast Cancer.

While hundreds of germline genetic variants have been associated with breast cancer risk, the mechanisms underlying the impacts of most of these variants on breast cancer remain uncertain. Metabolomics may offer valuable insights into the mechanisms underlying genetic risks of breast cancer. Among 143 cancer-free female participants, we used linear regression analyses to explore associations between the genetic risk of breast cancer, as determined by a previously developed polygenic risk score (PRS) that included 266 single-nucleotide polymorphisms (SNPs), and 223 measures of metabolites obtained from blood samples using nuclear magnetic resonance (NMR). A false discovery rate of 10% was applied to account for multiple comparisons. PRS was statistically significantly associated with 45 metabolite measures. These were primarily measures of very low-density lipoproteins (VLDLs) and high-density lipoproteins (HDLs), including triglycerides, cholesterol, and phospholipids. For example, the strongest effect was observed with the percent ratio of medium VLDL triglycerides to total lipids (0.53 unit increase in mean-standardized ln-transformed percent ratio per unit increase in PRS; q = 0.1). While larger-scale studies are needed to confirm these results, this exploratory study presents biologically plausible findings that are consistent with previously reported associations between lipids and breast cancer risk. If confirmed, these lipids could be targeted for lifestyle and pharmaceutical interventions among women at increased genetic risk of breast cancer.

Very-low-density lipoprotein triglyceride and free fatty acid plasma kinetics in women with high or low brown adipose tissue volume and overweight/obesity

Although a high amount of brown adipose tissue (BAT) is associated with low plasma triglyceride concentration, the mechanism responsible for this relationship in people is not clear. Here, we evaluate the interrelationships among BAT, very-low-density lipoprotein triglyceride (VLDL-TG), and free fatty acid (FFA) plasma kinetics during thermoneutrality in women with overweight/obesity who had a low (<20mL) or high (≥20mL) volume of cold-activated BAT (assessed by using positron emission tomography in conjunction with 2-deoxy-2-[18F]-fluoro-glucose). We find that plasma TG and FFA concentrations are lower and VLDL-TG and FFA plasma clearance rates are faster in women with high BAT than low BAT volume, whereas VLDL-TG and FFA appearance rates in plasma are not different between the two groups. These findings demonstrate that women with high BAT volume have lower plasma TG and FFA concentrations than women with low BAT volumes because of increased VLDL-TG and FFA clearance rates. This study was registered at ClinicalTrials.gov (NCT02786251).

Excess Triglycerides in Very Low-Density Lipoprotein (VLDL) Estimated from VLDL-Cholesterol could be a Useful Biomarker of Metabolic Dysfunction Associated Steatotic Liver Disease in Patients with Type 2 Diabetes.

We report that small dense low-density lipoprotein cholesterol (sdLDL-C) levels are sensitive biomarkers of metabolic dysfunction-associated steatotic liver disease (MASLD). Since triglyceride (TG)-rich very low-density lipoprotein (VLDL) is a precursor of sdLDL and is overproduced by MASLD, the composition of VLDL may be more directly associated with MAFLD than sdLDL-C or plasma TG. To identify TG-rich VLDL, this author proposed "Excess TG" and examined its association with MASLD. Patients with type 2 diabetes (n=1295), excluding fasting hypertriglyceridemia (TG ≥ 400 mg/dL) and heavy drinkers were examined. Liver steatosis and visceral fat area (VFA) were evaluated using CT. VLDL-C was calculated as the total C minus direct LDL-C minus HDL-C. The average VLDL-TG level can be estimated using VLDL-C×5, according to the principle of the Friedewald equation for LDL-C. Thus, VLDL-TG was estimated as VLDL-C×5, and Excess TG was calculated as plasma TG minus VLDL-C×5. Patients with MASLD were younger, more likely to be men and drinkers, and had higher VFA, TG, sdLDL-C, and excess TG, while VLDL-C was comparable. Excess TG was found to be the most sensitive lipid parameter for identifying MASLD, independent of sdLDL-C, TG, TG/VLDL-C, and VFA. The odds ratios for MASLD were 2.4-, 3.7-, and 3.9-fold higher for Excess TG ranges of 0-24, 25-49, and ≥ 50 mg/dL, respectively, relative to ＜0 mg, and a close relationship remained significant after adjustment for lipid- and adiposity-related parameters. Excess TG in VLDL was strongly associated with MASLD beyond TG and sdLDL-C levels, which may reflect the presence of TG-rich VLDL.

Maternal metabolic profiling across body mass index groups: An exploratory longitudinal study.

Increased BMI has been identified as a risk factor for most pregnancy complications, but the underlying metabolic factors mediating the detrimental effects of BMI are largely unknown. We aimed to compare metabolic profiles in overweight/obese women (body mass index [BMI] ≥ 25 kg/m2 ) and normal weight/underweight women (BMI < 25 kg/m2 ) across gestation. We also explored how gestational weight gain (GWG) affected maternal metabolic profiles. Exploratory nested case-control study based on a prospective longitudinal cohort of women who were healthy prior to pregnancy and gave birth at Oslo University Hospital from 2002 to 2008. The sample consisted of 48 women who were overweight/obese and 59 normal-weight/underweight women. Plasma samples from four time points in pregnancy (weeks 14-16, 22-24, 30-32 and 36-38) were analyzed by nuclear magnetic resonance spectroscopy and 91 metabolites were measured. Linear regression models were fitted for each of the metabolites at each time point. Overweight or obese women had higher levels of lipids in very-low-density lipoprotein (VLDL), total triglycerides, triglycerides in VLDL, total fatty acids, monounsaturated fatty acids, saturated fatty acids, leucine, valine, and total branched-chain amino acids in pregnancy weeks 14-16 compared to underweight and normal-weight women. Docosahexaenoic acid and degree of unsaturation were significantly lower in overweight/obese women in pregnancy weeks 36-38. In addition, overweight or obese women had higher particle concentration of XXL-VLDL and glycoprotein acetyls (GlycA) at weeks 14-16 and 30-32. GWG did not seem to affect the metabolic profile, regardless of BMI group when BMI was treated as a dichotomous variable, ≥25 kg/m2 (yes/no). Overweight or obese women had smaller pregnancy-related metabolic alterations than normal-weight/underweight women. There was a trend toward higher triglyceride and VLDL particle concentration in overweight/obese women. As this was a hypothesis-generating study, the similarities with late-onset pre-eclampsia warrant further investigation. The unfavorable development of fatty acid composition in overweight/obese women, with possible implication for the offspring, should also be studied further in the future.

Severe hypertriglyceridemia and its correlation with clinical atherosclerosis and pancreatitis investigated in beta-quantification

Abstract Background Severe hypertriglyceridemia is commonly associated with pancreatitis and questionably atherosclerotic cardiovascular disease (ASCVD). However, while some patients develop those sequelae at relatively low levels, others with extreme levels remain spared from it. Purpose This study serves to validate, whether the distribution of triglycerides (TG), either in very-low-density lipoprotein (VLDL) particles or chylomicrons (CM) can be decisive for the development of clinical disease. Methods Patients with TG levels above 500 mg/dL, who routinely visited our lipid outpatient clinic between 2016 and 2021 with their lipid panel done via beta-quantification were retrospectively investigated for the presence of clinical pancreatitis and ASCVD via electronic chart-review. Results In total, 221 patients met eligibility criteria, with a mean age of 48 years, BMI of 29.7, with 72.4% males. Median TG and low-density lipoprotein cholesterol levels were 827 mg/dL and 62 mg/dL, respectively. A history of pancreatitis was present in 13.7% of patients, while ASCVD was present in 20.7%. TG distribution, described as the ratio of TG in VLDL per TG in CM showed no significant differences with regards to pancreatitis incidence (p=0.393). For ASCVD, the ratio was significantly higher in those with ASCVD vs those without (3.54 vs 2.25) (p=0.008). CM-dominant TG distribution was present in 19.5% of patients and was associated with higher total TG levels (2122 mg/dL vs 825 mg/dL), compared to VLDL-dominant TG distribution. Conclusion A higher relative amount of TGs in VLDL compared to CM, as most prevalent pattern, was associated with ASCVD, while very high levels of TG frequently relate to hyperchylomicronemia.

The early effects of sleeve gastrectomy on postprandial chylomicron triglycerides during the progression of type 2 diabetes

BackgroundIt remains unclear whether early sleeve gastrectomy (SG) improves postprandial very-low-density lipoprotein (VLDL) as well as chylomicron triglycerides (TGs) in a weight-independent manner in patients with or without type 2 diabetes (DM). Herein we investigated the early effects of SG on postprandial VLDL and chylomicron kinetics. MethodsA liquid meal test was performed before and after 1 week of SG. The plasma was collected for postprandial triglyceride-rich lipoprotein kinetics analyses, including VLDLs and chylomicrons, isolated by high-speed ultracentrifugation. Lipidomics and metabolomics were used to profile lipid and metabolite compositions of plasma and postprandial chylomicrons. De novo fatty acid synthesis in intestinal epithelial cells treated with chylomicron metabolites was examined using RT-PCR, immunoblotting, and free fatty acid measurement. ResultsWe found that patients with DM had markedly higher VLDL TGs than patients without DM, and such an increase was still retained after SG. In contrast, SG significantly decreased postprandial chylomicron TGs, but surprisingly, the degree of the reduction in patients with DM was less prominent than in patients without DM, confirmed by untargeted lipidomics analysis. Moreover, 5 unique metabolites potentially linked to de novo fatty acid synthesis from the pathway analysis were discovered by further metabolomic analysis of postprandial chylomicrons from patients with DM who underwent SG and verified by In vitro intestinal epithelial cell culture experiments. ConclusionsSG in 1 week did not impact postprandial VLDL but decreased chylomicron TGs. Patients with DM keep higher postprandial chylomicron TG concentrations than patients without it after SG, potentially through some unique metabolites that increase intestinal fatty acid synthesis. These results implicate the timing for SG to reach lower intestinal fatty acid synthesis and postprandial chylomicron TG production is prior to the diagnosis of DM to potentially reduce cardiovascular risks.

Exercise increases myocardial free fatty acid oxidation in subjects with metabolic dysfunction-associated fatty liver disease

Background and aimsMetabolic dysfunction-associated fatty liver disease (MAFLD) is associated with dyslipidemia and may promote cardiac lipotoxicity. Myocardial free fatty acids (FFA) oxidation (MOFFA) is normal in pre-diabetes, but reduced in heart failure. We hypothesized that during exercise MOFFA, very low-density lipoprotein triglycerides (VLDL-TG) secretion, hepatic FFA utilization, and lactate production differ among obese subjects with and without MAFLD. MethodsNine obese subjects with MAFLD and 8 matched subjects without MAFLD (Control) without a history of heart failure and cardiovascular disease were compared before and after 90-min exercise at 50% Peak oxygen consumption. Basal and exercise induced cardiac and hepatic FFA oxidation, uptake and re-esterification and VLDL-TG secretion were measured using [11C]palmitate positron-emission tomography and [1–14C]VLDL-TG. ResultsIn the heart, increased MOFFA was observed after exercise in MAFLD, whereas MOFFA decreased in Control (basal vs exercise, MAFLD: 4.1 (0.8) vs 4.8 (0.8) μmol·100 ml−1 min−1; Control: 4.9 (1.8) vs 4.0 (1.1); μmol·100 ml−1 min−1, mean (SD), p < 0.048). Hepatic FFA fluxes were significantly lower in MAFLD than Control and increased ≈ two-fold in both groups. VLDL-TG secretion was 50% greater in MAFLD at rest and similarly suppressed during exercise. Plasma lactate increased significantly less in MAFLD than Control during exercise. ConclusionsUsing robust tracer-techniques we found that obese subjects with MAFLD do not downregulate MOFFA during exercise compared to Control, possibly due to diminished lactate supply. Hepatic FFA fluxes are significantly lower in MAFLD than Control, but increase similarly with exercise. VLDL-TG export remains greater in MAFLD compared to Control. Basal and post-exercise myocardial and hepatic FFA, VLDL-TG and lactate metabolism is abnormal in subjects with MAFLD compared to Control.

Hyperlipidaemia elicits an atypical, T helper 1-like CD4+ T-cell response: a key role for very low-density lipoprotein.

Hyperlipidemia and T cell driven inflammation are important drivers of atherosclerosis, the main underlying cause of cardiovascular disease. Here, we detailed the effects of hyperlipidemia on T cells. In vitro, exposure of human and murine CD4+ T cells to very low-density lipoprotein (VLDL), but not to low-density lipoprotein (LDL) resulted in upregulation of Th1 associated pathways. VLDL was taken up via a CD36-dependent pathway and resulted in membrane stiffening and a reduction in lipid rafts. To further detail this response in vivo, T cells of mice lacking the LDL receptor (LDLr), which develop a strong increase in VLDL cholesterol and triglyceride levels upon high cholesterol feeding were investigated. CD4+ T cells of hyperlipidemic Ldlr-/- mice exhibited an increased expression of the C-X-C-chemokine receptor 3 (CXCR3) and produced more interferon-γ (IFN-γ). Gene set enrichment analysis identified IFN-γ-mediated signaling as the most upregulated pathway in hyperlipidemic T cells. However, the classical Th1 associated transcription factor profile with strong upregulation of Tbet and Il12rb2 was not observed. Hyperlipidemia did not affect levels of the CD4+ T cell's metabolites involved in glycolysis or other canonical metabolic pathways but enhanced amino acids levels. However, CD4+ T cells of hyperlipidemic mice showed increased cholesterol accumulation and an increased arachidonic acid (AA) to docosahexaenoic acid (DHA) ratio, which was associated with inflammatory T cell activation. Hyperlipidemia, and especially its VLDL component induces an atypical Th1 response in CD4+ T cells. Underlying mechanisms include CD36 mediated uptake of VLDL, and an altered AA/DHA ratio.

Mediterranean Diet, Energy Restriction, Physical Activity, and Atherogenicity of Very-Low Density Lipoproteins: Findings from Two Randomized Controlled Trials.

Some very-low density lipoprotein (VLDL) properties may render them more pro-atherogenic. We aimed to assess whether a Mediterranean diet (MedDiet) or an energy-reduced MedDiet with increased physical activity improves them. In a sample of the PREvención con DIeta MEDiterránea (PREDIMED) study, a 1-year intervention with MedDiet with extra-virgin olive oil (n = 89) or nuts (MedDiet-Nuts; n = 79) is compared with a low-fat diet (n=90). In the PREDIMED-Plus study, a 1-year intervention with energy-reduced MedDiet and physical activity (n = 103) is compared with an ad libitum MedDiet (n = 101). VLDL levels of apolipoprotein C-I, C-III, triglycerides, and cholesterol; the apolipoprotein E-/C-I ratio; and VLDL ex-vivo triglyceride transfer are measured. In PREDIMED participants in both MedDiet groups combined, VLDL apolipoprotein C-III levels are nominally reduced (-0.023 SD units, 95% CI -0.44 to -0.014, p = 0.037). VLDL triglyceride transfer is nominally increased in the MedDiet-Nuts group (+0.39 SD units, 95% CI 0.012-0.78, p = 0.045). In PREDIMED-Plus, no inter-group differences are detected. In older adults at high cardiovascular risk, MedDiet is associated with lower VLDL atherogenicity versus a low-fat diet. No differences are seen after an energy-reduced MedDiet with physical activity.

Cholesterol Content of Very-Low-Density Lipoproteins Is Associated with 1-Year Mortality in Acute Heart Failure Patients.

Considering the relationship between the extent of metabolic derangement and the disease severity in heart failure, we hypothesized that the lipid content of very-low-density lipoprotein (VLDL) may have prognostic value for 1 year mortality in acute heart failure (AHF). Baseline serum levels of VLDL cholesterol (VLDL-C), VLDL triglycerides (VLDL-TG), VLDL phospholipids (VLDL-PL), and VLDL apolipoprotein B (VLDL-apoB) were measured using NMR spectroscopy. We calculated the ratios of the respective VLDL lipids and VLDL apoB (VLDL-C/VLDL-apoB, VLDL-TG/VLDL-apoB, and VLDL-PL/VLDL-apoB), as estimators of the cholesterol, triglyceride, and phospholipid content of VLDL particles and tested their association with mortality. Out of 315 AHF patients, 118 (37.5%) patients died within 1 year after hospitalization for AHF. Univariable Cox regression analyses revealed a significant inverse association of VLDL-C/VLDL-apoB (hazard ratio (HR) 0.43, 95% confidence interval (CI) 0.29–0.64, p < 0.001), VLDL-TG/VLDL-apoB (HR 0.79, 95% CI 0.71–0.88, p < 0.001), and VLDL-PL/VLDL-apoB (HR 0.37, 95% CI 0.25–0.56, p < 0.001) with 1 year mortality. Of the tested parameters, only VLDL-C/VLDL-apoB remained significant after adjustment for age and sex, as well as other clinical and laboratory parameters that showed a significant association with 1 year mortality in the univariable analyses. We conclude that cholesterol content of circulating VLDL (VLDL-C/VLDL-apoB) might be of prognostic value in AHF.

Leptin increases hepatic triglyceride export via a vagal mechanism in humans

Recombinant human leptin (metreleptin) reduces hepatic lipid content in patients with lipodystrophy and overweight patients with non-alcoholic fatty liver disease and relative hypoleptinemia independent of its anorexic action. In rodents, leptin signaling in the brain increases very-low-density lipoprotein triglyceride (VLDL-TG) secretion and reduces hepatic lipid content via the vagus nerve. In this randomized, placebo-controlled crossover trial (EudraCT Nr. 2017-003014-22), we tested whether a comparable mechanism regulates hepatic lipid metabolism in humans. A single metreleptin injection stimulated hepatic VLDL-TG secretion (primary outcome) and reduced hepatic lipid content in fasted, lean men (n= 13, age range 20-38 years) but failed to do so in metabolically healthy liver transplant recipients (n= 9, age range 26-62 years) who represent a model for hepatic denervation. In an independent cohort of lean men (n= 10, age range 23-31 years), vagal stimulation by modified sham feeding replicated the effects of metreleptin on VLDL-TG secretion. Therefore, we propose that leptin has anti-steatotic properties that are independent of food intake by stimulating hepatic VLDL-TG export via a brain-vagus-liver axis.

Macrophages take up VLDL-sized emulsion particles through caveolae-mediated endocytosis and excrete part of the internalized triglycerides as fatty acids.

Triglycerides are carried in the bloodstream as part of very low-density lipoproteins (VLDLs) and chylomicrons, which represent the triglyceride-rich lipoproteins. Triglyceride-rich lipoproteins and their remnants contribute to atherosclerosis, possibly by carrying remnant cholesterol and/or by exerting a proinflammatory effect on macrophages. Nevertheless, little is known about how macrophages process triglyceride-rich lipoproteins. Here, using VLDL-sized triglyceride-rich emulsion particles, we aimed to study the mechanism by which VLDL triglycerides are taken up, processed, and stored in macrophages. Our results show that macrophage uptake of VLDL-sized emulsion particles is dependent on lipoprotein lipase (LPL) and requires the lipoprotein-binding C-terminal domain but not the catalytic N-terminal domain of LPL. Subsequent internalization of VLDL-sized emulsion particles by macrophages is carried out by caveolae-mediated endocytosis, followed by triglyceride hydrolysis catalyzed by lysosomal acid lipase. It is shown that STARD3 is required for the transfer of lysosomal fatty acids to the ER for subsequent storage as triglycerides, while NPC1 likely is involved in promoting the extracellular efflux of fatty acids from lysosomes. Our data provide novel insights into how macrophages process VLDL triglycerides and suggest that macrophages have the remarkable capacity to excrete part of the internalized triglycerides as fatty acids.

Triglyceride Metabolism Modifies Lipoprotein(a) Plasma Concentration.

Lipoprotein(a) (Lp(a)) is a significant cardiovascular risk factor. Knowing the mechanisms that regulate its concentration can facilitate the development of Lp(a)-lowering drugs. This study analyzes the relationship between triglycerides (TGs) and Lp(a) concentrations, cross-sectionally and longitudinally, and the influence of the number and composition of TG-rich lipoproteins, and the APOE genotype. Data from Aragon Workers Health Study (AWHS) (n = 5467), National Health and Nutrition Examination Survey III phase 2 (n = 3860), and Hospital Universitario Miguel Servet (HUMS) (n = 2079) were used for cross-sectional TG and Lp(a) relationship. Lp(a) intrasubject variation was studied in AWHS participants and HUMS patients with repeated measurements. TG-rich lipoproteins were quantified by nuclear magnetic resonance in a subsample from AWHS. Apolipoproteins B and E were quantified by Luminex in very low-density lipoprotein (VLDL) isolated by ultracentrifugation, from HUMS samples. APOE genotyping was carried in AWHS and HUMS participants. Regression models adjusted for age and sex were used to study the association. The 3 studies showed an inverse relationship between TG and Lp(a). Increased VLDL number, size, and TG content were associated with significantly lower Lp(a). There was an inverse association between the apoE concentration in VLDL and Lp(a). No significant association was observed for apolipoprotein (apo)B. Subjects carrying the apoE2/E2 genotype had significantly lower levels of Lp(a). Our results show an inverse relationship Lp(a)-TG. Subjects with larger VLDL size have lower Lp(a), and lower values of Lp(a) were present in patients with apoE-rich VLDL and apoE2/E2 subjects. Our results suggest that bigger VLDLs and VLDLs enriched in apoE are inversely involved in Lp(a) plasma concentration.

Lipoprotein profiles associated with exposure to poly- and perfluoroalkyl substances (PFASs) in the EuroMix human biomonitoring study

Exposure to per- and polyfluoroalkyl substances (PFASs) is associated with increased blood cholesterol. Although elevated cholesterol is a well-established risk factor for cardiovascular diseases (CVD), it is not clear whether PFASs affect this risk. Lipoprotein subclasses are emerging biomarkers for disease risk and lipoprotein profiling may provide an insight to physiological implications of PFAS exposure. We explored the association between serum PFAS concentrations and lipoprotein subclasses in a cross-sectional study. We determined the concentrations and lipid composition of the major subclasses of lipoproteins in plasma samples from 127 adult participants of the EuroMix human biomonitoring study by nuclear magnetic resonance (NMR). Serum concentrations of 17 PFASs showed a detection frequency between 30 and 100% and were included in further analyses. We examined the associations between PFAS concentrations and lipoprotein subclasses by linear mixed-effect regression models, adjusted for confounders. In the adjusted models, positive associations were found between several PFASs and cholesterol concentrations in large to medium sized HDL and medium sized LDL particles. We found a 4–12% increase in HDL cholesterol per interquartile range (IQR) increase for several PFASs. In women the associations with PFNA, PFUnDA, PFDoDA and PFOS were significant after adjustment for multiple comparisons. Similar magnitude of change was observed between longer chained PFASs and LDL cholesterol, and a few of these associations reached significance for cholesterol in large to medium LDL particle sizes in women. No significant associations with plasma triglycerides were observed. However, most PFASs tended to be associated with reduction in VLDL (very low-density lipoproteins) particle number and VLDL triglyceride. Findings from this exploratory study, suggest that background PFAS exposures influence particle size distributions and lipid composition of plasma lipoprotein subclasses, and that these effects may be more prominent in women. A two-points lipoprofiling for all subjects indicated both low intra-individual variability and good analytical reproducibility.

276-OR: Impaired Glucagon-Mediated Suppression of VLDL-TG Secretion in Individuals with MAFLD

Background: Individuals with Metabolic Dysfunction-Associated Fatty Liver Disease (MAFLD) have both elevated triglyceride and glucagon levels, although glucagon suppresses the very-low-density lipoprotein-triglyceride (VLDL-TG) secretion in lean individuals. Therefore, we hypothesized that the glucagon-mediated suppression of VLDL-TG secretion is impaired in MAFLD, with greater impairment in individuals with steatohepatitis (MASH) than individuals with simple steatosis (MAFL) . Methods: 28 participants were included in the study (seven overweight control subjects (CON) , patients with MAFL, and with MASH) . We performed a somatostatin-clamp with replacement doses of growth hormone, insulin (0.2 mU/kg/min) and glucagon to suppress endogenous hormone production. Participants were studied during low-dose glucagon infusion (0.65 ng/kg/min, t=0─270 min, LowG) followed by a high-dose glucagon infusion (1.5 ng/kg/min, t=270─450 min, HighG) . We used 14C-labeled VLDL-TG and 3H-labeled glucose tracers to evaluate VLDL-TG secretion and endogenous glucose production (EGP) . Results: HighG suppressed VLDL-TG secretion compared to LowG in all groups, but with a greater reduction in VLDL-TG secretion in CON than in the MAFLD groups (CON 36% (±SD 19%) ; MAFL 10% (±SD 8%) ; MASH 13% (±SD 16%) , p&amp;lt;0.01) . Thus, the suppression was similarly mitigated in both MAFLD groups. This led to insignificant reductions in plasma VLDL-TG, TG and VLDL-TG oxidation rates in the CON group, in contrast to stable or increasing values in the MAFLD groups (CON vs. MAFLD delta-values, p&amp;lt;0.01, p=0.02 and p=0.04) , respectively) . EGP was similar in all groups throughout the LowG period and rose similarly when increasing glucagon infusion rate. Conclusions: Patients with MAFLD, MAFL as well as MASH, have an impaired glucagon-mediated suppression of VLDL-TG secretion which is a probable contributing factor to the dyslipidemia and the increased cardiovascular risk observed in individuals with MAFLD. NCT04042142 Disclosure S.Heebøll: None. J.R.Risikesan: None. H.Gronbaek: Other Relationship; ARLA, Research Support; AbbVie Inc., Intercept Pharmaceuticals, Inc., Speaker's Bureau; AstraZeneca, Novartis AG. E.Søndergaard: None. S.Nielsen: None. Funding Independent Research Fund Denmark (9039-00177B and -00278B) and Aase og Ejnar Danielsens Fond

Hepatic MDM2 Causes Metabolic Associated Fatty Liver Disease by Blocking Triglyceride-VLDL Secretion via ApoB Degradation.

Dysfunctional triglyceride‐very low‐density lipoprotein (TG‐VLDL) metabolism is linked to metabolic‐associated fatty liver disease (MAFLD); however, the underlying cause remains unclear. The study shows that hepatic E3 ubiquitin ligase murine double minute 2 (MDM2) controls MAFLD by blocking TG‐VLDL secretion. A remarkable upregulation of MDM2 is observed in the livers of human and mouse models with different levels of severity of MAFLD. Hepatocyte‐specific deletion of MDM2 protects against high‐fat high‐cholesterol diet‐induced hepatic steatosis and inflammation, accompanied by a significant elevation in TG‐VLDL secretion. As an E3 ubiquitin ligase, MDM2 targets apolipoprotein B (ApoB) for proteasomal degradation through direct protein–protein interaction, which leads to reduced TG‐VLDL secretion in hepatocytes. Pharmacological blockage of the MDM2‐ApoB interaction alleviates dietary‐induced hepatic steatohepatitis and fibrosis by inducing hepatic ApoB expression and subsequent TG‐VLDL secretion. The effect of MDM2 on VLDL metabolism is p53‐independent. Collectively, these findings suggest that MDM2 acts as a negative regulator of hepatic ApoB levels and TG‐VLDL secretion in MAFLD. Inhibition of the MDM2‐ApoB interaction may represent a potential therapeutic approach for MAFLD treatment.

Abstract P020: Triglyceride-lowering Lipoprotein Lipase Gene Variants And Improvements In Lipids And Lipoprotein Subspecies Defined By Apolipoprotein C-iii: Pounds Lost Trial

Introduction: Triglyceride-lowering variants in the lipoprotein lipase ( LPL ) gene have been associated with a lower risk of coronary heart disease. Lipoproteins are heterogeneous, and lipoprotein subspecies containing apolipoprotein C-III (ApoCIII) have adverse effects on cardiovascular disease (CVD) risk. Hypothesis: We examined associations of triglyceride-lowering LPL gene variants with long-term improvement in lipids and lipoprotein subspecies with ApoCIII in patients with obesity. We also investigated whether the LPL gene variants were significantly related to the improvement of the 10-year CVD risk estimated using the Framingham risk score in the participants of a weight-loss dietary intervention trial. Methods: This study included 382 overweight and obese adults of white European ancestry in a 2-year weight-loss dietary intervention, the POUNDS Lost trial. We evaluated changes in lipids (triglycerides and cholesterol) and lipoproteins (such as very-low-density lipoprotein (VLDL), low-density lipoprotein (LDL), and high-density lipoprotein (HDL)) subfractions defined by the presence or absence of ApoCIII from baseline to 2 years after the intervention. A genetic risk score of LPL (LPL-GRS) was calculated by summing triglyceride-lowering alleles of five independent single nucleotide polymorphisms (SNPs). We calculated the Framingham risk score and estimated changes in the 10-year CVD risk after the intervention. Results: At baseline, higher scores of triglyceride-lowering LPL-GRS were significantly associated with higher levels of HDL cholesterol (p= 0.0065) and lower levels of triglycerides (p= 0.017). Higher LPL-GRS was also associated with more decreases in total triglycerides (p= 0.028) and triglycerides in VLDL with ApoCIII (p=0.018) at 2 years. The LPL-GRS was also predictive of 2-year improvements in other atherogenic lipoprotein subtypes, such as cholesterol in VLDL with ApoCIII (p= 0.037) and cholesterol in LDL with ApoCIII (p= 0.027) after the intervention. Further, the LPL-GRS was significantly associated with a 2-year reduction of the estimated CVD risk, regardless of the initial risk status (p=0.034). Conclusion: The triglyceride-lowering LPL-GRS was significantly predictive of improvements in unfavorable lipid profiles, including lipoprotein subtypes containing ApoCIII after consuming a weight-loss diet in patients with obesity. The reduction of the estimated CVD risk after the dietary intervention was predicted by the LPL-GRS at the pre-intervention.

Liver-Specific Deletion of Mouse Tm6sf2 Promotes Steatosis, Fibrosis, and Hepatocellular Cancer.

Human transmembrane 6 superfamily 2 (TM6SF2) variant rs58542926 is associated with NAFLD and HCC. However, conflicting reports in germline Tm6sf2 knockout mice suggest no change or decreased very low density lipoprotein (VLDL) secretion and either unchanged or increased hepatic steatosis, with no increased fibrosis. We generated liver-specific Tm6Sf2 knockout mice (Tm6 LKO) to study VLDL secretion and the impact on development and progression of NAFLD. Two independent lines of Tm6 LKO mice exhibited spontaneous hepatic steatosis. Targeted lipidomic analyses showed increased triglyceride species whose distribution and abundance phenocopied findings in mice with liver-specific deletion of microsomal triglyceride transfer protein. The VLDL triglyceride secretion was reduced with small, underlipidated particles and unchanged or increased apolipoprotein B. Liver-specific adeno-associated viral, serotype 8 (AAV8) rescue using either wild-type or mutant E167K-Tm6 reduced hepatic steatosis and improved VLDL secretion. The Tm6 LKO mice fed a high milk-fat diet for 3weeks exhibited increased steatosis and fibrosis, and those phenotypes were further exacerbated when mice were fed fibrogenic, high fat/fructose diets for 20weeks. In two models of HCC, either neonatal mice injected with streptozotocin (NASH/STAM) and high-fat fed or with diethylnitrosamine injection plus fibrogenic diet feeding, Tm6 LKO mice exhibited increased steatosis, greater tumor burden, and increased tumor area versus Tm6 flox controls. Additionally, diethylnitrosamine-injected and fibrogenic diet-fed Tm6 LKO mice administered wild-type Tm6 or E167K-mutant Tm6 AAV8 revealed significant tumor attenuation, with tumor burden inversely correlated with Tm6 protein levels. Liver-specific Tm6sf2 deletion impairs VLDL secretion, promoting hepatic steatosis, fibrosis, and accelerated development of HCC, which was mitigated with AAV8- mediated rescue.