Isotope Labeling and Biochemical Assessment of Liver-Triacylglycerol in Patients with Different Levels of Histologically-Graded Liver Disease.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) prevalence is rapidly growing, and fatty liver has been found in a quarter of the US population. Increased liver lipids, particularly those derived from the pathway of de novo lipogenesis (DNL), have been identified as a hallmark feature in individuals with high liver fat. This has led to much activity in basic science and drug development in this area. No studies to date have investigated the contribution of DNL across a spectrum of disease, although it is clear that inhibition of DNL has been shown to reduce liver fat. OBJECTIVES: The purpose of this study was to determine whether liver lipid synthesis increases across the continuum of liver injury. METHODS: Individuals (n = 49) consumed deuterated water for 10 d before their scheduled bariatric surgeries to label DNL; blood and liver tissue samples were obtained on the day of the surgery. Liver lipid concentrations were quantitated, and levels of protein and gene expression assessed. RESULTS: Increased liver DNL, measured isotopically, was significantly associated with liver fatty acid synthase protein content (R = 0.470, P = 0.003), total steatosis assessed by histology (R = 0.526, P = 0.0008), and the fraction of DNL fatty acids in plasma very low-density lipoprotein-triacylglycerol (R = 0.747, P < 0.001). Regression analysis revealed a parabolic relationship between fractional liver DNL (percent) and NAFLD activity score (R = 0.538, P = 0.0004). CONCLUSION: These data demonstrate that higher DNL is associated with early to mid stages of liver disease, and this pathway may be an effective target for the treatment of NAFLD and nonalcoholic steatohepatitis. This study was registered at clinicaltrials.gov as NCT03683589.

Lymphatics - not just a chylomicron conduit.

PURPOSE OF REVIEW: Lymphatics are known to have active, regulated pumping by smooth muscle cells that enhance lymph flow, but whether active regulation of lymphatic pumping contributes significantly to the rate of appearance of chylomicrons (CMs) in the blood circulation (i.e., CM production rate) is not currently known. In this review, we highlight some of the potential mechanisms by which lymphatics may regulate CM production. RECENT FINDINGS: Recent data from our lab and others are beginning to provide clues that suggest a more active role of lymphatics in regulating CM appearance in the circulation through various mechanisms. Potential contributors include apolipoproteins, glucose, glucagon-like peptide-2, and vascular endothelial growth factor-C, but there are likely to be many more. SUMMARY: The digested products of dietary fats absorbed by the small intestine are re-esterified and packaged by enterocytes into large, triglyceride-rich CM particles or stored temporarily in intracellular cytoplasmic lipid droplets. Secreted CMs traverse the lamina propria and are transported via lymphatics and then the blood circulation to liver and extrahepatic tissues, where they are stored or metabolized as a rich energy source. Although indirect data suggest a relationship between lymphatic pumping and CM production, this concept requires more experimental evidence before we can be sure that lymphatic pumping contributes significantly to the rate of CM appearance in the blood circulation.

Lactation alters the relationship between liver lipid synthesis and hepatic fat stores in the postpartum period.

In mothers who are nursing their infants, increased clearance of plasma metabolites into the mammary gland may reduce ectopic lipid in the liver. No study to date has investigated the role of lactation on liver lipid synthesis in humans, and we hypothesized that lactation would modify fatty acid and glucose handling to support liver metabolism in a manner synchronized with the demands of milk production. Lactating (n = 18) and formula-feeding women (n = 10) underwent metabolic testing at 6-week postpartum to determine whether lactation modified intrahepatic triacylglycerols (IHTGs), measured by proton magnetic resonance spectroscopy. Subjects ingested oral deuterated water to measure fractional de novo lipogenesis (DNL) in VLDL-TG during fasting and during an isotope-labeled clamp at an insulin infusion rate of 10 mU/m2/min. Compared with formula-feeding women, we found that lactating women exhibited lower plasma VLDL-TG concentrations, similar IHTG content and similar contribution of DNL to total VLDL-TG production. These findings suggest that lactation lowers plasma VLDL-TG concentrations for reasons that are unrelated to IHTG and DNL. Surprisingly, we determined that the rate of appearance of nonesterified fatty acids was not related to IHTG in either group, and the expected positive association between DNL and IHTG was only significant in formula-feeding women. Further, in lactating women only, the higher the prolactin concentration, the lower the IHTG, while greater DNL strongly associated with elevations in VLDL-TG. In conclusion, we suggest that future studies should investigate the role of lactation and prolactin in liver lipid secretion and metabolism.

Fatty Acid Synthase Inhibitor TVB-2640 Reduces Hepatic de Novo Lipogenesis in Males With Metabolic Abnormalities.

BACKGROUND AND AIMS: Elevated hepatic de novo lipogenesis (DNL) is a key distinguishing characteristic of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis. In rodent models of NAFLD, treatment with a surrogate of TVB-2640, a pharmacological fatty acid synthase inhibitor, has been shown to reduce hepatic fat and other biomarkers of DNL. The purpose of this phase I clinical study was to test the effect of the TVB-2640 in obese men with certain metabolic abnormalities that put them at risk for NAFLD. APPROACH AND RESULTS: Twelve subjects (mean ± SEM, 42 ± 2 years, body mass index 37.4 ± 1.2 kg/m2 , glucose 103 ± 2 mg/dL, triacylglycerols 196 ± 27 mg/dL, and elevated liver enzymes) underwent 10 days of treatment with TVB-2640 at doses ranging from 50-150 mg/day. Food intake was controlled throughout the study. Hepatic DNL was measured before and after an oral fructose/glucose bolus using isotopic labeling with 1-13 C1 -acetate intravenous infusion, followed by measurement of labeled very low-density lipoprotein palmitate via gas chromatography mass spectometry. Substrate oxidation was measured by indirect calorimetry. Across the range of doses, fasting DNL was reduced by up to 90% (P = 0.003). Increasing plasma concentrations of TVB-2640 were associated with progressive reductions in the percent of fructose-stimulated peak fractional DNL (R2  = -0.749, P = 0.0003) and absolute DNL area under the curve 6 hours following fructose/glucose bolus (R2  = -0.554, P = 0.005). For all subjects combined, alanine aminotransferase was reduced by 15.8 ± 8.4% (P = 0.05). Substrate oxidation was unchanged, and safety monitoring revealed that the drug was well tolerated, without an increase in plasma triglycerides. Alopecia occurred in 2 subjects (reversed after stopping the drug), but otherwise no changes were observed in fasting glucose, insulin, ketones, and renal function. CONCLUSION: These data support the therapeutic potential of a fatty acid synthase inhibitor, TVB-2640 in particular, in patients with NAFLD and nonalcoholic steatohepatitis.

Traditional Versus Suspended Push-up Muscle Activation in Athletes and Sedentary Women.

Syed-Abdul, MM, Soni, DS, Miller, WM, Johnson, RJ, Barnes, JT, Pujol, TJ, and Wagganer, JD. Traditional versus suspended push-up muscle activation in athletes and sedentary women. J Strength Cond Res 32(7): 1816-1820, 2018-Many strength training programs incorporate push-up exercises, which primarily activate upper-body muscles. Past data support the fact that shoulder girdle muscles (i.e., triceps (T) and anterior deltoids [AD]) exhibit greater electromyography (EMG) activity when a push-up is performed on an unstable (i.e., suspended [SP]) vs. stable (i.e., traditional [TD]) surface (). Sixty-nine healthy female volunteers (soccer players [SO], n = 24; gymnasts [GY], n = 21; sedentary [SE], n = 24) performed three TD and three SP push-ups. Muscle activation, expressed as absolute integral (mV), was measured using EMG analysis. Significant increases in muscle activation were exhibited by GY (TD: p < 0.01 and SP: p < 0.001) and SO (TD: p < 0.05 and SP: p < 0.05) compared to SE for the T muscle. Only SO (p < 0.05) exhibited significantly higher muscle activation during the SP versus TD. For the AD, values were significantly higher for SO (TD: p < 0.001 and SP: p < 0.001) and GY (TD: p < 0.01 and SP: p < 0.01) compared to the SE group. In addition, significantly higher values were exhibited by SO compared with GY during TD push-ups (p < 0.01). Both the SO (p < 0.05) and GY (p < 0.05) group exhibited significantly higher values during SP versus TD push-ups. Finally, values were significantly higher for the AD compared to the T muscle only in the SO group during TD (p < 0.01) and SP (p < 0.05) push-ups. Data from this study for trained women (i.e., SO) are consistent with previous studies, whereas for untrained women (i.e., SE) the findings differed during TD and SP push-ups for both muscles. Differences were also observed between female SO and GY are unexplainable and therefore need further investigation.

Glucagon-like peptide-2 mobilization of intestinal lipid does not require canonical enterocyte chylomicron synthetic machinery.

BACKGROUND & AIMS: Dietary triglycerides (TG) retained in the intestine after a meal can be mobilized many hours later by glucagon-like peptide-2 (GLP-2) in humans and animal models, despite the well-documented absence of expression of the GLP-2 receptor on enterocytes. In this study, we examined the site of GLP-2 action to mobilize intestinal lipids and enhance chylomicron production. METHODS: In mesenteric lymph duct-cannulated rats, we assessed GLP-2-stimulated lymph flow rate, TG concentration, TG output, and apoB48 abundance 5 h after an intraduodenal lipid bolus, in the presence of a validated GLP-2 antagonist or vehicle. Additionally, the same GLP-2-stimulated parameters were examined in the presence or absence of cis-Golgi disruption by Brefeldin A (BFA). RESULTS: Compared to placebo, GLP-2 administration increased lymph flow by 2.8-fold (P < 0.001), cumulative lymph volume by 2.69-fold (P < 0.001) and total TG output 2-fold (P = 0.015). GLP-2 receptor antagonism markedly diminished GLP-2's ability to stimulate lymph flow, cumulative lymph volume and total TG output, demonstrating the dependence of GLP-2 stimulation of lymph flow and TG output on its receptor activation. In contrast, disruption of the cis-Golgi apparatus with Brefeldin A did not diminish the GLP-2-response of lymph flow i.e., increased lymph flow by 2.7-fold (P = 0.001), lymph volume by 2.9-fold (P = 0.001), and total TG output i.e., increased by 2.5-fold (P = 0.003). CONCLUSIONS: GLP-2 mobilizes enteral lipid at a site distal to the Golgi, acting via its receptor. Since GLP-2 receptors are not expressed on enterocytes, GLP-2 likely mobilizes intestinal lipid residing extracellularly, either in the lamina propria or in the lymphatics.

The Utility and Diagnostic Accuracy of Transient Elastography in Adults with Morbid Obesity: A Prospective Study.

Patients with morbid obesity are at high risk for nonalcoholic fatty liver disease (NAFLD) complicated by liver fibrosis. The clinical utility of transient elastography (TE) by Fibroscan in patients with morbid obesity (body mass index (BMI) ≥ 40 kg/m2) is not well-defined. We examined the diagnostic accuracy of Fibroscan in predicting significant liver fibrosis (fibrosis stage ≥2) in morbidly obese patients (BMI ≥ 40 kg/m2). Patients scheduled for bariatric surgery were prospectively enrolled. Intraoperative liver biopsy, liver-stiffness measurement (LSM) by Fibroscan (XL probe), and biochemical evaluation were all performed on the same day. The endpoint was significant liver fibrosis defined as fibrosis stage ≥2 based on the Nonalcoholic Steatohepatitis Clinical Research Network. The optimal LSM cutoff value for detecting significant fibrosis was determined by using the Youden Index method. Routine clinical, laboratory, and elastography data were analyzed by stepwise logistic regression analysis to identify predictors of significant liver fibrosis and build a predictive model. An optimal cutoff point of the new model's regression formula for predicting significant fibrosis was determined by using the Youden index method. One hundred sixty-seven patients (mean age, 46.4 years) were included, of whom 83.2% were female. Histological assessment revealed the prevalence of steatohepatitis and significant fibrosis of 40.7% and 11.4%, respectively. The median LSM was found to be significantly higher in the significant fibrosis group compared to those in the no or non-significant fibrosis group (18.2 vs. 7.7 kPa, respectively; p = 0.0004). The optimal LSM cutoff for predicting significant fibrosis was 12.8 kPa, with an accuracy of 71.3%, sensitivity of 73.7%, specificity of 70.9%, positive predictive value of 24.6%, negative predictive value of 95.5%, and ROC area of 0.723 (95% CI: 0.62-0.83). Logistic regression analysis identified three independent predictors of significant fibrosis: LSM, hemoglobin A1c, and alkaline phosphatase. A risk score was developed by using these three variables. At an optimal cutoff value of the regression formula, the risk score had an accuracy of 79.6% for predicting significant fibrosis, sensitivity of 89.5%, specificity of 78.4%, positive predictive value of 34.7%, negative predictive value of 98.3%, and ROC area of 0.855 (95% CI: 0.76-0.95). Fibroscan utility in predicting significant liver fibrosis in morbidly obese subjects is limited with accuracy of 71.3%. A model incorporating hemoglobin A1c and alkaline phosphatase with LSM improves accuracy in detecting significant fibrosis in this patient population.

Comparative analysis of BIA, IBC and DXA for determining body fat in American Football players.

BACKGROUND: Body composition is frequently measured by sports, fitness, and healthcare professionals. Dual-Energy X-ray Absorptiometry (DXA) analysis is a validated measurement of body composition and is considered a criterion or "gold-standard" measurement. However, due to long scan times, accessibility and cost, conducting DXA scans on larger athletes (i.e., football players) is difficult. Hence fitness professionals, notably strength and conditioning coaches, typically use other methods to measure body composition. The aim of this study was to assess the accuracy of the Bioelectrical Impedance Analysis (BIA) and Integrative Body Composition (IBC) techniques to DXA body fat percent (BF%) in collegiate American Football players. METHODS: Participants performed all three modes of body composition measurement: DXA, BIA (BIA-A [athlete]and BIA-NA [non-athlete modes]), and IBC, on the same day during early morning hours in a fasted state. RESULTS: The BF% measured via all methods significantly correlated with BF% measured via DXA (i.e., BIA-A [P<0.001, r=0.903], BIA-NA [P<0.001, r=0.891], and IBC [P<0.001, r=0.867]). However, values obtained via BIA-A (athlete) (P<0.001) and IBC (P<0.001) methods under predicted BF%. CONCLUSIONS: BIA and IBC can be used as an alternative to DXA for measuring BF% in American Football players. The BIA-A and IBC under predicted BF% compare to DXA, therefore, a correction formula can be utilized by coaches and athletes to predict BF% more accurately compared to IBC and BIA-A methods in American Football players.

The Tailgate Study: Differing metabolic effects of a bout of excessive eating and drinking.

INTRODUCTION: Excess energy intake by spectators at a sporting event (i.e., a tailgate) might cause acute negative health effects. However, limited data exist regarding the effects of overeating and alcohol consumption on lipid metabolism and the potential to gain intrahepatic triacylglycerols (IHTG). We tested the hypothesis that overconsumption of food and alcohol would significantly increase both hepatic de novo lipogenesis (DNL) and IHTG. METHODS: Eighteen males (mean ± SD, age: 31.4 ± 7.3 years, BMI: 32.1 ± 5.9 kg/m2) were given alcoholic drinks to elevate blood alcohol for 5 h, while highly palatable food was presented. Blood samples were collected and DNL in TG-rich lipoproteins (TRL) was measured by GC/MS, IHTG was measured via MRS (n = 15), and substrate oxidation was measured via indirect calorimetry. RESULTS: Subjects consumed 5087 ± 149 kcal (191 ± 25% excess of total daily energy needs including 171 ± 24 g alcohol), which increased plasma insulin, glucose, TG, and decreased NEFA (ANOVA p ≤ 0.003 for all). Both DNL and TRL-TG increased (p < 0.001), while IHTG did not change in the group as a whole (p = 0.229). Individual subject data revealed remarkably differing responses for IHTG (nine increased, five decreased, one did not change). Despite maintaining equal breath alcohol levels, subjects with IHTG elevations exhibited higher DNL, consumed 90% less alcohol (p = 0.048), tended to consume more carbohydrates, and exhibited lower whole-body fat oxidation (not significant) compared to those whose IHTG was reduced. DISCUSSION: This study demonstrates that acute excess energy intake may have differing effects on an individual's DNL and IHTG, and dietary carbohydrate may influence DNL more than alcohol.

Lactation Versus Formula Feeding: Insulin, Glucose, and Fatty Acid Metabolism During the Postpartum Period.

Milk production may involve a transient development of insulin resistance in nonmammary tissues to support redistribution of maternal macronutrients to match the requirements of the lactating mammary gland. In the current study, adipose and liver metabolic responses were measured in the fasting state and during a two-step (10 and 20 mU/m2/min) hyperinsulinemic-euglycemic clamp with stable isotopes, in 6-week postpartum women who were lactating (n = 12) or formula-feeding (n = 6) their infants and who were closely matched for baseline characteristics (e.g., parity, body composition, and intrahepatic lipid). When controlling for the low insulin concentrations of both groups, the lactating women exhibited a fasting rate of endogenous glucose production (EGP) that was 2.6-fold greater and a lipolysis rate that was 2.3-fold greater than the formula-feeding group. During the clamp, the groups exhibited similar suppression rates of EGP and lipolysis. In the lactating women only, higher prolactin concentrations were associated with greater suppression rates of lipolysis and lower intrahepatic lipid and plasma triacylglycerol concentrations. These data suggest that whole-body alterations in glucose transport may be organ specific and facilitate nutrient partitioning during lactation. Recapitulating a shift toward noninsulin-mediated glucose uptake could be an early postpartum strategy to enhance lactation success in women at risk for delayed onset of milk production.

Leptin decreases de novo lipogenesis in patients with lipodystrophy.

De novo lipogenesis (DNL) plays a role in the development of hepatic steatosis. In humans with lipodystrophy, reduced adipose tissue causes lower plasma leptin, insulin resistance, dyslipidemia, and ectopic triglyceride (TG) accumulation. We hypothesized that recombinant leptin (metreleptin) for 6 months in 11 patients with lipodystrophy would reduce DNL by decreasing insulin resistance and glycemia, thus reducing circulating TG and hepatic TG. The percentage of TG in TG-rich lipoprotein particle (TRLP-TG) derived from DNL (%DNL) was measured by deuterium incorporation from body water into palmitate. At baseline, DNL was elevated, similar to levels previously shown in obesity-associated nonalcoholic fatty liver disease (NAFLD). After metreleptin, DNL decreased into the normal range. Similarly, absolute DNL (TRLP-TG × %DNL) decreased by 88% to near-normal levels. Metreleptin improved peripheral insulin sensitivity (hyperinsulinemic-euglycemic clamp) and lowered hemoglobin A1c and hepatic TG. Both before and after metreleptin, DNL positively correlated with insulin resistance, insulin doses, and hepatic TG, supporting the hypothesis that hyperinsulinemia stimulates DNL and that elevated DNL is integral to the pathogenesis of lipodystrophy-associated NAFLD. These data suggest that leptin-mediated improvement in insulin sensitivity increases clearance of blood glucose by peripheral tissues, reduces hepatic carbohydrate flux, and lowers insulinemia, resulting in DNL reductions and improvements in hepatic steatosis and dyslipidemia.

Changes in Food Cravings and Eating Behavior after a Dietary Carbohydrate Restriction Intervention Trial.

Compared to low-fat diets, low-carbohydrate (CHO) diets cause weight loss (WL) over a faster time frame; however, it is unknown how changes in food cravings and eating behavior contribute to this more rapid WL in the early phases of dieting. We hypothesized that reductions in food cravings and improved eating behaviors would be evident even after a relatively short (4-week) duration of CHO-restriction, and that these changes would be associated with WL. Adult participants (n = 19, 53% males, mean ± SD: BMI = 34.1 ± 0.8 kg/m2; age 40.6 ± 1.9 years) consumed a CHO-restricted diet (14% CHO, 58% fat, 28% protein) for 4 weeks. Before and after the intervention, specific and total cravings were measured with the Food Craving Inventory (FCI) and eating behaviors assessed with the Three-Factor Eating questionnaire. Food cravings were significantly reduced at week 4, while women had significantly greater reductions in sweet cravings than men. Dietary restraint was significantly increased by 102%, while disinhibiton and hunger scores were reduced (17% and 22%, respectively, p < 0.05). Changes in cravings were unrelated to changes in body weight except for the change in high-fat cravings where those who lost the most weight experienced the least reductions in fat cravings (r = -0.458, p = 0.049). Changes in dietary restraint were inversely related to several FCI subscales. A short-term, low-CHO diet was effective in reducing food cravings. These data suggest that in subjects that have successfully lost weight on a low-CHO diet, those who craved high-fat foods at the onset were able to satisfy their cravings-potentially due to the high-fat nature of this restricted diet.

Effect of carbohydrate restriction-induced weight loss on aortic pulse wave velocity in overweight men and women.

Increased aortic stiffness, measured by carotid-to-femoral pulse wave velocity (PWV), is an independent predictor of cardiovascular disease, and past data have shown that low-fat and low-energy diets, fed for 8-24 weeks, lower PWV. The purpose of this study was to determine whether a reduction in PWV would be achieved by dietary carbohydrate (CHO) restriction, shown to bring about weight loss over a shorter timeframe. Men (n = 10, age: 41.8 ± 10.2 years, BMI: 34.2 ± 3.0 kg/m2 (mean ± SD)) and women (n = 10, age: 38.6 ± 6.1 years, BMI: 33.5 ± 3.8 kg/m2) with characteristics of insulin resistance and the metabolic syndrome consumed a structured, CHO-restricted diet for 4 weeks (energy deficit, 645 kcal/day). For the whole group, subjects lost 5.4% ± 0.5% (P < 0.001) of body weight and experienced significant reductions in blood pressure (6%-8%), plasma insulin (34%), and triglycerides (34%). PWV was reduced by 6% ± 2% (7.1 ± 0.2 m/s to 6.7 ± 0.2 m/s, P = 0.008) and surprisingly, in women, it fell significantly (from 7.2 ± 0.3 m/s to 6.3 ± 0.3 m/s, P = 0.028), while no changes were observed in men (7.2 ± 0.3 vs. 7.0 ± 0.3 m/s, P = 0.144). This is the first study to demonstrate that weight loss can improve PWV in as little as 4 weeks and that dietary CHO restriction may be an effective treatment for reducing aortic stiffness in women. Future studies are needed to establish the mechanisms by which dietary CHO restriction may confer more cardiovascular benefits to women than to men.