Proteomic Analysis of Egg Yolk Proteins During Embryonic Development in Wanxi White Goose.

To investigate the alterations of yolk protein during embryonic development in Wanxi white goose, the egg yolk protein composition at days 0, 4, 7, 14, 18, and 25 of incubation (D0, D4, D7, D14, D18, and D25) was analyzed by two-dimensional gel electrophoresis combined with mass spectrometry. A total of 65 spots representing 11 proteins with significant abundance changes were detected. Apolipoprotein B-100, vitellogenin-1, vitellogenin-2-like, riboflavin-binding protein, and serotransferrin mainly participated in nutrient (lipid, riboflavin, and iron ion) transport, and vitellogenin-2-like showed a lower abundance after D14. Ovomucoid-like were involved in endopeptidase inhibitory activity and immunoglobulin binding and exhibited a higher expression after D18, suggesting a potential role in promoting the absorption of immunoglobulin and providing passive immune protection for goose embryos after D18. Furthermore, myosin-9 and actin (ACTB) were involved in the tight junction pathway, potentially contributing to barrier integrity. Serum albumin mainly participated in cytolysis and toxic substance binding. Therefore, the high expression of serum albumin, myosin-9, and ACTB throughout the incubation might protect the developing embryo. Apolipoprotein B-100, vitellogenin-1, vitellogenin-2-like, riboflavin-binding protein, and serotransferrin might play a crucial role in providing nutrition for embryonic development, and VTG-2-like was preferentially degraded/absorbed.

ANGPTL3 deficiency impairs lipoprotein production and produces adaptive changes in hepatic lipid metabolism.

Angiopoietin-like protein 3 (ANGPTL3) is a hepatically secreted protein and therapeutic target for reducing plasma triglyceride-rich lipoproteins and low-density lipoprotein (LDL) cholesterol. Although ANGPTL3 modulates the metabolism of circulating lipoproteins, its role in triglyceride-rich lipoprotein assembly and secretion remains unknown. CRISPR-associated protein 9 (CRISPR/Cas9) was used to target ANGPTL3 in HepG2 cells (ANGPTL3-/-) whereupon we observed ∼50% reduction of apolipoprotein B100 (ApoB100) secretion, accompanied by an increase in ApoB100 early presecretory degradation via a predominantly lysosomal mechanism. Despite defective particle secretion in ANGPTL3-/- cells, targeted lipidomic analysis did not reveal neutral lipid accumulation in ANGPTL3-/- cells; rather ANGPTL3-/- cells demonstrated decreased secretion of newly synthesized triglycerides and increased fatty acid oxidation. Furthermore, RNA sequencing demonstrated significantly altered expression of key lipid metabolism genes, including targets of peroxisome proliferator-activated receptor α, consistent with decreased lipid anabolism and increased lipid catabolism. In contrast, CRISPR/Cas9 LDL receptor (LDLR) deletion in ANGPTL3-/- cells did not result in a secretion defect at baseline, but proteasomal inhibition strongly induced compensatory late presecretory degradation of ApoB100 and impaired its secretion. Additionally, these ANGPTL3-/-;LDLR-/- cells rescued the deficient LDL clearance of LDLR-/- cells. In summary, ANGPTL3 deficiency in the presence of functional LDLR leads to the production of fewer lipoprotein particles due to early presecretory defects in particle assembly that are associated with adaptive changes in intrahepatic lipid metabolism. In contrast, when LDLR is absent, ANGPTL3 deficiency is associated with late presecretory regulation of ApoB100 degradation without impaired secretion. Our findings therefore suggest an unanticipated intrahepatic role for ANGPTL3, whose function varies with LDLR status.

PCSK7: A novel regulator of apolipoprotein B and a potential target against non-alcoholic fatty liver disease.

BACKGROUND: Epidemiological evidence links the proprotein convertase subtilisin/kexin 7 (PCSK7) to triglyceride (TG) metabolism. We associated the known PCSK7 gain-of-function non-coding SNP rs236918 with higher levels of plasma apolipoprotein B (apoB) and the loss-of-function coding variant p.Pro777Leu (SNP rs201598301) with lower apoB and TG. Herein, we aimed to unravel the in vivo role of liver PCSK7. METHODS: We biochemically defined the functional role of PCSK7 in lipid metabolism using hepatic cell lines and Pcsk7-/- mice. Our findings were validated following subcutaneous administration of hepatocyte-targeted N-acetylgalactosamine (GalNAc)-antisense oligonucleotides (ASOs) against Pcsk7. RESULTS: Independent of its proteolytic activity, membrane-bound PCSK7 binds apoB100 in the endoplasmic reticulum and enhances its secretion. Mechanistically, the loss of PCSK7/Pcsk7 leads to apoB100 degradation, triggering an unfolded protein response, autophagy, and ß-oxidation, eventually reducing lipid accumulation in hepatocytes. Non-alcoholic fatty liver disease (NAFLD) was induced by a 12-week high fat/fructose/cholesterol diet in wild type (WT) and Pcsk7-/- mice that were then allowed to recover on a 4-week control diet. Pcsk7-/- mice recovered more effectively than WT mice from all NAFLD-related liver phenotypes. Finally, subcutaneous administration of GalNAc-ASOs targeting hepatic Pcsk7 to WT mice validated the above results. CONCLUSIONS: Our data reveal hepatic PCSK7 as one of the major regulators of apoB, and its absence reduces apoB secretion from hepatocytes favoring its ubiquitination and degradation by the proteasome. This results in a cascade of events, eventually reducing hepatic lipid accumulation, thus supporting the notion of silencing PCSK7 mRNA in hepatocytes for targeting NAFLD.

Improved isolation of extracellular vesicles by removal of both free proteins and lipoproteins.

Extracellular vesicles (EVs) are released by all cells into biofluids such as plasma. The separation of EVs from highly abundant free proteins and similarly sized lipoproteins remains technically challenging. We developed a digital ELISA assay based on Single Molecule Array (Simoa) technology for ApoB-100, the protein component of several lipoproteins. Combining this ApoB-100 assay with previously developed Simoa assays for albumin and three tetraspanin proteins found on EVs (Ter-Ovanesyan, Norman et al., 2021), we were able to measure the separation of EVs from both lipoproteins and free proteins. We used these five assays to compare EV separation from lipoproteins using size exclusion chromatography with resins containing different pore sizes. We also developed improved methods for EV isolation based on combining several types of chromatography resins in the same column. We present a simple approach to quantitatively measure the main impurities of EV isolation in plasma and apply this approach to develop novel methods for enriching EVs from human plasma. These methods will enable applications where high-purity EVs are required to both understand EV biology and profile EVs for biomarker discovery.

LncRP11-675F6.3 responds to rapamycin treatment and reduces triglyceride accumulation via interacting with HK1 in hepatocytes by regulating autophagy and VLDL-related proteins.

Long noncoding RNAs (lncRNAs) have been widely proven to be involved in liver lipid homeostasis. Herein, we identify an upregulated lncRNA named lncRP11-675F6.3 in response to rapamycin treatment using a microarray in HepG2 cells. Knockdown of lncRP11-675F6. 3 leads to a significant reduction in apolipoprotein 100 (ApoB100), microsomal triglyceride transfer protein (MTTP), ApoE and ApoC3 with increased cellular triglyceride level and autophagy. Furthermore, we find that ApoB100 is obviously colocalized with GFP-LC3 in autophagosomes when lncRP11-675F6. 3 is knocked down, indicating that elevated triglyceride accumulation likely related to autophagy induces the degradation of ApoB100 and impairs very low-density lipoprotein (VLDL) assembly. We then identify and validate that hexokinase 1 (HK1) acts as the binding protein of lncRP11-675F6.3 and mediates triglyceride regulation and cell autophagy. More importantly, we find that lncRP11-675F6.3 and HK1 attenuate high fat diet induced nonalcoholic fatty liver disease (NAFLD) by regulating VLDL-related proteins and autophagy. In conclusion, this study reveals that lncRP11-675F6.3 is potentially involved in the downstream of mTOR signaling pathway and the regulatory network of hepatic triglyceride metabolism in cooperation with its interacting protein HK1, which may provide a new target for fatty liver disorder treatment.

Differentially Regulated Apolipoproteins and Lipid Profiles as Novel Biomarkers for Polypoidal Choroidal Vasculopathy and Neovascular Age-Related Macular Degeneration.

Lipid dyshomeostasis has been implicated in the pathogenesis of various retinal and choroidal vascular diseases. This study aims to investigate whether apolipoprotein (apo) mediated differential regulation of lipid metabolism contributes to the phenotypes of polypoidal choroidal vasculopathy (PCV) and neovascular age-related macular degeneration (nAMD). This study involved 148 subjects including 53 patients with PCV, 44 patients with nAMD, and 51 age-, sex-matched subjects with normal fundus controls. Routine blood biochemistry profile was evaluated. Apolipoproteins was estimated by Luminex technology. After controlling for age, gender, body mass index, duration of hypertension and type 2 diabetes mellitus, apoB/non-high density lipoprotein cholesterol (HDL-C) (p=0.015) was an independent risk factor for nAMD, apoB was an independent risk factor for PCV(p=0.011), compared with control. Low-density lipoprotein cholesterol (LDL-C) was significantly higher in patients with PCV when compared with nAMD (p=0.037). Furthermore, apoB/non-HDL, LDL-C, triglycerides and were significantly correlated with the pathogenesis of subgroups of PCV and nAMD. We concluded that lipid profiles and apos are differential regulated in PCV, nAMD and their subtypes, indicating different pathogenicity contributed to the different phenotypes of PCV and nAMD. Non-pachy PCV shares pathological similarities with nAMD, which is highly correlated with age-related atherosclerosis.

Associations between insulin-like growth factor binding protein-2 and lipoprotein kinetics in men.

Low circulating concentrations of insulin-like growth factor binding protein-2 (IGFBP-2) have been associated with dyslipidemia, notably with high triglyceride (TG) levels. However, the determinants by which IGFBP-2 influences lipoprotein metabolism, especially that of TG-rich lipoproteins (TRLs), are poorly understood. Here, we aimed to assess the relationships between IGFBP-2 levels and lipoprotein production and catabolism in human subjects. Fasting IGFBP-2 concentrations were measured in the plasma of 219 men pooled from previous lipoprotein kinetics studies. We analyzed production rate and fractional catabolic rates of TRLapoB-48, and LDL-, IDL-, and VLDLapoB-100 by multicompartmental modeling of l-[5,5,5-D3] leucine enrichment data after a 12 h primed constant infusion in individuals kept in a constant nutritional steady state. Subjects had an average BMI of 30 kg/m2, plasma IGFBP-2 levels of 157 ng/ml, and TG of 2.2 mmol/l. After adjustments for age and BMI, IGFBP-2 levels were negatively associated with plasma TG (r = -0.29; P < 0.0001) and positively associated with HDL-cholesterol (r = 0.26; P < 0.0001). In addition, IGFBP-2 levels were positively associated with the fractional catabolic rate of VLDLapoB-100 (r = 0.20; P < 0.01) and IDLapoB-100 (r = 0.19; P < 0.05) and inversely with the production rate of TRLapoB-48 (r = -0.28; P < 0.001). These correlations remained statistically significant after adjustments for age, BMI, and the amount of fat given during the tracer infusion. These findings show that the association between low plasma IGFBP-2 and high TG concentrations could be due to both an impaired clearance of apoB-100-containing VLDL and IDL particles and an increased production of apoB-48-containing chylomicrons. Additional studies are necessary to investigate whether and how IGFBP-2 directly impacts the kinetics of TRL.

Physiological interindividual variability in endogenous estradiol concentration does not influence adipose tissue and hepatic lipid kinetics in women.

Objective: Increased triglyceride (TG) and apolipoprotein B-100 (apoB-100) concentrations in plasma are important risk factors for cardiovascular disease in women. Administration of some estrogen preparations raises plasma TG and apoB-100 concentrations by increasing hepatic very low-density lipoprotein (VLDL) TG and apoB-100 secretion rates. However, the influence of physiological variation in endogenous estradiol on VLDL-TG and VLDL-apoB-100 metabolism and on free fatty acid (FFA) release into plasma (the major source of fatty acids for VLDL-TG production) is not known. Design and methods: We measured basal VLDL-TG, VLDL-apoB-100, and plasma FFA kinetics by using stable isotopically labeled tracers in 36 eumenorrheic, premenopausal women (age: 33 ± 2 years, BMI: 31 ± 1 kg/m2; mean ± s.e.m.) during the follicular phase of the menstrual cycle; participants were divided into two groups based on low (n = 18) or high (n = 18) plasma estradiol concentrations (defined as below or above the median value of 140 pmol/L in the whole group). Results: Mean plasma estradiol concentration was >3-fold higher in the high-estradiol than in the low-estradiol group (299 ± 37 and 96 ± 7 pmol/L, P < 0.001); there was no difference in plasma progesterone concentrations between the two groups (P = 0.976). There were no significant differences in plasma FFA concentration, FFA rate of appearance in plasma, VLDL-TG and VLDL-apoB-100 concentrations, hepatic VLDL-TG and VLDL-apoB-100 secretion rates, VLDL-TG and VLDL-apoB-100 plasma clearance rates, and mean residence times (all P ≥ 0.45). No significant associations were found between plasma estradiol concentration and FFA, VLDL-TG, and VLDL-apoB-100 concentrations and kinetics (all P > 0.19). Conclusions: Plasma estradiol concentration is not an important correlate of basal plasma FFA, VLDL-TG, and VLDL-apoB-100 kinetics in premenopausal women.

Sortilin restricts secretion of apolipoprotein B-100 by hepatocytes under stressed but not basal conditions.

Genetic variants at the SORT1 locus in humans, which cause increased SORT1 expression in the liver, are significantly associated with reduced plasma levels of LDL cholesterol and apolipoprotein B (apoB). However, the role of hepatic sortilin remains controversial, as genetic deletion of sortilin in mice has resulted in variable and conflicting effects on apoB secretion. Here, we found that Sort1-KO mice on a chow diet and several Sort1-deficient hepatocyte lines displayed no difference in apoB secretion. When these models were challenged with high-fat diet or ER stress, the loss of Sort1 expression resulted in a significant increase in apoB-100 secretion. Sort1-overexpression studies yielded reciprocal results. Importantly, carriers of SORT1 variant with diabetes had larger decreases in plasma apoB, TG, and VLDL and LDL particle number as compared with people without diabetes with the same variants. We conclude that, under basal nonstressed conditions, loss of sortilin has little effect on hepatocyte apoB secretion, whereas, in the setting of lipid loading or ER stress, sortilin deficiency leads to increased apoB secretion. These results are consistent with the directionality of effect in human genetics studies and suggest that, under stress conditions, hepatic sortilin directs apoB toward lysosomal degradation rather than secretion, potentially serving as a quality control step in the apoB secretion pathway in hepatocytes.

Apo(a) and ApoB Interact Noncovalently Within Hepatocytes: Implications for Regulation of Lp(a) Levels by Modulation of ApoB Secretion.

BACKGROUND: Elevated plasma Lp(a) (lipoprotein(a)) levels are associated with increased risk for atherosclerotic cardiovascular disease and aortic valve stenosis. However, the cell biology of Lp(a) biosynthesis remains poorly understood, with the locations of the noncovalent and covalent steps of Lp(a) assembly unclear and the nature of the apoB-containing particle destined for Lp(a) unknown. We, therefore, asked if apo(a) and apoB interact noncovalently within hepatocytes and if this impacts Lp(a) biosynthesis. METHODS: Using human hepatocellular carcinoma cells expressing 17K (17 kringle) apo(a), or a 17KΔLBS7,8 variant with a reduced ability to bind noncovalently to apoB, we performed coimmunoprecipitation, coimmunofluorescence, and proximity ligation assays to document intracellular apo(a):apoB interactions. We used a pulse-chase metabolic labeling approach to measure apo(a) and apoB secretion rates. RESULTS: Noncovalent complexes containing apo(a)/apoB are present in lysates from cells expressing 17K but not 17KΔLBS7,8, whereas covalent apo(a)/apoB complexes are absent from lysates. 17K and apoB colocalized intracellularly, overlapping with staining for markers of endoplasmic reticulum trans-Golgi, and early endosomes, and less so with lysosomes. The 17KΔLBS7,8 had lower colocalization with apoB. Proximity ligation assays directly documented intracellular 17K/apoB interactions, which were dramatically reduced for 17KΔLBS7,8. Treatment of cells with PCSK9 (proprotein convertase subtilisin/kexin type 9) enhanced, and lomitapide reduced, apo(a) secretion in a manner dependent on the noncovalent interaction between apo(a) and apoB. Apo(a) secretion was also reduced by siRNA-mediated knockdown of APOB. CONCLUSIONS: Our findings explain the coupling of apo(a) and Lp(a)-apoB production observed in human metabolic studies using stable isotopes as well as the ability of agents that inhibit apoB biosynthesis to lower Lp(a) levels.

Liver damage promotes pro-inflammatory T-cell responses against apolipoprotein B-100.

OBJECTIVES: Liver-derived apolipoprotein B-100 (ApoB100) is an autoantigen that is recognized by atherogenic CD4+ T cells in cardiovascular disease (CVD). CVD is a major mortality risk for patients with chronic inflammatory liver diseases. However, the impact of liver damage for ApoB100-specific T-cell responses is unknown. METHODS: We identified ApoB100-specific T cells in blood from healthy controls, nonalcoholic fatty liver disease (NAFLD) patients, and CVD patients by activation-induced marker expression and analyzed their differentiation pattern in correlation to the lipid profile and liver damage parameters in a cross-sectional study. To assess the induction of extrahepatic ApoB100-specific T cells upon transient liver damage in vivo, we performed hydrodynamic tail vein injections with diphtheria toxin A (DTA)-encoding plasmid in human ApoB100-transgenic mice. RESULTS: Utilizing immunodominant ApoB100-derived peptides, we found increased ApoB100-specific T-cell populations in NAFLD and CVD patients compared to healthy controls. In a peptide-specific manner, ApoB100 reactivity in healthy controls was accompanied by expression of the regulatory T (Treg)-cell transcription factor FOXP3. In contrast, FOXP3 expression decreased, whereas expression of pro-inflammatory cytokine interleukin (IL)-17A increased in ApoB100-specific T cells from NAFLD and CVD patients. Dyslipidemia and liver damage parameters in blood correlated with reduced FOXP3 expression and elevated IL-17A production in ApoB100-specific T-cell populations, respectively. Moreover, DTA-mediated transient liver damage in human ApoB100-transgenic mice accumulated IL-17a-expressing ApoB100-specific T cells in the periphery. CONCLUSION: Our results show that liver damage promotes pro-inflammatory ApoB100-specific T-cell populations, thereby providing a cellular mechanism for the increased CVD risk in liver disease patients.

Aqueous proteins help predict the response of patients with neovascular age-related macular degeneration to anti-VEGF therapy.

BackgroundTo reduce the treatment burden for patients with neovascular age-related macular degeneration (nvAMD), emerging therapies targeting vascular endothelial growth factor (VEGF) are being designed to extend the interval between treatments, thereby minimizing the number of intraocular injections. However, which patients will benefit from longer-acting agents is not clear.MethodsEyes with nvAMD (n = 122) underwent 3 consecutive monthly injections with currently available anti-VEGF therapies, followed by a treat-and-extend protocol. Patients who remained quiescent 12 weeks from their prior treatment entered a treatment pause and were switched to pro re nata (PRN) treatment (based on vision, clinical exam, and/or imaging studies). Proteomic analysis was performed on aqueous fluid to identify proteins that correlate with patients' response to treatment.ResultsAt the end of 1 year, 38 of 122 eyes (31%) entered a treatment pause (≥30 weeks). Conversely, 21 of 122 eyes (17%) failed extension and required monthly treatment at the end of year 1. Proteomic analysis of aqueous fluid identified proteins that correlated with patients' response to treatment, including proteins previously implicated in AMD pathogenesis. Interestingly, apolipoprotein-B100 (ApoB100), a principal component of drusen implicated in the progression of nonneovascular AMD, was increased in treated patients who required less frequent injections. ApoB100 expression was higher in AMD eyes compared with controls but was lower in eyes that develop choroidal neovascularization (CNV), consistent with a protective role. Accordingly, mice overexpressing ApoB100 were partially protected from laser-induced CNV.FundingThis work was supported by the National Eye Institute, National Institutes of Health grants R01EY029750, R01EY025705, and R01 EY27961; the Research to Prevent Blindness, Inc.; the Alcon Research Institute; and Johns Hopkins University through the Robert Bond Welch and Branna and Irving Sisenwein professorships in ophthalmology.ConclusionAqueous biomarkers could help identify patients with nvAMD who may not require or benefit from long-term treatment with anti-VEGF therapy.

Hsp40s play distinct roles during the initial stages of apolipoprotein B biogenesis.

Apolipoprotein B (ApoB) is the primary component of atherogenic lipoproteins, which transport serum fats and cholesterol. Therefore elevated levels of circulating ApoB are a primary risk factor for cardiovascular disease. During ApoB biosynthesis in the liver and small intestine under nutrient-rich conditions, ApoB cotranslationally translocates into the endoplasmic reticulum (ER) and is lipidated and ultimately secreted. Under lipid-poor conditions, ApoB is targeted for ER-associated degradation (ERAD). Although prior work identified select chaperones that regulate ApoB biogenesis, the contributions of cytoplasmic Hsp40s are undefined. To this end, we screened ApoB-expressing yeast and determined that a class A ER-associated Hsp40, Ydj1, associates with and facilitates the ERAD of ApoB. Consistent with these results, a homologous Hsp40, DNAJA1, functioned similarly in rat hepatoma cells. DNAJA1-deficient cells also secreted hyperlipidated lipoproteins in accordance with attenuated ERAD. In contrast to the role of DNAJA1 during ERAD, DNAJB1-a class B Hsp40-helped stabilize ApoB. Depletion of DNAJA1 and DNAJB1 also led to opposing effects on ApoB ubiquitination. These data represent the first example in which different Hsp40s exhibit disparate effects during regulated protein biogenesis in the ER and highlight distinct roles that chaperones can play on a single ERAD substrate.

Short-Term Effects of Supplemental L-Arginine, Diosmin, Troxerutin, and Hesperidin in Diabetic Patients: A Pilot Study.

BACKGROUND AND AIMS: Inflammatory, oxidative stress, and endothelial dysfunction play a key role in the pathogenesis of long-term cardiovascular complications in patients with diabetes. The present observational prospective study is aimed at evaluating the effects of micronutrients and phytochemicals contained in the dietary supplement Flebotrofine® (AMNOL Chimica Biologica) on biochemical markers of inflammation, endothelial dysfunction, and glycemic control in patients with diabetes. METHODS: 105 type 1 or type 2 diabetes patients regularly took a daily dose of the dietary supplement Flebotrofine® for three consecutive months, and haematological and biochemical parameters were checked at baseline, after three months of treatment, and one month after its suspension. Statistical comparison of the laboratory parameters was performed using the two-tailed ANOVA test for repeated samples with a statistical significance level set at p < 0.05. RESULTS: The daily use of Flebotrofine® did not change the glycemic metabolic compensation of enrolled patients. After three months of regular Flebotrofine® intake, the plasma levels of the antioxidant ß-carotene and of arginine were significantly higher compared with the baseline values, with a decrease in the ADMA/arginine ratio. In contrast, apolipoprotein B, ApoB/ApoA1 ratio, and platelet and leukocyte counts significantly dropped. CONCLUSION: The daily use of Flebotrofine® might be a valid supplement of arginine, the precursor of NO, and essential in the prevention of endothelial dysfunction. The regular intake of arginine and phytochemicals also improved the antioxidant and antithrombotic profile of enrolled patients. Therefore, Flebotrofine® could be a useful dietary supplement to prevent long-term complications in patients with diabetes.

GP73 is a TBC-domain Rab GTPase-activating protein contributing to the pathogenesis of non-alcoholic fatty liver disease without obesity.

The prevalence of non-obese nonalcoholic fatty liver disease (NAFLD) is increasing worldwide with unclear etiology and pathogenesis. Here, we show GP73, a Golgi protein upregulated in livers from patients with a variety of liver diseases, exhibits Rab GTPase-activating protein (GAP) activity regulating ApoB export. Upon regular-diet feeding, liver-GP73-high mice display non-obese NAFLD phenotype, characterized by reduced body weight, intrahepatic lipid accumulation, and gradual insulin resistance development, none of which can be recapitulated in liver-GAP inactive GP73-high mice. Common and specific gene expression signatures associated with GP73-induced non-obese NAFLD and high-fat diet (HFD)-induced obese NAFLD are revealed. Notably, metformin inactivates the GAP activity of GP73 and alleviates GP73-induced non-obese NAFLD. GP73 is pathologically elevated in NAFLD individuals without obesity, and GP73 blockade improves whole-body metabolism in non-obese NAFLD mouse model. These findings reveal a pathophysiological role of GP73 in triggering non-obese NAFLD and may offer an opportunity for clinical intervention.

Effects of apolipoprotein B on lifespan and risks of major diseases including type 2 diabetes: a mendelian randomisation analysis using outcomes in first-degree relatives.

BACKGROUND: Apolipoprotein B (apoB) is emerging as the crucial lipoprotein trait for the role of lipoprotein lipids in the aetiology of coronary heart disease. In this study, we evaluated the effects of genetically predicted apoB on outcomes in first-degree relatives. METHODS: Data on lipoprotein lipids and disease outcomes in first-degree relatives were obtained from the UK Biobank study. We did a univariable mendelian randomisation analysis using a weighted genetic instrument for apoB. For outcomes with which apoB was associated at a false discovery rate (FDR) of less than 5%, multivariable mendelian randomisation analyses were done, including genetic instruments for LDL cholesterol and triglycerides. Associations between apoB and self-reported outcomes in first-degree relatives were characterised for 12 diseases (including heart disease, stroke, and hypertension) and parental vital status together with age at death. Estimates were inferred causal effects per 1 SD elevated lipoprotein trait (for apoB, 1 SD=0·24 g/L). Replication of estimates for lifespan and type 2 diabetes was done using conventional two-sample mendelian randomisation with summary estimates from genome-wide association study consortia. FINDINGS: In univariable mendelian randomisation, genetically elevated apoB in participants was identified to lead to a shorter lifespan in parents (fathers: 0·89 years of life lost per 1 SD higher apoB in offspring, 95% CI 0·63-1·16, FDR-adjusted p=4·0 × 10-10; mothers: 0·48 years of life lost per 1 SD higher apoB in offspring, 0·25-0·71, FDR-adjusted p=1·7 × 10-4). The effects were strengthened to around 2 years of life lost in multivariable mendelian randomisation and were replicated in conventional two-sample mendelian randomisation (odds ratio [OR] of surviving to the 90th centile of lifespan: 0·38 per 1 SD higher apoB in offspring, 95% CI 0·22-0·65). Genetically elevated apoB caused higher risks of heart disease in all first-degree relatives and a higher risk of stroke in mothers. Findings in first-degree relatives were replicated in two-sample multivariable mendelian randomisation, which identified apoB to increase (OR 2·32 per 1 SD higher apoB, 95% CI 1·49-3·61) and LDL cholesterol to decrease (0·34 per 1 SD higher LDL cholesterol, 0·21-0·54) the risk of type 2 diabetes. INTERPRETATION: Higher apoB shortens lifespan, increases risks of heart disease and stroke, and in multivariable analyses that account for LDL cholesterol, increases risk of diabetes. FUNDING: British Heart Foundation, UK Medical Research Council, and UK Research and Innovation.

Ellagic Acid Suppresses ApoB Secretion and Enhances ApoA-1 Secretion from Human Hepatoma Cells, HepG2.

The effect of ellagic acid (EA), a naturally occurring polyphenolic compound, on the secretion of apolipoproteins from human hepatocytes, HepG2, was investigated. The levels of apoB and apoA-1 secreted in the cell culture medium were determined by sandwich ELISA. EA did not affect cell viability at the tested concentrations (up to 50 µM). EA suppressed the secretion of apoB and enhanced that of apoA-1 from HepG2 cells. However, cellular apoB levels were increased, suggesting that EA inhibited the trafficking of apoB during the process of secretion. In contrast, the increase in the cellular levels of apoA-1 was consistent with its secreted levels. These results indicate that EA inhibits the secretion of apoB from hepatocytes and increases the secretion of apoA-1. Both of these effects are beneficial for lipoprotein metabolism in the prevention of lifestyle-related diseases. The detailed mechanism underlying these effects of EA on lipoprotein metabolism should be elucidated in the future, but this naturally occurring polyphenolic compound might be antihyperlipidemic. Based on these results, EA is suggested as a candidate food-derived compound for the prevention of hyperlipidemia.

Down-regulation of RBP4 indicates a poor prognosis and correlates with immune cell infiltration in hepatocellular carcinoma.

Recent research has indicated that metabolically related genes play crucial roles in the pathogenesis of hepatocellular carcinoma (HCC). We evaluated the associations between novel biomarkers and retinol-binding protein 4 (RBP4) for predicting clinical HCC outcomes, hub-related genes, pathway regulation, and immune cells infiltration. Bioinformatic analyses based on data from The Cancer Genome Atlas were performed using online analysis tools. RBP4 expression was low in HCC and was also down-regulated in pan-cancers compared with normal tissues. RBP4 expression was also significantly different based on age (41-60 years old versus 61-80 years old), and low RBP4 expression levels were associated with advanced tumor stages and grades. Higher RBP4 expression was associated with better overall survival time in HCC patients, and we identified a deletion-mutation rate of 1.4% in RBP4. We also identified ten co-expressed genes most related to RBP4 and explored the relationships between six hub genes (APOB, FGA, FGG, SERPINC1, APOA1, and F2) involved in RBP4 regulation. A pathway enrichment analysis for RBP4 indicated complement and coagulation cascades, metabolic pathways, antibiotic biosynthesis pathways, peroxisome proliferator-activated receptor signaling pathways, and pyruvate metabolism pathways. These results suggest that RBP4 may be a novel biomarker for HCC prognosis, and an indicator of low immune response to the disease.

Changes in PCSK 9 and apolipoprotein B100 in Niemann-Pick disease after enzyme replacement therapy with olipudase alfa.

BACKGROUND: Enzyme replacement therapy (ERT) with olipudase alfa, a recombinant human acid sphingomyelinase (rhASM), is being developed to treat patients with ASM deficiency (ASMD), commonly known as Niemann-Pick disease (NPD) types A or B. This study assessed the effect of ERT on lipid parameters and inflammatory markers. METHODS: Serum and plasma samples from five adults with NPD type B (NPD-B) who received olipudase alfa ERT for 26 weeks were analysed. We also collected fasting blood samples from fifteen age- and sex-matched participants as reference and comparison group. We measured fasting lipid profile, apolipoproteins B48 and B100 (apoB48 and apoB100), apolipoprotein A1 (apoA1), proprotein convertase subtilisin/klexin type 9 (PCSK9) mass, oxidised low-density lipoprotein (oxLDL), small dense low-density lipoprotein cholesterol (sdLDL-C) and tumour necrosis factor α (TNF-α). RESULTS: Patients with NPD-B, compared with age and sex matched reference group, had higher triglycerides, PCSK9, apoB48, oxLDL and TNF-α and lower high density lipoprotein cholesterol (HDL-C) and apoA1. Treatment with ERT was associated with improved lipid parameters including total cholesterol, triglycerides, low density lipoprotein cholesterol (LDL-C), sdLDL-C, oxLDL and apoB100. Though there was an increase in apoA1, HDL-C was slightly reduced. TNF-α showed a reduction. ApoB100 decreased in parallel with a decrease in total serum PCSK9 mass after ERT. CONCLUSION: This study demonstrated that patients with NPD-B had a proatherogenic lipid profile and higher circulating TNF-α compared to reference group. There was an improvement in dyslipidaemia after olipudase alfa. It was possible that reductions in LDL-C and apoB100 were driven by reductions in TNF-α and PCSK9 following ERT.

The Apolipoprotein B/A1 Ratio is Associated With Metabolic Syndrome Components, Insulin Resistance, Androgen Hormones, and Liver Enzymes in Women With Polycystic Ovary Syndrome.

Aim: To evaluate the association between the apolipoprotein B/A1 ratio (ApoB/ApoA1) and metabolic and endocrine parameters in women with polycystic ovary syndrome (PCOS). Methods: This study was a secondary analysis of the Acupuncture and Clomiphene for Chinese Women with Polycystic Ovary Syndrome trial (PCOSAct), and 957 subjects with available ApoB and ApoA1 measurements were included. Tests for linear trends and linear regression were used to assess the relation between the ApoB/ApoA1 ratio and metabolic and endocrine parameters. Logistic regression was used to estimate the association between the ratio and risk of metabolic syndrome (MetS) and insulin resistance (IR). The receiver operating characteristics (ROC) curve was used to determine the predictive value of the ApoB/ApoA1 ratio for MetS and IR. Results: The results showed that the ApoB/ApoA1 ratio was positively associated with waist circumference, systolic blood pressure, total cholesterol, triglycerides, low-density lipoprotein, fasting plasma glucose, fasting insulin, homeostatic model assessment-insulin resistance, high free testosterone, high free androgen index, alanine transferase, aspartate transferase, and higher prevalence of MetS and IR, but was negatively correlated with high-density lipoprotein and sex hormone-binding globulin after adjusting for age and body mass index. Logistic regression showed that compared with the ApoB/ApoA1 ratio in first quartile, those in the fourth quartile demonstrated a higher risk of MetS (OR: 24.48, 95%CI: 8.54-70.15, P trend <0.001) and IR (OR: 1.78, 95%CI: 1.10-2.87, P trend <0.05) after adjusting for confounding factors. ROC curve results showed that the AUCMetS was 0.84 (95%CI: 0.81-0.86) and had 86.8% sensitivity and 70.3% specificity with a threshold value of 0.64, and the AUCIR was 0.68 (95%CI: 0.64-0.71) and had 74.3% sensitivity and 58.2% specificity with a threshold value of 0.56. Conclusions: Increased ApoB/ApoA1 ratio was associated with worse MetS components, IR, and elevated androgen hormones and liver enzymes. The ratio might be a useful tool to screen for MetS and IR in PCOS patients.