Dapagliflozin Enhances Arterial and Venous Compliance During Exercise in Heart Failure With Preserved Ejection Fraction: Insights From the Cardiac and Metabolic Effects of Dapagliflozin in Heart Failure With Preserved Ejection Fraction Trial.

BACKGROUND: Systemic arterial compliance and venous capacitance are typically impaired in patients with heart failure with preserved ejection fraction (HFpEF), contributing to hemodynamic congestion with stress. Sodium-glucose cotransporter-2 inhibitors reduce hemodynamic congestion and improve clinical outcomes in patients with HFpEF, but the mechanisms remain unclear. This study tested the hypothesis that Dapagliflozin would improve systemic arterial compliance and venous capacitance during exercise in patients with HFpEF. METHODS: In this secondary analysis from the Cardiac and Metabolic Effects of Dapagliflozin in Heart Failure With Preserved Ejection Fraction Trial, 37 patients with HFpEF (mean age 68 ± 9 years, women 65%) underwent invasive hemodynamic exercise testing with simultaneous echocardiography at baseline and following treatment for 24 weeks with Dapagliflozin or placebo. Radial artery pressure (BP) was measured continuously using a fluid-filled catheter with transformation to aortic pressure, central hemodynamics were measured using high-fidelity micromanometers, and stressed blood volume was estimated from hemodynamic indices fit to a comprehensive cardiovascular model. RESULTS: There was no statistically significant effect of Dapagliflozin on resting BP, but Dapagliflozin reduced systolic BP during peak exercise (estimated treatment difference [ETD], -18.8 mm Hg [95% CI, -33.9 to -3.7] P=0.016). Reduction in BP was related to improved exertional total arterial compliance (ETD, 0.06 mL/mm Hg/m2 [95% CI, 0.003-0.11] P=0.039) and aortic root characteristic impedance (ETD, -2.6 mm Hg/mL*sec [95% CI: -5.1 to -0.03] P=0.048), with no significant effect on systemic vascular resistance. Dapagliflozin reduced estimated stressed blood volume at rest and during peak exercise (ETD, -292 mm Hg [95% CI, -530 to -53] P=0.018), and improved venous capacitance evidenced by a decline in ratio of estimated stressed blood volume to total blood volume (ETD, -7.3% [95% CI, -13.3 to -1.3] P=0.020). Each of these effects of Dapagliflozin at peak exercise were also observed during matched 20W exercise intensity. Improvements in total arterial compliance and estimated stressed blood volume were correlated with decreases in body weight, and reduction in systolic BP with treatment was correlated with the change in estimated stressed blood volume during exercise (r=0.40, P=0.019). Decreases in BP were correlated with reduction in pulmonary capillary wedge pressure during exercise (r=0.56, P<0.001). CONCLUSIONS: In patients with HFpEF, treatment with Dapagliflozin improved systemic arterial compliance and venous capacitance during exercise, while reducing aortic characteristic impedance, suggesting a reduction in arterial wall stiffness. These vascular effects may partially explain the clinical benefits with sodium-glucose cotransporter-2 inhibitors in HFpEF. REGISTRATION: URL: https://www.clinicaltrials.gov; Unique identifier: NCT04730947.

Similar insulin regulation of splanchnic FFA and VLDL-TG in men with nonalcoholic hepatic steatosis and steatohepatitis.

This study aimed to determine whether obese men with nonalcoholic fatty liver disease (NAFLD) display differences between those with simple steatosis versus steatohepatitis (NASH) in splanchnic and hepatic FFA and VLDL-triglycerides (VLDL-TG) balances. The study involved 17 obese men with biopsy-proven NAFLD (9 with NASH and 8 with simple steatosis). We used hepatic vein catheterization in combination with [3H]palmitate and [14C]VLDL-TG tracers to measure splanchnic palmitate and VLDL-TG uptake and release rates during basal and hyperinsulinemic conditions. Indocyanine green was used to measure splanchnic plasma flow. Splanchnic palmitate uptake was similar in the two groups and significantly reduced during hyperinsulinemia (NASH: 62 (48-77) versus 38 (18-58) µmol/min; simple steatosis: 62 (46-78) versus 45 (25-65) µmol/min, mean (95% CI), basal versus clamp periods, respectively, P = 0.02 time-effect). Splanchnic palmitate release was also comparable between groups and nonsignificantly diminished during hyperinsulinemia. The percent palmitate delivered to the liver originating from visceral adipose tissue lipolysis was similar and unchanged by hyperinsulinemia. Splanchnic uptake and release of VLDL-TG were similar between groups. Hyperinsulinemia suppressed VLDL-TG release (P <0.05 time-effect) in both groups. Insulin-mediated glucose disposal was similar in the two groups (P = 0.54). Obese men with NASH and simple steatosis have similar splanchnic uptake and release of FFA and VLDL-TG and a similar proportion of FFA from visceral adipose tissue lipolysis delivered to the liver. These results demonstrate that the splanchnic balances of FFA and VLDL-TG do not differ between obese men with NASH and those with simple steatosis.

Cardiac and Metabolic Effects of Dapagliflozin in Heart Failure With Preserved Ejection Fraction: The CAMEO-DAPA Trial.

BACKGROUND: Sodium-glucose cotransporter-2 inhibitors reduce risk of hospitalization for heart failure in patients who have heart failure with preserved ejection fraction (HFpEF), but the hemodynamic mechanisms underlying these benefits remain unclear. This study sought to determine whether treatment with dapagliflozin affects pulmonary capillary wedge pressure (PCWP) at rest and during exercise in patients with HFpEF. METHODS: This was a single-center, double-blinded, randomized, placebo-controlled trial testing the effects of 10 mg of dapagliflozin once daily in patients with HFpEF. Patients with New York Heart Association class II or III heart failure, ejection fraction ≥50%, and elevated PCWP during exercise were recruited. Cardiac hemodynamics were measured at rest and during exercise using high-fidelity micromanometers at baseline and after 24 weeks of treatment. The primary end point was a change from baseline in rest and peak exercise PCWPs that incorporated both measurements, and was compared using a mixed-model likelihood ratio test. Key secondary end points included body weight and directly measured blood and plasma volumes. Expired gas analysis was performed evaluate oxygen transport in tandem with arterial lactate sampling. RESULTS: Among 38 patients completing baseline assessments (median age 68 years; 66% women; 71% obese), 37 completed the trial. Treatment with dapagliflozin resulted in reduction in the primary end point of change in PCWP at rest and during exercise at 24 weeks relative to treatment with placebo (likelihood ratio test for overall changes in PCWP; P<0.001), with lower PCWP at rest (estimated treatment difference [ETD], -3.5 mm Hg [95% CI, -6.6 to -0.4]; P=0.029) and maximal exercise (ETD, -5.7 mm Hg [95% CI, -10.8 to -0.7]; P=0.027). Body weight was reduced with dapagliflozin (ETD, -3.5 kg [95% CI, -5.9 to -1.1]; P=0.006), as was plasma volume (ETD, -285 mL [95% CI, -510 to -60]; P=0.014), but there was no significant effect on red blood cell volume. There were no differences in oxygen consumption at 20-W or peak exercise, but dapagliflozin decreased arterial lactate at 20 W (-0.70 ± 0.77 versus 0.37 ± 1.29 mM; P=0.006). CONCLUSIONS: In patients with HFpEF, treatment with dapagliflozin reduces resting and exercise PCWP, along with the favorable effects on plasma volume and body weight. These findings provide new insight into the hemodynamic mechanisms of benefit with sodium-glucose cotransporter-2 inhibitors in HFpEF. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT04730947.

Effects of acute changes in fasting glucose and free fatty acid concentrations on indices of ß-cell function and glucose metabolism in subjects without diabetes.

Elevated fasting free fatty acids (FFAs) and fasting glucose are additively associated with impaired glucose tolerance (IGT) and decreased ß-cell function [quantified as disposition index (DI)]. We sought to examine how changes in fasting FFA and glucose alter islet function. We studied 10 subjects with normal fasting glucose (NFG) and normal glucose tolerance (NGT) on two occasions. On one occasion, Intralipid and glucose were infused overnight to mimic conditions present in IFG/IGT. In addition, we studied seven subjects with IFG/IGT on two occasions. On one occasion, insulin was infused to lower overnight FFA and glucose concentrations to those observed in people with NFG/NGT. The following morning, a labeled mixed meal was used to measure postprandial glucose metabolism and ß-cell function. Elevation of overnight fasting FFA and glucose in NFG/NGT did not alter peak or integrated glucose concentrations (2.0 ± 0.1 vs. 2.0 ± 0.1 Mol per 5 h, Saline vs. Intralipid/glucose, P = 0.55). Although overall ß-cell function quantified by the Disposition Index was unchanged, the dynamic component of ß-cell responsivity (ϕd) was decreased by Intralipid and glucose infusion (9 ± 1 vs. 16 ± 3 10-9, P = 0.02). In people with IFG/IGT, insulin did not alter postprandial glucose concentrations or indices of ß-cell function. Endogenous glucose production and glucose disappearance were also unchanged in both groups. We conclude that acute, overnight changes in FFA, and glucose concentrations do not alter islet function or glucose metabolism in prediabetes.NEW & NOTEWORTHY This experiment studied the effect of changes in overnight concentrations of free fatty acids (FFAs) and glucose on ß-cell function and glucose metabolism. In response to elevation of these metabolites, the dynamic component of the ß-cell response to glucose was impaired. This suggests that in health overnight hyperglycemia and FFA elevation can deplete preformed insulin granules in the ß-cell.

Endothelin-1 as a novel target for the prevention of metabolic dysfunction with intermittent hypoxia in male participants.

We examined the effect of intermittent hypoxia (IH, a hallmark feature of sleep apnea) on adipose tissue lipolysis and the role of endothelin-1 (ET-1) in this response. We hypothesized that IH can increase ET-1 secretion and plasma free fatty acid (FFA) concentrations. We further hypothesized that inhibition of ET-1 receptor activation with bosentan could prevent any IH-mediated increase in FFA. To test this hypothesis, 16 healthy male participants (32 ± 5 yr, 26 ± 2 kg/m2) were exposed to 30 min of IH in the absence (control) and presence of bosentan (62.5 mg oral twice daily for 3 days prior). Arterial blood samples for ET-1, epinephrine, and FFA concentrations, as well as abdominal subcutaneous adipose tissue biopsies (to assess transcription of cellular receptors/proteins involved in lipolysis), were collected. Additional proof-of-concept studies were conducted in vitro using primary differentiated human white preadipocytes (HWPs). We show that IH increased circulating ET-1, epinephrine, and FFA (P < 0.05). Bosentan treatment reduced plasma epinephrine concentrations and blunted IH-mediated increases in FFA (P < 0.01). In adipose tissue, bosentan had no effect on cellular receptors and proteins involved in lipolysis (P > 0.05). ET-1 treatment did not directly induce lipolysis in differentiated HWP. In conclusion, IH increases plasma ET-1 and FFA concentrations. Inhibition of ET-1 receptors with bosentan attenuates the FFA increase in response to IH. Based on a lack of a direct effect of ET-1 in HWP, we speculate the effect of bosentan on circulating FFA in vivo may be secondary to its ability to reduce sympathoadrenal tone.

Pathophysiologic importance of visceral adipose tissue in women with heart failure and preserved ejection fraction.

AIMS: Central obesity is a major risk factor for heart failure with preserved ejection fraction (HFpEF), particularly in women, but the mechanisms remain unclear. We hypothesized that sex-specific differences in visceral adipose tissue (VAT) content would differentially relate to haemodynamic severity of HFpEF in women and men. METHODS AND RESULTS: Abdominal computed tomography (CT) and invasive haemodynamic exercise testing were performed in 105 subjects with HFpEF (63 women) and 105 age-, sex-, and body mass index-matched controls. Visceral adipose tissue area was quantified by CT. As compared with control women, VAT area was 34% higher in women with HFpEF (186 ± 112 vs. 139 ± 72 cm2, P = 0.006), while VAT area was not significantly different in men with or without HFpEF (294 ± 158 vs. 252 ± 92 cm2, P = 0.1). During exercise, pulmonary capillary wedge pressure (PCWP) increased markedly and to similar extent in both men and women with HFpEF. Women with increased VAT area displayed 33% higher PCWP during exercise compared with women with normal VAT area (28 ± 10 vs. 21 ± 10 mmHg, P = 0.001), whereas exercise PCWP was similar in men with or without excess VAT (24 ± 9 vs. 25 ± 6, P = 0.89). In women, each 100 cm2 increase in VAT area was associated with a 4.0 mmHg higher PCWP (95% CI 2.1, 6.0 mmHg; P < 0.0001), but there was no such relationship in men (interaction P = 0.009). CONCLUSIONS: These data suggest that accumulation of excess VAT plays a distinct and important role in the pathophysiology of HFpEF preferentially in women. Further research is needed to better understand the mechanisms and treatment implications for visceral fat in HFpEF.

Red blood cell triglycerides-a unique pool that incorporates plasma-free fatty acids and relates to metabolic health.

Most research into red blood cell (RBC) lipids focuses on membrane phospholipids and their relationships to metabolic conditions and diet. Triglycerides (TGs) exist in most cells; the TG-fatty acids serve as readily available fuel for oxidative phosphorylation. Because RBCs lack mitochondria, they would not be expected to store fatty acids in TG. We followed up on a previous in vitro study that found FFA can be incorporated into RBC-TG by testing whether intravenously infused [U-13C]palmitate could be detected in RBC-TG. We also quantified RBC-TG fatty acid concentrations and profiles as they relate to plasma FFA and lipid concentrations. We found that 1) RBC-TG concentrations measured by glycerol and LC/MS were correlated (r = 0.77; P < 0.001) and averaged <50 nmol/ml RBC; 2) RBC-TG concentrations were stable over 18 h; 3) [U-13C]palmitate was detectable in RBC-TG from half the participants; 4) RBC-TGs were enriched in saturated fatty acids and depleted in unsaturated fatty acid compared with plasma FFA and previously reported RBC membrane phospholipids; 5) RBC-TG fatty acid profiles differed significantly between obese and nonobese adults; 6) weight loss altered the RBC-TG fatty acid profile in the obese group; and 7) the RBC-TG fatty acid composition correlated with plasma lipid concentrations. This is the first report showing that plasma FFA contributes to RBC-TG in vivo, in humans, and that the RBC-TG fatty acid profile is related to metabolic health. The storage of saturated fatty acids in RBC-TG stands in stark contrast to the highly unsaturated profile reported in RBC membrane phospholipids.

Early-life exposure to the Chinese famine, genetic susceptibility and the risk of type 2 diabetes in adulthood.

AIMS/HYPOTHESIS: Early famine exposure has been related to the development of type 2 diabetes; however, little is known about whether the genetic background modifies this association. We aimed to investigate the joint effects of famine exposure at different stages of early life and genetic susceptibility on diabetes risk in adulthood. METHODS: The study included 8350 participants from the Survey on Prevalence in East China for Metabolic Diseases and Risk Factors (SPECT-China) who were born around the time of the Chinese Great Famine. We determined famine exposure subgroups according to the birth year as nonexposed (1963-1974), fetal-exposed (1959-1962), childhood-exposed (1949-1958), and adolescence-exposed (1941-1948). We developed a genetic risk score of 21 variants previously associated with type 2 diabetes in East Asians. Hierarchical logistic models were used to examine the association of famine exposure and genetic risk with diabetes. RESULTS: The age-standardised prevalence of diabetes in nonexposed, fetal-exposed, childhood-exposed and adolescence-exposed subgroups was 13.0%, 18.2%, 15.1% and 13.2%, respectively. Compared with nonexposed participants, fetal-exposed participants showed an increased risk of diabetes in adulthood (OR 1.47; 95% CI 1.13, 1.93). A higher genetic risk score was associated with an increased risk of diabetes (OR 1.23; 95% CI 1.15, 1.31 per SD increment). The association between famine exposure and diabetes was consistent across genetic risk strata (all p for interaction >0.05). When considered jointly, fetal- or childhood-exposed participants at high genetic risk (highest tertile of genetic risk score) had 2.60-fold (95% CI 1.71, 3.93) and 1.95-fold (95% CI 1.24, 3.05) higher risks of diabetes, respectively, compared with nonexposed participants at low genetic risk (lowest tertile). CONCLUSIONS/INTERPRETATIONS: Prenatal exposure to famine was associated with an increased risk of type 2 diabetes in Chinese adults independent of genetic risk score using 21 variants common in the East Asian population. Famine exposure and genetic susceptibility may exhibit an additive effect on diabetes development.

Regulation of direct adipose tissue free fatty acid storage during mixed meal ingestion and high free fatty acid concentration conditions.

We found that direct free fatty acid (FFA) storage (fatty acid cycling back into adipose tissue) in leg vs. abdominal subcutaneous fat is related to regional differences in adipose tissue diacylglycerol acyltransferase (DGAT) activity under high-FFA conditions and to differences in adipose tissue acyl-CoA synthetase (ACS)activity under meal ingestion conditions. We also found that direct FFA storage rates in leg fat were significantly less in physically active than sedentary adults. Direct FFA storage into adipocytes relates to body fat distribution. Adipose tissue CD36, ACS, and DGAT may account for some of the between-depot and interindividual variability in FFA storage. These studies were to test whether CD36, ACS, or DGAT might be important for direct palmitate storage under meal ingestion or high-FFA conditions. We measured upper (UBSQ) and lower body subcutaneous (LBSQ) adipose tissue FFA storage rates by infusing palmitate tracers intravenously and performing adipose biopsies under hypoinsulinemic (high-FFA) and mixed-meal conditions. We recruited five postmenopausal women, physically active males (5) and females (5), and sedentary males (5) and females (5). We found that 1) the ratio of UBSQ to LBSQ DGAT activity predicted the ratio of palmitate storage [adjusted R = 0.25, F = 8.0, P = 0.01, 95% CI (0.07, 0.48)] under high-FFA conditions; 2) the ratio of UBSQ to LBSQ ACS activity predicted the ratio of palmitate storage under meal conditions [adjusted R = 0.18, F = 6.3, P = 0.02, 95% CI (0.12, 1.28)]; 3) LBSQ direct palmitate storage rates were significantly less in physically active than sedentary and 4) adipose tissue CD36 protein content, ACS, or DGAT activities did not independently predict palmitate storage rates. We conclude that physically active adults have lesser fatty acid cycling back into adipose tissue and that adipose ACS and DGAT may affect competition between UBSQ and LBSQ adipose for direct palmitate storage.

Adipose tissue macrophage populations and inflammation are associated with systemic inflammation and insulin resistance in obesity.

Obesity is accompanied by numerous systemic and tissue-specific derangements, including systemic inflammation, insulin resistance, and mitochondrial abnormalities in skeletal muscle. Despite growing recognition that adipose tissue dysfunction plays a role in obesity-related disorders, the relationship between adipose tissue inflammation and other pathological features of obesity is not well-understood. We assessed macrophage populations and measured the expression of inflammatory cytokines in abdominal adipose tissue biopsies in 39 nondiabetic adults across a range of body mass indexes (BMI 20.5-45.8 kg/m2). Skeletal muscle biopsies were used to evaluate mitochondrial respiratory capacity, ATP production capacity, coupling, and reactive oxygen species production. Insulin sensitivity (SI) and ß cell responsivity were determined from test meal postprandial glucose, insulin, c-peptide, and triglyceride kinetics. We examined the relationships between adipose tissue inflammatory markers, systemic inflammatory markers, SI, and skeletal muscle mitochondrial physiology. BMI was associated with increased adipose tissue and systemic inflammation, reduced SI, and reduced skeletal muscle mitochondrial oxidative capacity. Adipose-resident macrophage numbers were positively associated with circulating inflammatory markers, including tumor necrosis factor-α (TNFα) and C-reactive protein (CRP). Local adipose tissue inflammation and circulating concentrations of TNFα and CRP were negatively associated with SI, and circulating concentrations of TNFα and CRP were also negatively associated with skeletal muscle oxidative capacity. These results demonstrate that obese humans exhibit increased adipose tissue inflammation concurrently with increased systemic inflammation, reduced insulin sensitivity, and reduced muscle oxidative capacity and suggest that adipose tissue and systemic inflammation may drive obesity-associated metabolic derangements.NEW AND NOTEWORTHY Adipose inflammation is proposed to be at the nexus of the systemic inflammation and metabolic derangements associated with obesity. The present study provides evidence to support adipose inflammation as a central feature of the pathophysiology of obesity. Adipose inflammation is associated with systemic and peripheral metabolic derangements, including increased systemic inflammation, reduced insulin sensitivity, and reduced skeletal muscle mitochondrial respiration.

Adipose tissue macrophage burden, systemic inflammation, and insulin resistance.

Adipose tissue inflammation, as defined by macrophage accumulation, is proposed to cause insulin resistance and systemic inflammation. Because the strength of this relationship for humans is unclear, we tested whether adipose tissue macrophage (ATM) burden is correlated with these health indicators. Using immunohistochemistry, we measured abdominal subcutaneous CD68+ (total ATM), CD14+ (proinflammatory/M1), and CD206+ (anti-inflammatory/M2) ATM in 97 volunteers (BMI 20-38 kg/m2, in addition to body composition, adipocyte size, homeostasis model assessment of insulin resistance, ADIPO-IR, adipose tissue insulin resistance measured by palmitate, plasma lipids, TNF, and IL-6 concentrations. There were several significant univariate correlations between metabolic parameters to IL-6 and ATM per 100 adipocytes, but not ATM per gram tissue; adipocyte size was a confounding variable. We used matching strategies and multivariate regression analyses to investigate the relationships between ATM and inflammatory/metabolic parameters independent of adipocyte size. Matching approaches revealed that the groups discordant for CD206 but concordant for adipocyte size had significantly different fasting insulin and IL-6 concentrations. However, groups discordant for adipocyte size but concordent for ATM differeded in that visceral fat, plasma triglyceride, glucose, and TNF concentrations were greater in those with large adipocytes. Multivariate regression analysis indicated that indexes of insulin resistance and fasting triglycerides were predicted by body composition; the predictive value of ATM per 100 adipocytes or per gram tissue was variable between males and females. We conclude that the relationship between ATM burden and metabolic/inflammatory variables is confounded by adipocyte size/body composition and that ATM do not predict insulin resistance, systemic inflammation, or dyslipidemia. ATM may primarily play a role in tissue remodeling rather than in metabolic pathology.

Low-level laser therapy for weight reduction: a randomized pilot study.

Obesity and overweight is a global health crisis and novel methods of treatment are needed to address it. Low-level laser therapy (LLLT) is a currently available non-invasive procedure for lysing excess fat, but there is a lack of consensus exists on LLLT frequency and limited research from studies of LLLT. The purpose of this pilot study is to compare the effect of three of the most common LLLT frequencies on weight, waist circumference, body fat percentage, and quality of life. Sixty overweight (body mass index (BMI) 25-29.9 kg/m2) adult participants were randomized to 12 LLLT treatments: (1) three times weekly for 4 weeks, (2) twice weekly for 6 weeks, or (3) once weekly for 12 weeks. All participants attended an in-person visit at baseline and at weeks 4, 6, 12, and 26. Participants were recruited September 30, 2016 through to August 27, 2017. The majority of the 60 participants were female (90%) with an average age of 43.7 years (± 9.2 years). Most participants (98%) completed 10 or more of the 12 LLLT treatments. When comparing across treatment groups, the greatest reductions from baseline were observed in those assigned to twice weekly for 6 weeks in weight (1 ± 1.7 (±SD) kg by week 6), waist circumference (- 2.0 ± 3.2 in. by week 6 and - 1.5 ± 3.2 in. by week 26), body mass index (- 0.4 ± 0.6 kg/m2), and body fat mass (- 1.1 ± 1.6 kg). This group also had the most significant improvement from baseline in quality of life (+ 0.5 ± 0.8 by week 6), body satisfaction (+ 0.2 ± 0.4 by week 6 and week 26), and body appreciation (+ 0.2 ± 0.3 by week 6 and + 0.3 ± 0.3 by week 26). LLLT twice weekly for 6 weeks could be proposed as the optimal frequency and duration for the management of body weight. Trial registration: https://clinicaltrials.gov/ct2/show/NCT02877004. Registered August 24, 2016.

Subcutaneous adipose tissue free fatty acid uptake measured using positron emission tomography and adipose biopsies in humans.

Positron emission tomography (PET) radiopharmaceuticals can noninvasively measure free fatty acid (FFA) uptake into adipose tissue. We studied 29 volunteers to test whether abdominal and femoral subcutaneous adipose tissue FFA uptake measured using [1-11C]palmitate PET agrees with FFA storage rates measured using an intravenous bolus of [1-14C]palmitate and adipose biopsies. The dynamic left ventricular cavity PET images combined with blood sample radioactivity corrected for the 11CO2 content were used to create the blood time activity curve (TAC), and the constant (Ki) was determined using Patlak analysis of the TACs generated for regions of interest in abdominal subcutaneous fat. These data were used to calculate palmitate uptake rates in abdominal subcutaneous adipose tissue (µmol·kg-1·min-1). Immediately after the dynamic imaging, a static image of the thigh was taken to measure the standardized uptake value (SUV) in thigh adipose tissue, which was scaled to each participant's abdominal adipose tissue SUV to calculate thigh adipose palmitate uptake rates. Abdominal adipose palmitate uptake using PET [1-11C]palmitate was correlated with, but significantly (P < 0.001) greater than, FFA storage measured using [1-14C]palmitate and adipose biopsy. Thigh adipose palmitate measured using PET calculation was positively correlated (R2 = 0.44, P < 0.0001) with and not different from the biopsy approach. The relative differences between PET measured abdominal subcutaneous adipose tissue palmitate uptake and biopsy-measured palmitate storage were positively correlated (P = 0.03) with abdominal subcutaneous fat. We conclude that abdominal adipose tissue FFA uptake measured using PET does not equate to adipose FFA storage measured using biopsy techniques.

Preliminary evidence for reduced adipose tissue inflammation in vegetarians compared with omnivores.

BACKGROUND: There are links between obesity and inflammation that may relate activation of pro-inflammatory pathways by dietary factors. Because dietary fat intake of vegetarians is thought to be more beneficial than that of omnivores, we hypothesized that obese vegetarians would have less adipose tissue inflammation and lower intramyocellular ceramide concentrations than equally obese omnivores. METHODS: Eight obese vegetarian (1 male) and 8 obese omnivore volunteers (1 male) completed a Food Frequency Questionnaire, underwent body composition measures, subcutaneous adipose tissue and muscle biopsies. We used immunohistochemistry to measure adipose macrophage (ATM) and senescent cells. Plasma free fatty acid (FFA), adipose FA and muscle ceramide profiles were measured using liquid chromatography/mass spectrometry. Student t tests were used for the comparison of primary outcomes; univariate regression analysis was used to test for associations between dietary patterns and ATMs (secondary analysis). RESULTS: There were no differences in age (38 ± 8 vs. 39 ± 8 years), BMI (32.2 ± 2.6 vs. 33.3 ± 1.9 kg/m2) or percent body fat (44 ± 8 vs. 45 ± 4) between the vegetarians and omnivores. Vegetarians consumed 42% (P = 0.02) less saturated fat and 50% (P = 0.04) less cholesterol than the omnivores. Plasma FFA of vegetarians had lesser proportions of palmitic acid (24 ± 3 vs. 29 ± 4%, P = 0.02) and vegetarians had fewer femoral pro-inflammatory ATMs than omnivores (3.6 ± 2.8 vs. 7.9 ± 4.4 per 100 adipocytes, respectively; P = 0.02). Omnivores had 50% greater (P = 0.01) expression of TNF mRNA in abdominal fat. We found no significant between group differences in muscle ceramide concentrations. CONCLUSIONS: Although the sample size is small, these results may indicate that dietary patterns play a role in adipose tissue inflammation, as reflected by reduced number of femoral ATMs in obese vegetarians than obese omnivores.

Free fatty acid flux measured using [1-11C]palmitate positron emission tomography and [U-13C]palmitate in humans.

PET radiopharmaceuticals can noninvasively measure free fatty acid (FFA) tissue uptake. Investigators often use PET scan-derived data to calculate FFA flux. We tested whether the [1-11C]palmitate PET measures of palmitate flux provide results equivalent to a continuous infusion of [U-13C]palmitate. Nine volunteers participated in study 1 to evaluate whether a rapidly (10-20 s) given bolus of [1-11C]palmitate affects calculated flux results. Thirty volunteers participated in study 2, which was identical to study 1 except that the [1-11C]palmitate bolus was given over 1 min. Volunteers in both studies also received a continuous intravenous infusion of [U-13C]palmitate. Plasma palmitate concentrations and enrichment were measured by liquid chromatography-mass spectrometry. The PET/CT images were analyzed on a workstation running PMOD. Palmitate flux was estimated using PET time-activity curve (TAC) data from regions of interest in the left ventricle (LV) and aorta both with and without hybrid TACs that employed the 11CO2-corrected data for the first 5 min and the 11CO2-corrected blood radioactivity for the remainder of the PET scan. Palmitate flux in study 1 measured with PET [1-11C]palmitate and [U-13C]palmitate were not correlated, and the PET [1-11C]palmitate flux was significantly less than the [U-13C]palmitate measured flux. In study 2, the palmitate flux using PET [1-11C]palmitate hybrid LV models provided closer mean estimates of [U-13C]palmitate measured flux. The best PET calculation approaches predicted 64% of the interindividual variance in [U-13C]palmitate measured flux. Palmitate kinetics measured using [1-11C]palmitate/PET do not provide the same palmitate kinetic results as the continuous infusion [U-13C]palmitate approach.

Effects of meal ingestion on intramyocellular ceramide concentrations and fractional de novo synthesis in humans.

We investigated the effects of meal ingestion on intramyofibrillar (IMF) and subsarcolemmal (SS) ceramide metabolism in volunteers ranging from lean to obese. Thirty-eight women and men underwent a steady-state meal ingestion protocol that included a 6.5-h infusion of [U-13C]palmitate and muscle biopsies 1.5 and 6.5 h after starting the tracer infusion. We measured IMF and SS sphingolipid concentrations and the contribution of plasma palmitate to intramyocellular C16:0 ceramide by use of LC-MS-MS. In response to meal ingestion SS C24 ceramide concentrations, but not C14-C20 concentrations, increased significantly. IMF ceramide concentrations did not change. The increases in SS C24 ceramides were negatively related to parameters of insulin resistance. The fractional contribution of plasma palmitate to intramyocellular C16:0 ceramides in both IMF and SS fractions was inversely related to overweight status (ß = -0.432, P = 0.0095 and ß = -0.443, P = 0.0058, respectively). These data indicate that meal ingestion has differing effects on SS ceramide subspecies and suggest that the fractional de novo synthesis of intramyocellular ceramide from plasma palmitate in the postprandial condition is reduced in those who are overweight.

Histomorphometric analyses of human adipose tissues using intact, flash-frozen samples.

Histomorphometric analyses of adipose tissue usually require formalin fixation of fresh samples. Our objective was to determine if intact, flash-frozen whole adipose tissue samples stored at - 80 °C could be used for measurements developed for fresh-fixed adipose tissues. Portions of adipose tissue samples were either formalin-fixed immediately upon sampling or flash-frozen and stored at - 80 °C and then formalin-fixed during the thawing process. Mean adipocyte diameter was measured. Immunohistochemistry was performed on additional samples to identify macrophage subtypes (M1, CD14 + and M2, CD206 +) and total (CD68 +) number. All slides were counterstained using haematoxylin and eosin (H&E). Visual inspection of H&E-stained adipose tissue slides performed in a blinded fashion showed little or no sign of cell breakage in 74% of frozen-fixed samples and in 68% of fresh-fixed samples (p > 0.5). There was no difference in the distribution frequencies of adipocyte sizes in fresh-fixed vs. frozen-fixed tissues in both depots (p > 0.9). Mean adipocyte size from frozen-fixed samples correlated significantly and positively with adipocyte size from fresh-fixed samples (r = 0.74, p < 0.0001, for both depots). The quality of staining/immunostaining and appearance of tissue architecture were comparable in fresh-fixed vs. frozen-fixed samples. In conclusion, intact flash-frozen adipose tissue samples stored at - 80 °C can be used to perform techniques conventionally applied to fresh-fixed samples. This approach allows for retrospective studies with frozen human adipose tissue samples.

The Soluble Fiber α-Cyclodextrin Does Not Increase the Fecal Losses of Dietary Fat in Adults-A Double-Blind, Randomized, Placebo-Controlled, Crossover Trial.

Background: α-Cyclodextrin (α-CD), a soluble dietary fiber, may improve abnormal plasma lipids and promote weight loss. Preliminary evidence suggests that it may exert these effects by binding dietary fat and reducing absorption; this has not been tested in humans. Objective: The primary objective was to test whether supplemental α-CD increases fecal content of dietary lipid in humans. Methods: This was a randomized, double-blind, placebo-controlled, crossover study completed at the Mayo Clinic. Eight healthy volunteers, 5 premenopausal women and 3 men ages 23-54 y with body mass index (BMI; kg/m2) 18-27, underwent 2 separate study visits with a ≥2-wk washout period. The first morning of each visit volunteers consumed a standardized breakfast (14.5% protein, 27.5% fat, 60% carbohydrate, and 1.5 kcal/mL) containing [14C]tripalmitin and [3H]triolein with 2 g of α-CD or placebo, followed by 2 g of α-CD or placebo per meal for 2 more days. Volunteers consumed 100 g/d of dietary fat. Feces were collected for 72 h after the labeled breakfast to measure radiotracer content and total fecal fat. Radiotracer appearance in plasma TGs was measured at intervals after the labeled meal as a secondary outcome. Results: Virtually no 3H radiotracer, but an average of ∼20% of the 14C radiotracer was recovered in fecal lipids, with no difference between α-CD and placebo. Total fecal fat content and radiotracer appearance in postprandial plasma TGs did not differ between the α-CD and placebo treatments. Plasma appearance of 14C-TG was 37% ± 14% less (P < 0.0001) than 3H-TG. Conclusions: α-CD supplementation did not increase loss of dietary lipid in stool or total fecal fat compared with placebo in healthy adults. Greater stool loss and lesser appearance in plasma TGs of tripalmitin-derived [14C] compared with triolein-derived [3H] TGs imply different metabolic handling of these 2 dietary fat tracers. This trial was registered at www.clinicaltrials.gov as NCT03002168.

JAK inhibition alleviates the cellular senescence-associated secretory phenotype and frailty in old age.

Chronic, low grade, sterile inflammation frequently accompanies aging and age-related diseases. Cellular senescence is associated with the production of proinflammatory chemokines, cytokines, and extracellular matrix (ECM) remodeling proteases, which comprise the senescence-associated secretory phenotype (SASP). We found a higher burden of senescent cells in adipose tissue with aging. Senescent human primary preadipocytes as well as human umbilical vein endothelial cells (HUVECs) developed a SASP that could be suppressed by targeting the JAK pathway using RNAi or JAK inhibitors. Conditioned medium (CM) from senescent human preadipocytes induced macrophage migration in vitro and inflammation in healthy adipose tissue and preadipocytes. When the senescent cells from which CM was derived had been treated with JAK inhibitors, the resulting CM was much less proinflammatory. The administration of JAK inhibitor to aged mice for 10 wk alleviated both adipose tissue and systemic inflammation and enhanced physical function. Our findings are consistent with a possible contribution of senescent cells and the SASP to age-related inflammation and frailty. We speculate that SASP inhibition by JAK inhibitors may contribute to alleviating frailty. Targeting the JAK pathway holds promise for treating age-related dysfunction.