Reduced Plasma Triglyceride Research Articles

Interventional hepatic apoC-III knockdown improves atherosclerotic plaque stability and remodeling by triglyceride lowering.

Apolipoprotein C-III (apoC-III) is a critical regulator of triglyceride metabolism and correlates positively with hypertriglyceridemia and cardiovascular disease (CVD). It remains unclear if therapeutic apoC-III lowering reduces CVD risk and if the CVD correlation depends on the lipid-lowering or antiinflammatory properties. We determined the impact of interventional apoC-III lowering on atherogenesis using an apoC-III antisense oligonucleotide (ASO) in 2 hypertriglyceridemic mouse models where the intervention lowers plasma triglycerides and in a third lipid-refractory model. On a high-cholesterol Western diet apoC-III ASO treatment did not alter atherosclerotic lesion size but did attenuate advanced and unstable plaque development in the triglyceride-responsive mouse models. No lesion size or composition improvement was observed with apoC-III ASO in the lipid-refractory mice. To circumvent confounding effects of continuous high-cholesterol feeding, we tested the impact of interventional apoC-III lowering when switching to a cholesterol-poor diet after 12 weeks of Western diet. In this diet switch regimen, apoC-III ASO treatment significantly reduced plasma triglycerides, atherosclerotic lesion progression, and necrotic core area and increased fibrous cap thickness in lipid-responsive mice. Again, apoC-III ASO treatment did not alter triglyceride levels, lesion development, and lesion composition in lipid-refractory mice after the diet switch. Our findings suggest that interventional apoC-III lowering might be an effective strategy to reduce atherosclerosis lesion size and improve plaque stability when lipid lowering is achieved.

Orally administered zingerone does not mitigate alcohol-induced hepatic oxidative stress in growing Sprague Dawley rat pups

Neonatal alcohol exposure (NAE) can induce oxidative stress. We determined whether zingerone (ZO), a phytochemical with anti-oxidant activity, can mitigate the negative impact of neonatal alcohol-induced oxidative stress. Seventy ten-day-old Sprague-Dawley rat pups (35 male, 35 female) were randomly assigned and administered the following treatment regimens daily from postnatal day (PND) 12–21: group 1 – nutritive milk (NM), group 2 – NM +1 g/kg ethanol (Eth), group 3 – NM + 40 mg/kg ZO, group 4 – NM + Eth + ZO. Growth performance, blood glucose and plasma triglycerides (TGs), total cholesterol, HDL-cholesterol, leptin and insulin concentration were determined. Cytochrome p450E21(CYP2E1) and thiobarbituric acid (TBARS); markers of hepatic oxidative stress and catalase, superoxide dismutase (SOD) and total glutathione (GSH), anti-oxidant markers of the pups were determined. Oral administration of ethanol (NM + Eth), zingerone (NM + ZO) and combined ethanol and zingerone (NM + Eth + ZO) did not affect the growth performance and insulin and leptin concentration of the rats (p > 0.05). Ethanol significantly reduced plasma TGs concentration of female rats (p = 0.04 vs control). However, ethanol and/or its combination with zingerone decreased hepatic GSH (p = 0.02 vs control) and increased CYP2E1 (p = 0.0002 vs control) activity in male rat pups. Zingerone had no effect (p > 0.05 vs control) on the rats' CYP2E1, GSH, SOD and catalase activities. Neonatal alcohol administration elicited hepatic oxidative stress in male rat pups only, showing sexual dimorphism. Zingerone (NM + ZO) prevented an increase in CYP2E1 activity and a decrease in GSH concentration but did not prevent the alcohol-induced hepatic oxidative stress in the male rat pups.

ANGPTL4 silencing via antisense oligonucleotides reduces plasma triglycerides and glucose in mice without causing lymphadenopathy

Angiopoietin-like 4 (ANGPTL4) is an important regulator of plasma triglyceride (TG) levels and an attractive pharmacological target for lowering plasma lipids and reducing cardiovascular risk. Here, we aimed to study the efficacy and safety of silencing ANGPTL4 in the livers of mice using hepatocyte-targeting GalNAc-conjugated antisense oligonucleotides (ASOs). Compared with injections with negative control ASO, four injections of two different doses of ANGPTL4 ASO over 2 weeks markedly downregulated ANGPTL4 levels in liver and adipose tissue, which was associated with significantly higher adipose LPL activity and lower plasma TGs in fed and fasted mice, as well as lower plasma glucose levels in fed mice. In separate experiments, injection of two different doses of ANGPTL4 ASO over 20 weeks of high-fat feeding reduced hepatic and adipose ANGPTL4 levels but did not trigger mesenteric lymphadenopathy, an acute phase response, chylous ascites, or any other pathological phenotypes. Compared with mice injected with negative control ASO, mice injected with ANGPTL4 ASO showed reduced food intake, reduced weight gain, and improved glucose tolerance. In addition, they exhibited lower plasma TGs, total cholesterol, LDL-C, glucose, serum amyloid A, and liver TG levels. By contrast, no significant difference in plasma alanine aminotransferase activity was observed. Overall, these data suggest that ASOs targeting ANGPTL4 effectively reduce plasma TG levels in mice without raising major safety concerns.

1354-P: Alternate-Day Intermittent Fasting Improves Diabetes and Protects Beta-Cell Function in Polygenic Mouse Models of T2DM

Type 2 diabetes (T2DM) , caused by the interaction of multiple genes and environmental factors, is characterized by hyperglycemia, insulin secretion deficiency and insulin resistance. Chronic hyperglycemia induces β-cell dysfunction and loss of β-cell mass/identity, increased apoptosis and β-cell dedifferentiation. Intermittent fasting (IF) , either alternate-day fasting (ADF) or time-restricted feeding, commonly used regimens for weight-loss, also induces metabolic benefits including reduced blood glucose, improved insulin sensitivity, reduced adiposity, inflammation and oxidative-stress, and increased fatty-acid oxidation; however, the mechanisms underlying these effects remain elusive. KK and KKAy, mouse models of polygenic T2DM spontaneously develop hyperglycemia, glucose intolerance, glucosuria, impaired insulin secretion and insulin resistance. To determine the long-term effects of IF on T2DM, 6-weeks old KK and KKAy mice were subjected to ADF for 16-weeks. While KKAy mice fed ad-libitum demonstrated severe hyperglycemia (˜500mg/dL) , increased plasma insulin, impaired glucose tolerance and insulin resistance at 8 weeks of age, KK mice showed blood glucose levels of ˜200mg/dL, but progressively became as severely diabetic as KKAy mice by 22-weeks. Strikingly, both KK and KKAy mice subjected to ADF showed reduced blood glucose and plasma insulin levels, decreased body weight gain despite increased food intake, reduced plasma triglycerides and cholesterol, and improved insulin sensitivity. They also demonstrated enhanced expression of the β-cell transcription factors PDX1 and NKX6.1, suggesting protection from loss of β-cell identity/dedifferentiation by IF. In addition, respiratory exchange ratio (RER) and movement were significantly enhanced in the ADF group, indicating peripheral benefits of IF. These results have important implications as an optional intervention for preservation of β-cell mass and function in T2DM. Disclosure S.Patel: None. Z.Yan: None. M.S.Remedi: None. Funding National Institutes of Health (R01DK123163)

Combined Effects of High-Intensity Aerobic Exercise Training and Ziziphus jujuba Extract on Tissue Nesfatin-1 in Rats.

Nesfatin-1 is involved in metabolic/feeding regulation and prevention of cardiovascular disease. Previous studies have shown that exercise and herb supplementation can influence nesfatin-1 concentration. The present study investigated the effects of high-intensity training (HIT) and Ziziphus jujuba (ZJ) extract on tissue nesfatin-1 in rats. Twenty-eight female rats were randomly assigned to one of four groups i.e. 1) Saline-Control (SC), 2) Saline-High Intensity Training (ST), 3) Ziziphus jujuba-Control (ZJC), and 4) Ziziphus jujuba-High Intensity Training (ZJT). Rats performed exercise on a treadmill and/or administered supplements intragastrically for 6 weeks, depending on group category. Seventy-two hours after the last training session, rats were anesthetized. Blood, hypothafi 2lamus tissue, heart and gastrocnemius muscles were sent to the laboratory for analyses. Significantly higher nesfatin-1 gene expression and concentration and ATP concentration were found in trained rat. HIT increased plasma High Density Lipoprotein (HDL) and insulin concentration and reduced plasma Triglyceride (TG) and cortisol. ZJ increased tissue nesftain-1 gene expression and concentration while only increasing heart ATP. The combination of exercise and ZJ showed an additive effect compared to each intervention alone on hypothalamus, heart and gastrocnemius NUCB2 gene expression, heart and gastrocnemius nesfatin-1 concentration, plasma HDL and cortisol concentration. The authors recommend both interventions as a means to improve cardiovascular health in rats with further work needed to confirm similar findings in homo sapiens.

Role of Intestinal Stearoyl‐CoA Desaturase 1 in Whole‐Body Lipid Metabolism and Metabolic Health

Stearoyl‐CoA desaturase 1 (SCD1) is an ER‐resident enzyme that converts saturated fatty acids into monounsaturated fatty acids (MUFAs). These MUFAs are the preferred substrates for the synthesis of lipid species such as cholesterol esters and triglycerides. The intestine plays an important role in both lipid absorption and assimilation. Cholesterol and free fatty acids must be esterified into cholesterol esters (CE) and triglycerides (TG), respectively, for efficient secretion of chylomicrons for eventual lipid assimilation. Given the important role of SCD1 in modulating the esterification of both CE and TG, we hypothesized that intestinal SCD1 may regulate lipid secretion from the intestine. Our studies show that SCD1 protein and gene increases along the length of the intestine and this expression is induced by fasting followed by refeeding a high sucrose very low‐fat diet. To study the role of intestinal SCD1 in lipid metabolism, intestine‐specific knockout (iKO) mice were generated by crossing SCD1 floxed mice with mice expressing Cre recombinase under the control of the villin promoter. Our studies reveal that these mice have a 13% reduction in steady‐state plasma lipids, with a specific reduction in plasma triglycerides and free cholesterol. They also have reduced hepatic diacylglycerols and cholesterol esters, with a particular reduction in species containing the MUFA myristoleic acid (14:1). iKO mice have altered lipid composition in the distal intestine, where SCD1 is more highly expressed and regulated. In addition to these alterations in lipid handling, iKO mice have increases in plasma and hepatic bile acids, potentially due to an inability to efficiently esterify cholesterol. Downstream signaling via bile acids is elevated in iKO mice, resulting in activation of TGR5 signaling in brown adipose tissue and the ileum. Thus, our results indicate that deletion of intestinal SCD1 has significant impacts on whole‐body energy balance and lipid metabolism.

Synthetic Cannabidiol Reduced Body Mass and Visceral Adiposity but Not Blood Pressure in Rats With Advanced Metabolic Syndrome

Very little is known about the effects of CBD on age‐associated cardiovascular and metabolic dysfunction. The prevalence of metabolic syndrome (MetS) is 54.9% among people ≥60 years of age, which increases the risk of the development of associated pathologies like cardiovascular disease and type II diabetes. Additionally, synthetic CBD alternatives offer a chemically pure product, which is free of plant‐associated contaminants. To assess the effects of the synthetic CBD, H4CBD, on advanced metabolic dysfunction, a cohort of 41‐week‐old lean strain Long‐Evans Tokushima Otsuka (LETO) rats (n=8) and MetS strain Otsuka Long‐Evans Tokushima Fatty (OLETF) rats were administered sesame oil vehicle (n=8) or 200 mg H4CBD/kg (OLETF n=8) by oral gavage for 4 weeks. Animals were fed standard chow ad libitum and monitored for systolic blood pressure (SBP), activity, body mass (BM), food intake and urinary excretion. After 4 weeks, H4CBD tended to increase activity scores in OLETF (p=0.099) and reduced BM to control levels within the first week. Retroperitoneal and epidydimal fat masses were reduced 25% compared to untreated OLETF. However, the H4CBD‐induced loss of visceral adiposity was not associated with reduced arterial blood pressure. Clinical indicator for muscle wasting, urinary 3‐methylhistidine (3MH), ratioed to urinary creatinine (Cr) (3MH/Cr), suggested the primary mass lost was adipose. An oral glucose tolerance test (oGTT) was performed after 4 weeks of treatment. Glucose response was reduced (AUCglucose) 29% (p<0.001) by H4CBD treatment compared to control and reduced insulin resistance index (IRI) by 25% (p<0.05). However, H4CBD‐induced improvements to glucose tolerance did not associate with reduced basal blood glucose or plasma insulin, despite compensatory increases in insulin receptor (IR) protein expression (54%; p<0.05) and membrane‐bound glucose transporter protein 4 (GLUT4) protein expression (41.5%). The results suggest that a chronic, high dose of CBD may improve glucose tolerance, independent of insulin signaling, without caloric restriction via increased activity. H4CBD reduced plasma triglycerides by 45% (p<0.05) but not plasma NEFA suggesting that de novo TG synthesis was reduced. The results suggest that a chronic, high dose of CBD may drive fat loss via increased activity and reduced TG synthesis, but the mass loss was not sufficient to correct the associated hypertension or impaired substrate metabolism common in MetS for this duration of treatment and advanced stage of dysregulated metabolism.

Glycerol-3-phosphate phosphatase operates a glycerol shunt in pancreatic β-cells that controls insulin secretion and metabolic stress

ObjectiveThe recently identified glycerol-3-phosphate (Gro3P) phosphatase (G3PP) in mammalian cells, encoded by the PGP gene, was shown to regulate glucose, lipid and energy metabolism by hydrolyzing Gro3P and to control glucose-stimulated insulin secretion (GSIS) in β-cells, in vitro. However, whether G3PP regulates β-cell function and insulin secretion in vivo is not known. MethodsWe now examined the role of G3PP in the control of insulin secretion in vivo, β-cell function and glucotoxicity in inducible β-cell specific G3PP-KO (BKO) mice. Inducible BKO mice were generated by crossing floxed-G3PP mice with Mip-Cre-ERT (MCre) mice. All the in vivo studies were done using BKO and control mice fed normal diet and the ex vivo studies were done using pancreatic islets from these mice. ResultsBKO mice, compared to MCre controls, showed increased body weight, adiposity, fed insulinemia, enhanced in vivo GSIS, reduced plasma triglycerides and mild glucose intolerance. Isolated BKO mouse islets incubated at high (16.7 mM), but not at low or intermediate glucose (3 and 8 mM), showed elevated GSIS, Gro3P content as well as increased levels of metabolites and signaling coupling factors known to reflect β-cell activation for insulin secretion. BKO islets also showed reduced glycerol release and increased O2 consumption and ATP production at high glucose only. BKO islets chronically exposed to elevated glucose levels showed increased apoptosis, reduced insulin content and decreased mRNA expression of β-cell differentiation markers, Pdx-1, MafA and Ins-2. ConclusionsThe results demonstrate that β-cells are endowed with a “glycerol shunt”, operated by G3PP that regulates β-cell metabolism, signaling and insulin secretion in vivo, primarily at elevated glucose concentrations. We propose that the glycerol shunt plays a role in preventing insulin hypersecretion and excess body weight gain and contributes to β-cell mass preservation in the face of hyperglycemia.

Abstract P206: Proteomic And Metabolomic Signatures Of Plasma Triglyceride-related Trait Responses To Regular Exercise

Introduction: Elevated plasma triglycerides (TG) are associated with risk of cardiovascular disease and are modifiable through lifestyle interventions such as regular exercise. However, TG responses to regular exercise are characterized by significant inter-individual differences. Hypothesis: We hypothesized that baseline levels of circulating proteins and metabolites are associated with TG response to exercise and can predict exercise-induced changes in plasma TG traits. Methods: We measured circulating proteins (n=4979 proteins) and metabolites (n=300) in 650 Black and White adults of the HERITAGE Family Study who completed 20 weeks of exercise training and had complete data on TG traits. We investigated two TG-related traits that significantly improved with training: fasting TG and large TG-rich lipoprotein particle concentration (LTRLP). The association between baseline analyte values and exercise-induced changes in TG traits were examined using linear mixed models adjusted for age, sex, race, BMI, baseline trait value, and the random effect of family membership. Significance was determined as FDR<0.05. Molecular signatures of TG trait responses were generated via elastic net regression tuned using leave-one-out cross validation. Results: Regular exercise significantly reduced plasma TG (-1.03±1.3 mmol/L, p=0.01) and LTRLP (-0.24±2.2 nmol/L, p= 0.007), with their training-induced changes moderately correlated (r=0.4, p<0.0001). We identified largely distinct panels of baseline analytes associated with changes in TG and LTRLP ( Table 1 ). Similarly, elastic net regression yielded distinct models of 99 analytes for TG and 315 analytes for LTRLP responses to exercise with accuracy of RMSE=1.28 mmol/L for TG and 2.16 nmol/L for the LTRLP model. Conclusions: We identified panels of proteins and metabolites associated with exercise-induced changes in TG traits and demonstrate, within our data, circulating analytes hold promise for predicting the exercise responsiveness of plasma TG-related traits.

Edible Mushrooms as Novel Myco-Therapeutics: Effects on Lipid Level, Obesity and BMI.

Obesity, usually indicated by a body mass index of more than 30 kg/m2, is a worsening global health issue. It leads to chronic diseases, including type II diabetes, hypertension, and cardiovascular diseases. Conventional treatments for obesity include physical activity and maintaining a negative energy balance. However, physical activity alone cannot determine body weight as several other factors play a role in the overall energy balance. Alternatively, weight loss may be achieved by medication and surgery. However, these options can be expensive or have side effects. Therefore, dietary factors, including dietary modifications, nutraceutical preparations, and functional foods have been investigated recently. For example, edible mushrooms have beneficial effects on human health. Polysaccharides (essentially β-D-glucans), chitinous substances, heteroglycans, proteoglycans, peptidoglycans, alkaloids, lactones, lectins, alkaloids, flavonoids, steroids, terpenoids, terpenes, phenols, nucleotides, glycoproteins, proteins, amino acids, antimicrobials, and minerals are the major bioactive compounds in these mushrooms. These bioactive compounds have chemo-preventive, anti-obesity, anti-diabetic, cardioprotective, and neuroprotective properties. Consumption of edible mushrooms reduces plasma triglyceride, total cholesterol, low-density lipoprotein, and plasma glucose levels. Polysaccharides from edible mushrooms suppress mRNA expression in 3T3-L1 adipocytes, contributing to their anti-obesity properties. Therefore, edible mushrooms or their active ingredients may help prevent obesity and other chronic ailments.

Design and synthesis of novel spirocyclic carboxylic acids as potent and orally bioavailable DGAT1 inhibitors and their biological evaluation

A series of novel spirocyclic DGAT1 inhibitors containing the oxadiazole motif were designed and synthesized for biological evaluation. Several compounds exhibited potent diacylglycerol acyltransferase 1 (DGAT1) inhibitory activity. Optimization of the series led to the identification of five lead compounds 8, 9, 10, 11 and 12 that showed excellent in-vitro activity with IC50 values ranging from 7 to 20 nM against human DGAT1. All compounds demonstrated good druggability as well as microsomal stability and safety profiles such as hERG and CYP. Compound 12 significantly reduced plasma triglyceride levels in-vivo in the mouse model of acute lipid challenge. Significant reduction in plasma TG excursion was observed, thus indicating DGAT1 inhibition in-vivo.

Targeting ApoC3 Paradoxically Aggravates Atherosclerosis in Hamsters With Severe Refractory Hypercholesterolemia.

RationaleApoC3 plays a central role in the hydrolysis process of triglyceride (TG)-rich lipoproteins mediated by lipoprotein lipase (LPL), which levels are positively associated with the incidence of cardiovascular disease (CVD). Although targeting ApoC3 by antisense oligonucleotide (ASO), Volanesorsen markedly reduces plasma TG level and increase high-density lipoprotein cholesterol (HDL-C) in patients with hypertriglyceridemia (HTG), the cholesterol-lowering effect of ApoC3 inhibition and then the consequential outcome of atherosclerotic cardiovascular disease (ASCVD) have not been reported in patients of familial hypercholesterolemia (FH) with severe refractory hypercholesterolemia yet.ObjectiveTo investigate the precise effects of depleting ApoC3 on refractory hypercholesterolemia and atherosclerosis, we crossed ApoC3-deficient hamsters with a background of LDLR deficiency to generate a double knockout (DKO) hamster model (LDLR−/−, XApoC3−/−, DKO).Approach and ResultsOn the standard laboratory diet, DKO hamsters had reduced levels of plasma TG and total cholesterol (TC) relative to LDLR−/− hamsters. However, upon high-cholesterol/high-fat (HCHF) diet feeding for 12 weeks, ApoC3 deficiency reduced TG level only in female animals without affecting refractory cholesterol in the circulation, whereas apolipoprotein A1 (ApoA1) levels were significantly increased in DKO hamsters with both genders. Unexpectedly, loss of ApoC3 paradoxically accelerated diet-induced atherosclerotic development in female and male LDLR−/− hamsters but ameliorated fatty liver in female animals. Further analysis of blood biological parameters revealed that lacking ApoC3 resulted in abnormal platelet (PLT) indices, which could potentially contribute to atherosclerosis in LDLR−/− hamsters.ConclusionsIn this study, our novel findings provide new insight into the application of ApoC3 inhibition for severe refractory hypercholesterolemia and ASCVD.

Inactivity Causes Resistance to Improvements in Metabolism After Exercise.

Prolonged sitting prevents a 1-h bout of running from improving fat oxidation and reducing plasma triglycerides. This "exercise resistance" can be prevented by taking 8500 steps·d-1 or by interrupting 8 h of sitting with hourly cycle sprints. We hypothesize that there is an interplay between background physical activity (e.g., steps·d-1) and the exercise stimuli in regulating some acute and chronic adaptations to exercise.

Tyramine exerts hypolipidemic and anti-obesity effects in vivo

Obesity and dyslipidemia are conditions often associated with cardiovascular risk, inflammation, oxidative stress, and death. Thus, a new approach has been highlighted to promote research and development of pharmacological tools derived from natural sources. Among the most widely studied groups of substances, polyphenols such as tyramine stand out. This study investigated hypolipidemic and anti-obesity properties of tyramine. Oral toxicity evaluation, models of dyslipidemia and obesity were used. To induce dyslipidemia, Poloxamer-407 (P-407) was administered intraperitoneally. In the hypercholesterolemic and obesity model, specific diet and oral tyramine were provided. After 24h of P-407 administration, tyramine 2 mg/kg (T2) decreased triglycerides (TG) (2057.0 ± 158.5 mg/dL vs. 2838 ± 168.3 mg/dL). After 48h, TG were decreased by T2 (453.0 ± 35.47 vs. 760.2 ± 41.86 mg/dL) and 4 mg/kg (T4) (605.8 ± 26.61 760.2 ± 41.86 mg/dL). T2 reduced total cholesterol (TC) after 24h (309.0 ± 11.17 mg/dL vs. 399.7 ± 15.7 mg/dL); After 48h, 1 mg/kg (T1) (220.5 ± 12.78 mg/dL), T2 (205.8 ± 7.1 mg/dL) and T4 (216.8 ± 12.79 mg/dL), compared to P-407 (275.5 ± 12.1 mg/dL). The treatment decreased thiobarbituric acid reactive substances and nitrite in liver, increased superoxide dismutase, reduced the diet-induced dyslipidemia, decreasing TC around 15%. Tyramine reduced body mass, glucose, and TC after hypercaloric feed. Treatment with 5 mg/L (0.46 ± 0.04 ng/dL) and 10 mg/L (0.44 ± 0.02 ng/dL) reduced plasma insulin (1.18 ± 0.23 ng/dL). Tyramine increased adiponectin at 5 mg/L (1.02 ± 0.02 vs. 0.83 ± 0.02 ng/mL) and 10mg/L (0.96 ± 0.04 ng/mL). In conclusion, tyramine has low toxicity in rodents, has antioxidant effect, reduces plasma triglycerides and cholesterol levels. However, further studies should be conducted in rodents and non-rodents to better understand the pharmacodynamic and pharmacokinetic properties of tyramine.

(Pro)renin Receptor Inhibition Reduces Plasma Cholesterol and Triglycerides but Does Not Attenuate Atherosclerosis in Atherosclerotic Mice.

Objective: Elevated plasma cholesterol concentrations contributes to ischemic cardiovascular diseases. Recently, we showed that inhibiting hepatic (pro)renin receptor [(P)RR] attenuated diet-induced hypercholesterolemia and hypertriglyceridemia in low-density lipoprotein receptor (LDLR) deficient mice. The purpose of this study was to determine whether inhibiting hepatic (P)RR could attenuate atherosclerosis.Approach and Results: Eight-week-old male LDLR−/− mice were injected with either saline or N-acetylgalactosamine-modified antisense oligonucleotides (G-ASOs) primarily targeting hepatic (P)RR and were fed a western-type diet (WTD) for 16 weeks. (P)RR G-ASOs markedly reduced plasma cholesterol concentrations from 2,211 ± 146 to 1,128 ± 121 mg/dL. Fast protein liquid chromatography (FPLC) analyses revealed that cholesterol in very low-density lipoprotein (VLDL) and intermediate density lipoprotein (IDL)/LDL fraction were potently reduced by (P)RR G-ASOs. Moreover, (P)RR G-ASOs reduced plasma triglyceride concentrations by more than 80%. Strikingly, despite marked reduction in plasma lipid concentrations, atherosclerosis was not reduced but rather increased in these mice. Further testing in ApoE−/− mice confirmed that (P)RR G-ASOs reduced plasma lipid concentrations but not atherosclerosis. Transcriptomic analysis of the aortas revealed that (P)RR G-ASOs induced the expression of the genes involved in immune responses and inflammation. Further investigation revealed that (P)RR G-ASOs also inhibited (P)RR in macrophages and in enhanced inflammatory responses to exogenous stimuli. Moreover, deleting the (P)RR in macrophages resulted in accelerated atherosclerosis in WTD fed ApoE−/− mice.Conclusion: (P)RR G-ASOs reduced the plasma lipids in atherosclerotic mice due to hepatic (P)RR deficiency. However, augmented pro-inflammatory responses in macrophages due to (P)RR downregulation counteracted the beneficial effects of lowered plasma lipid concentrations on atherosclerosis. Our study demonstrated that hepatic (P)RR and macrophage (P)RR played a counteracting role in atherosclerosis.

Ghrelin is a signal to facilitate the utilization of fatty acids and save glucose by the liver, skeletal muscle, and adipose tissues in chicks

Ghrelin, classically known as a central appetite-stimulating hormone, has recently been recognized to play an important role in peripheral tissue energy metabolism. In chicken, contrary to mammal, ghrelin acts as an anorexia signal, increased by fasting and further elevated after refed. In the present study, the effect of ghrelin on glucose/lipid utilization by peripheral tissues was investigated. Injection of exogenous acyl ghrelin reduced plasma triglyceride and glucose levels of chickens at both fasting and fed status. In the in vitro cultured chicken primary hepatocytes, adipocytes, and myoblasts, ghrelin suppressed glucose uptake, stimulated fatty acids uptake and oxidation, and decreased TG content. In hepatocyte, ghrelin increased the activities of LPL and HL, and upregulated the expression levels of gene ACC, CPT1, and PPARα. Ghrelin treatment markedly increased the protein level of p-ACC, PPARγ, PGC1α, and CPT1 in hepatocytes, adipocytes and myoblasts. Inhibition of AMPK activity by Compound C had no influence on glucose uptake by hepatocyte, adipocyte, and myoblast, but further amplified the stimulated fatty acid uptake of adipocyte by ghrelin. The present result demonstrates that ghrelin facilitates the uptake and oxidation of fatty acid and cut down the utilization of glucose by the liver, muscle, and adipose tissues. The result suggests that ghrelin functions as a signal of fatty acid oxidation. The study provides a vital framework for understanding the intrinsic role of ghrelin as a crucial factor in the concerted regulation of metabolic substrate of hepatocytes, adipocytes, and myoblasts.

Lipid Profiles from Dried Blood Spots Reveal Lipidomic Signatures of Newborns Undergoing Mild Therapeutic Hypothermia after Hypoxic-Ischemic Encephalopathy.

Hypoxic-ischemic encephalopathy (HIE) is associated with perinatal brain injury, which may lead to disability or death. As the brain is a lipid-rich organ, various lipid species can be significantly impacted by HIE and these correlate with specific changes to the lipidomic profile in the circulation. Objective: To investigate the peripheral blood lipidomic signature in dried blood spots (DBS) from newborns with HIE. Using univariate analysis, multivariate analysis and sPLS-DA modelling, we show that newborns with moderate–severe HIE (n = 46) who underwent therapeutic hypothermia (TH) displayed a robust peripheral blood lipidomic signature comprising 29 lipid species in four lipid classes; namely phosphatidylcholine (PC), lysophosphatidylcholine (LPC), triglyceride (TG) and sphingomyelin (SM) when compared with newborns with mild HIE (n = 18). In sPLS-DA modelling, the three most discriminant lipid species were TG 50:3, TG 54:5, and PC 36:5. We report a reduction in plasma TG and SM and an increase in plasma PC and LPC species during the course of TH in newborns with moderate–severe HIE, compared to a single specimen from newborns with mild HIE. These findings may guide the research in nutrition-based intervention strategies after HIE in synergy with TH to enhance neuroprotection.

Abstract 9137: Isolated Liver Perfusion and Electron Microscopy Reveal Defect in Vldl Lipidation Associated With Inactivation of Tm6sf2

Background: A missense variant (E167K) in Transmembrane 6 Superfamily Member 2 (TM6SF2) is associated with lower plasma lipid levels and protection from coronary artery disease. Inactivation of Tm6sf2 in mice results in increased hepatic triglyceride (TG) content associated with a reduced rate of secretion of very low-density lipoprotein (VLDL)-TG, but not VLDL-Apolipoprotein B (ApoB). To determine the mechanism responsible for the hypolipidemic effect of Tm6sf2 inactivation, we performed liver perfusion and electron microscopy studies in wildtype (WT) and Tm6sf2 -/- mice. Methods: Ad libitum chow-fed male mice (11-13 weeks old, n=4/group) Tm6sf2 -/- and age- and sex-matched WT mice (C57BL/6N background) were anaesthetized with ketamine/xylazine and treated with heparin (4U/g body weight). Livers were harvested and perfused for 60 min without recirculation with buffer contained free fatty acids (0.8 mM). VLDL was isolated from the perfusate by ultracentrifugation ( d =1.006 g/ml) and VLDL-lipids were assayed using enzymatic assays (Sigma-Aldrich) and liquid chromatography mass-spectrometry (LC-MS). Amounts of VLDL-ApoB and -ApoE were compared between strains by immunoblotting. Electron microscopy was performed on liver sections that had been stained with imidazole/osmium tetroxide to visualize lipoproteins. Results: The secretion rates of VLDL-ApoB and -ApoE did not differ between the Tm6sf2 -/- and WT mice. Hepatic secretion of VLDL lipids (TG, cholesterol, phosphatidylcholine) per ApoB was reduced by 53%, 63% and 67%, respectively, in Tm6sf2 -/- mice compared to WT controls. Electron microscopy showed that the VLDL particles in the secretory pathway of Tm6sf2 -/- mice were smaller than those in the WT animals. Conclusions: Taken together these findings are consistent with TM6SF2 reducing plasma TG and cholesterol levels through limiting addition of neutral lipids to nascent VLDL particles in the hepatocyte prior to secretion.

The Potential of ANGPTL8 Antagonism to Simultaneously Reduce Triglyceride and Increase HDL-Cholesterol Plasma Levels.

Elevated triglyceride (TG) and reduced high-density lipoprotein-cholesterol (HDL-C) plasma levels are risk factors for atherosclerosis and cardiovascular disease. Therefore, a drug that simultaneously reduces TG and increases HDL-C plasma levels has the potential to prevent and treat these diseases. Angiopoietin-like 3 (ANGPTL3) regulates plasma TG and HDL-C levels by inhibiting lipoprotein lipase (LPL) and endothelial lipase (EL), respectively. ANGPTL3 inhibition of LPL requires complex formation with ANGPTL8, which is not required for its inhibition of EL. Therefore, the entire pool of plasma ANGPTL3 can be classified as ANGPTL8-associated ANGPTL3 and ANGPTL8-free ANGPTL3, where the former inhibits LPL and the latter inhibits EL. ANGPTL8 antibodies or inhibitors that block its interactions with ANGPTL3 can disrupt or preclude the ANGPTL3-8 complex formation, resulting in fewer ANGPTL3-8 complexes (reduced LPL inhibition), but more free ANGPTL3 (enhanced EL inhibition). Therefore, ANGPTL8 antagonism increases LPL activity while decreasing EL activity, thus leading to reduced plasma TG while simultaneously increasing HDL-C levels. In humans, carriers of ANGPTL8 truncating variants consistently have lower TG but higher HDL-C levels, supporting this hypothesis.

Recruiting a transcription factor in the liver to prevent atherosclerosis.

Hypertriglyceridemia is associated with obesity, diabetes, and atherosclerosis. While lipoprotein lipase (LPL) hydrolyzes triglyceride (TG) cargo into remnant lipoproteins with atherogenic properties, how remnant lipoprotein clearance relates to atherosclerosis in people with diabetes remains unclear. In this issue of the JCI, Shimizu-Albergine et al. examined the effects of the basic leucine zipper transcription factor CREBH, which induces genes that activate LPL in mouse models of type I diabetes. Overexpression of a CREBH fragment reduced apolipoprotein C3 (APOC3) levels, which reduced plasma TGs. Notably, the TGs were lowered by a mechanism that was independent of LPL, and atherosclerosis was alleviated by enhanced lipoprotein remnant clearance as opposed to increased lipolysis of TG-rich lipoprotein precursors. A proinflammatory mechanism likely underlies the atherogenicity of remnant lipoproteins. These findings suggest that modifying CREBH expression in the liver may ameliorate atherosclerosis and, perhaps, other diabetes complications.