Effects of canagliflozin on amputation risk in type 2 diabetes: the CANVAS Program.

AIMS/HYPOTHESIS: The primary analysis of the Canagliflozin cardioVascular Assessment Study (CANVAS) Program showed canagliflozin to have a beneficial effect on cardiovascular and renal outcomes in people with type 2 diabetes at high cardiovascular risk, but also an unexpected increased risk of major or minor lower extremity amputation. These secondary analyses explore this finding in more detail. METHODS: The effect of canagliflozin on amputation risk in the CANVAS Program was calculated for amputations of different types and proximate aetiologies and different canagliflozin doses. Univariate and multivariate associations of baseline characteristics with amputation risk were determined and proportional and absolute effects of canagliflozin were compared across subgroups. RESULTS: There were 187 (1.8%) participants with atraumatic lower extremity amputations (minor 71%, major 29%); as previously published, rates were 6.30 vs 3.37 per 1000 participant-years with canagliflozin vs placebo (HR 1.97 [95% CI 1.41, 2.75]). Risk was similar for ischaemic and infective aetiologies and for 100 mg and 300 mg doses. Overall amputation risk was strongly associated with baseline history of prior amputation (major or minor) (HR 21.31 [95% CI 15.40, 29.49]) and other established risk factors. No interactions between randomised treatment and participant characteristics explained the effect of canagliflozin on amputation risk. For every clinical subgroup studied, numbers of amputation events projected were smaller than numbers of major adverse cardiovascular events averted. CONCLUSIONS/INTERPRETATION: The CANVAS Program demonstrated that canagliflozin increased the risk of amputation (mainly minor) in this study population. Anticipated risk factors for amputation were identified, such as prior history of amputation, peripheral vascular disease and neuropathy, but no specific aetiological mechanism or at-risk subgroup for canagliflozin was identified.

Canagliflozin for Primary and Secondary Prevention of Cardiovascular Events: Results From the CANVAS Program (Canagliflozin Cardiovascular Assessment Study).

BACKGROUND: Canagliflozin is a sodium glucose cotransporter 2 inhibitor that significantly reduces the composite of cardiovascular death, nonfatal myocardial infarction, or nonfatal stroke in patients with type 2 diabetes mellitus and elevated cardiovascular risk. The comparative effects among participants with and without a history of cardiovascular disease (secondary versus primary prevention) were prespecified for evaluation. METHODS: The CANVAS Program (Canagliflozin Cardiovascular Assessment Study) randomly assigned 10 142 participants with type 2 diabetes mellitus to canagliflozin or placebo. The primary prevention cohort comprised individuals ≥50 years of age with ≥2 risk factors for cardiovascular events but with no prior cardiovascular event, and the secondary prevention cohort comprised individuals ≥30 years of age with a prior cardiovascular event. The primary end point was a composite of cardiovascular death, nonfatal myocardial infarction, or nonfatal stroke. Secondary outcomes included heart failure hospitalization and a renal composite (40% reduction in estimated glomerular filtration rate, renal replacement therapy, or renal death). RESULTS: Primary prevention participants (N=3486; 34%) were younger (63 versus 64 years of age), were more often female (45% versus 31%), and had a longer duration of diabetes mellitus (14 versus 13 years) compared with secondary prevention participants (N=6656; 66%). The primary end point event rate was higher in the secondary prevention group compared with the primary prevention group (36.9 versus 15.7/1000 patient-years, P<0.001). In the total cohort, the primary end point was reduced with canagliflozin compared with placebo (26.9 versus 31.5/1000 patient-years; hazard ratio [HR], 0.86; 95% confidence interval [CI], 0.75-0.97; P<0.001 for noninferiority, P=0.02 for superiority) with no statistical evidence of heterogeneity (interaction P value=0.18) between the primary (HR, 0.98; 95% CI, 0.74-1.30) and secondary prevention (HR, 0.82; 95% CI, 0.72-0.95) cohorts. Renal outcomes (HR, 0.59; 95% CI, 0.44-0.79 versus HR, 0.63; 95% CI, 0.39-1.02; interaction P value=0.73) and heart failure hospitalization (HR, 0.68; 95% CI, 0.51-0.90 versus HR, 0.64; 95% CI, 0.35-1.15; interaction P value=0.91) were similarly reduced in the secondary and primary prevention cohorts, respectively. Lower extremity amputations were similarly increased in the secondary and primary prevention cohorts (HR, 2.07; 95% CI, 1.43-3.00 versus HR, 1.52; 95% CI, 0.70-3.29; interaction P value=0.63). CONCLUSIONS: Patients with type 2 diabetes mellitus and prior cardiovascular events had higher rates of cardiovascular outcomes compared with the primary prevention patients. Canagliflozin reduced cardiovascular and renal outcomes with no statistical evidence of heterogeneity of the treatment effect across the primary and secondary prevention groups. Additional studies will provide further insights into the effects of canagliflozin in these patient populations. CLINICAL TRIAL REGISTRATION: URL: https://www.clinicaltrials.gov. Unique identifiers: NCT01032629 and NCT01989754.

Factors influencing longitudinal changes of circulating liver enzyme concentrations in subjects randomized to placebo in four clinical trials.

Liver enzyme concentrations are measured as safety end points in clinical trials to detect drug-related hepatotoxicity, but little is known about the epidemiology of these biomarkers in subjects without hepatic dysfunction who are enrolled in drug trials. We studied alanine and aspartate aminotransferase (ALT and AST) in subjects randomized to placebo who completed assessments over 36 mo in a cardiovascular outcome trial [the Stabilisation of Atherosclerotic Plaque by Initiation of Darapladib Therapy ("STABILITY") trial; n = 4,264; mean age: 64.2 yr] or over 12 mo in three trials that enrolled only subjects with type 2 diabetes (T2D) [the DIA trials; n = 308; mean age: 62.4 yr] to investigate time-dependent relationships and the factors that might affect ALT and AST, including body mass index (BMI), T2D, and renal function. Multivariate linear mixed models examined time-dependent relationships between liver enzyme concentrations as response variables and BMI, baseline T2D status, hemoglobin A1c levels, and renal function, as explanatory variables. At baseline, ALT was higher in individuals who were men, <65 yr old, and obese and who had glomerular filtration rate (GFR) >60 ml·min-1·1.73 m-2. ALT was not significantly associated with T2D at baseline, although it was positively associated with HbA1c. GFR had a greater impact on ALT than T2D. ALT concentrations decreased over time in subjects who lost weight but remained stable in individuals with increasing BMI. Weight change did not alter AST concentrations. We provide new insights on the influence of time, GFR, and HbA1c on ALT and AST concentrations and confirm the effect of sex, age, T2D, BMI, and BMI change in subjects receiving placebo in clinical trials. NEW & NOTEWORTHY Clinical trials provide high-quality data on liver enzyme concentrations from subjects randomized to placebo that can be used to investigate the epidemiology of these biomarkers. The adjusted models show the influence of sex, age, time, renal function, type 2 diabetes, HbA1c, and body mass index on alanine aminotransferase and aspartate aminotransferase concentrations and their relative importance. These factors need to be considered when assessing potential signals of hepatotoxicity in trials of new drugs and in clinical trials investigating subjects with nonalcoholic fatty liver disease.

Physiological Mechanisms of Weight Gain-Induced Steatosis in People With Obesity.

Weight gain is associated with an increase in intrahepatic triglycerides (IHTGs), and is the primary cause of nonalcoholic fatty liver disease in obese individuals. We combined imaging and stable isotope tracer techniques to evaluate the physiologic mechanisms of weight gain-induced steatosis in 27 obese people. Weight gain appeared to increase IHTG content by generating an imbalance between hepatic fatty acid availability and disposal, and resulted in increased hepatic de novo lipogenesis, decreased intrahepatic fatty acid oxidation, and inadequate increases in IHTG export via very low-density lipoprotein secretion. ClinicalTrials.gov ID NCT01184170.

Association between specific adipose tissue CD4+ T-cell populations and insulin resistance in obese individuals.

BACKGROUND & AIMS: An increased number of macrophages in adipose tissue is associated with insulin resistance and metabolic dysfunction in obese people. However, little is known about other immune cells in adipose tissue from obese people, and whether they contribute to insulin resistance. We investigated the characteristics of T cells in adipose tissue from metabolically abnormal insulin-resistant obese (MAO) subjects, metabolically normal insulin-sensitive obese (MNO) subjects, and lean subjects. Insulin sensitivity was determined by using the hyperinsulinemic euglycemic clamp procedure. METHODS: We assessed plasma cytokine concentrations and subcutaneous adipose tissue CD4(+) T-cell populations in 9 lean, 12 MNO, and 13 MAO subjects. Skeletal muscle and liver samples were collected from 19 additional obese patients undergoing bariatric surgery to determine the presence of selected cytokine receptors. RESULTS: Adipose tissue from MAO subjects had 3- to 10-fold increases in numbers of CD4(+) T cells that produce interleukin (IL)-22 and IL-17 (a T-helper [Th] 17 and Th22 phenotype) compared with MNO and lean subjects. MAO subjects also had increased plasma concentrations of IL-22 and IL-6. Receptors for IL-17 and IL-22 were expressed in human liver and skeletal muscle samples. IL-17 and IL-22 inhibited uptake of glucose in skeletal muscle isolated from rats and reduced insulin sensitivity in cultured human hepatocytes. CONCLUSIONS: Adipose tissue from MAO individuals contains increased numbers of Th17 and Th22 cells, which produce cytokines that cause metabolic dysfunction in liver and muscle in vitro. Additional studies are needed to determine whether these alterations in adipose tissue T cells contribute to the pathogenesis of insulin resistance in obese people.

Metabolic response to high-carbohydrate and low-carbohydrate meals in a nonhuman primate model.

We established a model of chronic portal vein catheterization in an awake nonhuman primate to provide a comprehensive evaluation of the metabolic response to low-carbohydrate/high-fat (LCHF; 20% carbohydrate and 65% fat) and high-carbohydrate/low-fat (HCLF; 65% carbohydrate and 20% fat) meal ingestion. Each meal was given 1 wk apart to five young adult (7.8 ± 1.3 yr old) male baboons. A [U-¹³C]glucose tracer was added to the meal, and a [6,6-²H2]glucose tracer was infused systemically to assess glucose kinetics. Plasma areas under the curve (AUCs) of glucose, insulin, and C-peptide in the femoral artery and of glucose and insulin in the portal vein were higher (P ≤ 0.05) after ingestion of the HCLF compared with the LCHF meal. Compared with the LCHF meal, the rate of appearance of ingested glucose into the portal vein and the systemic circulation was greater after the HCLF meal (P < 0.05). Endogenous glucose production decreased by ∼40% after ingestion of the HCLF meal but was not affected by the LCHF meal (P < 0.05). Portal vein blood flow increased (P < 0.001) to a similar extent after consumption of either meal. In conclusion, a LCHF diet causes minimal changes in the rate of glucose appearance in both portal and systemic circulations, does not affect the rate of endogenous glucose production, and causes minimal stimulation of C-peptide and insulin. These observations demonstrate that LCHF diets cause minimal perturbations in glucose homeostasis and pancreatic ß-cell activity.

Intrahepatic diacylglycerol content is associated with hepatic insulin resistance in obese subjects.

Data from studies in animal models indicate that certain lipid metabolites, particularly diacylglycerol, ceramide, and acylcarnitine, disrupt insulin action. We evaluated the relationship between the presence of these metabolites in the liver (assessed by mass spectrometry) and hepatic insulin sensitivity (assessed using a hyperinsulinemic-euglycemic clamp with stable isotope tracer infusion) in 16 obese adults (body mass index, 48 ± 9 kg/m²). There was a negative correlation between insulin-mediated suppression of hepatic glucose production and intrahepatic diacylglycerol (r = -0.609; P = .012), but not with intrahepatic ceramide or acylcarnitine. These data indicate that intrahepatic diacylglycerol is an important mediator of hepatic insulin resistance in obese people with nonalcoholic fatty liver disease.

Validation of a novel index to assess insulin resistance of adipose tissue lipolytic activity in obese subjects.

Insulin resistance in adipose tissue increases the release of free fatty acids into the circulation, which likely contributes to impaired insulin action in liver and skeletal muscle associated with obesity. However, reliable assessment of adipose tissue insulin resistance requires performing a hyperinsulinemic-euglycemic clamp procedure in conjunction with a fatty acid tracer infusion to determine insulin-mediated suppression of lipolytic rate. We developed a simpler method for evaluating adipose tissue insulin resistance in vivo, determined as the product of palmitate rate of appearance into the bloodstream and plasma insulin concentration during basal conditions. We validated our Adipose Tissue Insulin Resistance Index (ATIRI) by comparison with an assessment of adipose tissue insulin resistance determined by using the hyperinsulinemic-euglycemic clamp procedure in conjunction with a palmitate tracer infusion in 47 obese nondiabetic subjects (body mass index: 40.1 ± 9.3 kg/m(2)). We found the ATIRI correlated closely with adipose tissue insulin resistance assessed during the clamp procedure (r =-0.854, P < 0.001). These results demonstrate that the ATIRI provides a reliable index of adipose tissue insulin resistance in obese subjects.

Evidence for regulated monoacylglycerol acyltransferase expression and activity in human liver.

Intrahepatic lipid accumulation is extremely common in obese subjects and is associated with the development of insulin resistance and diabetes. Hepatic diacylglycerol and triacylglycerol synthesis predominantly occurs through acylation of glycerol-3-phosphate. However, an alternative pathway for synthesizing diacylglycerol from monoacylglycerol acyltransferases (MGAT) could also contribute to hepatic glyceride pools. MGAT activity and the expression of the three genes encoding MGAT enzymes (MOGAT1, MOGAT2, and MOGAT3) were determined in liver biopsies from obese human subjects before and after gastric bypass surgery. MOGAT expression was also assessed in liver of subjects with nonalcoholic fatty liver disease (NAFLD) or control livers. All MOGAT genes were expressed in liver, and hepatic MGAT activity was readily detectable in liver lysates. The hepatic expression of MOGAT3 was highly correlated with MGAT activity, whereas MOGAT1 and MOGAT2 expression was not, and knockdown of MOGAT3 expression attenuated MGAT activity in a liver-derived cell line. Marked weight loss following gastric bypass surgery was associated with a significant reduction in MOGAT2 and MOGAT3 expression, which were also overexpressed in NAFLD subjects. These data suggest that the MGAT pathway is active and dynamically regulated in human liver and could be an important target for pharmacologic intervention for the treatment of obesity-related insulin resistance and NAFLD.

Intrahepatic fat, not visceral fat, is linked with metabolic complications of obesity.

Visceral adipose tissue (VAT) is an important risk factor for obesity-related metabolic disorders. Therefore, a reduction in VAT has become a key goal in obesity management. However, VAT is correlated with intrahepatic triglyceride (IHTG) content, so it is possible that IHTG, not VAT, is a better marker of metabolic disease. We determined the independent association of IHTG and VAT to metabolic function, by evaluating groups of obese subjects, who differed in IHTG content (high or normal) but matched on VAT volume or differed in VAT volume (high or low) but matched on IHTG content. Stable isotope tracer techniques and the euglycemic-hyperinsulinemic clamp procedure were used to assess insulin sensitivity and very-low-density lipoprotein-triglyceride (VLDL-TG) secretion rate. Tissue biopsies were obtained to evaluate cellular factors involved in ectopic triglyceride accumulation. Hepatic, adipose tissue and muscle insulin sensitivity were 41, 13, and 36% lower (P < 0.01), whereas VLDL-triglyceride secretion rate was almost double (P < 0.001), in subjects with higher than normal IHTG content, matched on VAT. No differences in insulin sensitivity or VLDL-TG secretion were observed between subjects with different VAT volumes, matched on IHTG content. Adipose tissue CD36 expression was lower (P < 0.05), whereas skeletal muscle CD36 expression was higher (P < 0.05), in subjects with higher than normal IHTG. These data demonstrate that IHTG, not VAT, is a better marker of the metabolic derangements associated with obesity. Furthermore, alterations in tissue fatty acid transport could be involved in the pathogenesis of ectopic triglyceride accumulation by redirecting plasma fatty acid uptake from adipose tissue toward other tissues.

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

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

Insulin sensitivity is not associated with palmitoleate availability in obese humans.

We evaluated whether insulin resistance in obese people is associated with decreased plasma palmitoleate availability. Palmitoleate content (percentage and absolute concentrations) in FFA and VLDL was measured in obese subjects who were either insulin resistant (IR) or insulin sensitive (IS), based on assessment of multiorgan (skeletal muscle, liver, and adipose tissue) insulin sensitivity by using the hyperinsulinemic-euglycemic clamp procedure in conjunction with infusion of stable isotopically labeled tracers. Plasma palmitoleate concentration and the relative contribution of palmitoleate to total plasma FFA concentration in the IS group (0.018 ± 0.002 mmol/l and 4.4% ± 0.2%, respectively) were not significantly different than values in the IR group (0.023 ± 0.003 mmol/l and 4.4% ± 0.4%, respectively). Plasma VLDL-triglyceride palmitoleate concentration and the proportion of VLDL fatty acids as palmitoleate in the IS group (0.09 ± 0.02 mmol/l and 5.7 ± 0.3%, respectively) were also not significantly different than those in the IR group (0.16 ± 0.04 mmol/l and 5.0% ± 0.4%, respectively). These data demonstrate that decreased palmitoleate in plasma and in VLDL is not associated with insulin resistance in skeletal muscle, liver, or adipose tissue in obese people.

Dissociation between intrahepatic triglyceride content and insulin resistance in familial hypobetalipoproteinemia.

BACKGROUND & AIMS: Hepatic steatosis is associated with insulin resistance, but it is not clear whether increased intrahepatic triglyceride (IHTG) content causes the resistance or is a marker. Subjects with familial hypobetalipoproteinemia (FHBL) have high levels of IHTG because of a genetic defect in hepatic export of triglycerides, and provide a unique cohort to study the relationship between steatosis and insulin sensitivity. METHODS: One group of lean subjects with normal IHTG content (2.2% +/- 0.6% of liver volume) (n = 6), and 3 groups of overweight and obese subjects matched for body mass index, were studied: (1) normal IHTG content (3.3% +/- 0.5%; n = 6), (2) high IHTG content (21.4% +/- 2.6%) due to nonalcoholic fatty liver disease (NAFLD; n = 6), and (3) high IHTG content (18.1% +/- 2.2%) due to FHBL (n = 3). A hyperinsulinemic-euglycemic clamp procedure, in conjunction with glucose tracer infusion, was used to determine multiorgan insulin sensitivity. RESULTS: Hepatic insulin sensitivity (reciprocal of glucose rate of appearance [micromol x kg fat-free mass(-1) x min(-1)] x insulin [mU/L]) was greatest in the Lean group (2.0 +/- 0.4); it was the same among subjects with FHBL (0.8 +/- 0.1) and the group with normal IHTG content, matched for body mass index (0.7 +/- 0.1), but greater than the NAFLD group (0.3 +/- 0.1) (P < .01). Muscle insulin sensitivity (percent increase in glucose uptake during insulin infusion) was greatest in the Lean group (576% +/- 70%). Muscle insulin sensitivity was similar in subjects with FHBL and those with normal IHTG (319% +/- 77%, 326% +/- 27%, respectively), but greater than the NAFLD group (145% +/- 18%) (P < .01). CONCLUSIONS: Steatosis is dissociated from insulin resistance in FHBL, which suggests that increased IHTG content is a marker, not a cause, of metabolic dysfunction.

Surgical removal of omental fat does not improve insulin sensitivity and cardiovascular risk factors in obese adults.

BACKGROUND & AIMS: Visceral adipose tissue (VAT) is an important risk factor for the metabolic complications associated with obesity. Therefore, a reduction in VAT is considered an important target of obesity therapy. We evaluated whether reducing VAT mass by surgical removal of the omentum improves insulin sensitivity and metabolic function in obese patients. METHODS: We conducted a 12-month randomized controlled trial to determine whether reducing VAT by omentectomy in 22 obese subjects increased their improvement following Roux-en-Y gastric bypass (RYGB) surgery in hepatic and skeletal muscle sensitivity to insulin study 1. Improvement was assessed by using the hyperinsulinemic-euglycemic clamp technique. We also performed a 3-month, longitudinal, single-arm study to determine whether laparoscopic omentectomy alone, in 7 obese subjects with type 2 diabetes mellitus (T2DM), improved insulin sensitivity study 2. Improvement was assessed by using the Frequently Sampled Intravenous Glucose Tolerance Test. RESULTS: The greater omentum, which weighed 0.82 kg (95% confidence interval: 0.67-0.97), was removed from subjects who had omentectomy in both studies. In study 1, there was an approximate 2-fold increase in muscle insulin sensitivity (relative increase in glucose disposal during insulin infusion) and a 4-fold increase in hepatic insulin sensitivity 12 months after RYGB alone and RYGB plus omentectomy, compared with baseline values (P<.001). There were no significant differences between groups (P>.87) or group x time interactions (P>.36). In study 2, surgery had no effect on insulin sensitivity (P=.844) or use of diabetes medications. CONCLUSIONS: These results demonstrate that decreasing VAT through omentectomy, alone or in combination with RYGB surgery, does not improve metabolic function in obese patients.

Obesity and nonalcoholic fatty liver disease: biochemical, metabolic, and clinical implications.

Obesity is associated with an increased risk of nonalcoholic fatty liver disease (NAFLD). Steatosis, the hallmark feature of NAFLD, occurs when the rate of hepatic fatty acid uptake from plasma and de novo fatty acid synthesis is greater than the rate of fatty acid oxidation and export (as triglyceride within very low-density lipoprotein). Therefore, an excessive amount of intrahepatic triglyceride (IHTG) represents an imbalance between complex interactions of metabolic events. The presence of steatosis is associated with a constellation of adverse alterations in glucose, fatty acid, and lipoprotein metabolism. It is likely that abnormalities in fatty acid metabolism, in conjunction with adipose tissue, hepatic, and systemic inflammation, are key factors involved in the development of insulin resistance, dyslipidemia, and other cardiometabolic risk factors associated with NAFLD. However, it is not clear whether NAFLD causes metabolic dysfunction or whether metabolic dysfunction is responsible for IHTG accumulation, or possibly both. Understanding the precise factors involved in the pathogenesis and pathophysiology of NAFLD will provide important insights into the mechanisms responsible for the cardiometabolic complications of obesity.

Interaction between the UCP2 -866 G>A polymorphism, diabetes, and beta-blocker use among patients with acute coronary syndromes.

OBJECTIVE: UCP2 -866G>A (rs659366) has been implicated in cardiometabolic disease and represents a novel candidate gene for beta-blocker response, particularly among patients with diabetes. We assessed the function of -866G>A and its role as a modifier of beta-blocker treatment outcomes by diabetes status in an acute coronary syndrome (ACS) cohort. METHODS: ACS patients with genetic samples and 12 months of follow-up for cardiac rehospitalizations or death (n=468) were assessed. The influence of -866G>A on beta-blocker treatment outcomes was evaluated in those with diabetes and without. To assess functional correlates of -866G>A, we compared uncoupling protein 2 (UCP2) expression in the skeletal muscle of obese participants by genotype and compared the activity of UCP2 luciferase promoters with -866G and -866A alleles. RESULTS: An interaction between -866G>A and beta-blocker treatment was found in individuals with diabetes (P=0.002) but not those without (P=0.79). Among G/G individuals with diabetes, discharge beta-blocker use was associated with an 80% reduction in cardiac rehospitalization (adjusted hazard ratio: 0.20; 95% confidence interval: 0.04-1.02). In contrast, among A-carrier patients with diabetes, there was an 11-fold increase in cardiac rehospitalizations with discharge beta-blocker therapy (adjusted hazard ratio: 11.75; 95% confidence interval: 1.28-108.2). Promoter activity assays showed that -866G had greater cyclic AMP response element binding protein-responsiveness compared with -866A, and compared with -866A carriers G/G individuals exhibited increased UCP2 expression in the skeletal muscle. CONCLUSION: We identified a significant interaction between -866G>A and beta-blocker response among ACS patients with diabetes. Furthermore, -866G conferred greater gene transcriptional activity than -866A in cell lines and in obese patients. These findings may help us gain insight into the mechanisms underlying the beneficial and detrimental effects of beta-blockers in those with diabetes.

Liver, muscle, and adipose tissue insulin action is directly related to intrahepatic triglyceride content in obese subjects.

BACKGROUND & AIMS: Nonalcoholic fatty liver disease is associated with insulin resistance and diabetes. The purpose of this study was to determine the relationship between intrahepatic triglyceride (IHTG) content and insulin action in liver (suppression of glucose production), skeletal muscle (stimulation of glucose uptake), and adipose tissue (suppression of lipolysis) in nondiabetic obese subjects. METHODS: A euglycemic-hyperinsulinemic clamp procedure and stable isotopically labeled tracer infusions were used to assess insulin action, and magnetic resonance spectroscopy was used to determine IHTG content, in 42 nondiabetic obese subjects (body mass index, 36 +/- 4 kg/m(2)) who had a wide range of IHTG content (1%-46%). RESULTS: Hepatic insulin sensitivity, assessed as a function of glucose production rate and plasma insulin concentration, was inversely correlated with IHTG content (r = -0.599; P < .001). The ability of insulin to suppress fatty acid release from adipose tissue and to stimulate glucose uptake by skeletal muscle were also inversely correlated with IHTG content (adipose tissue: r = -0.590, P < .001; skeletal muscle: r = -0.656, P < .001). Multivariate linear regression analyses found that IHTG content was the best predictor of insulin action in liver, skeletal muscle, and adipose tissue, independent of body mass index and percent body fat, and accounted for 34%, 42%, and 44% of the variability in these tissues, respectively (P < .001 for each model). CONCLUSIONS: These results show that progressive increases in IHTG content are associated with progressive impairment of insulin action in liver, skeletal muscle, and adipose tissue in nondiabetic obese subjects. Therefore, nonalcoholic fatty liver disease should be considered part of a multiorgan system derangement in insulin sensitivity.

Alterations in adipose tissue and hepatic lipid kinetics in obese men and women with nonalcoholic fatty liver disease.

BACKGROUND & AIMS: Steatosis in patients with nonalcoholic fatty liver disease (NAFLD) is due to an imbalance between intrahepatic triglyceride (IHTG) production and export. The purpose of this study was to evaluate TG metabolism in adipose tissue and liver in NAFLD. METHODS: Fatty acid, VLDL-TG, and VLDL-apolipoprotein B-100 (apoB100) kinetics were assessed by using stable isotope tracers in 14 nondiabetic obese subjects with NAFLD (IHTG, 22.7% +/- 2.0%) and 14 nondiabetic obese subjects with normal IHTG content (IHTG, 3.4% +/- 0.4%), matched on age, sex, body mass index, and percent body fat. RESULTS: Compared with the normal IHTG group, the NAFLD group had greater rates of palmitate release from adipose tissue into plasma (85.4 +/- 6.6 and 114.1 +/- 8.1 micromol/min, respectively; P = .01) and VLDL-TG secretion (11.4 +/- 1.1 and 24.3 +/- 3.1 micromol/min, respectively; P = .001); VLDL-apoB100 secretion rates were not different between groups. The increase in VLDL-TG secretion was primarily due to an increased contribution from "nonsystemic" fatty acids, presumably derived from lipolysis of intrahepatic and intra-abdominal fat and de novo lipogenesis. VLDL-TG secretion rate increased linearly with increasing IHTG content in subjects with normal IHTG but reached a plateau when IHTG content was >/=10% (r = 0.618, P < .001). CONCLUSIONS: Obese persons with NAFLD have marked alterations in both adipose tissue (increased lipolytic rates) and hepatic (increased VLDL-TG secretion) TG metabolism. Fatty acids derived from nonsystemic sources are responsible for the increase in VLDL-TG secretion. However, the increase in hepatic TG export is not adequate to normalize IHTG content.

Methods for assessing intrahepatic fat content and steatosis.

PURPOSE OF REVIEW: Intrahepatic fat content is increasingly being recognized as an integral part of metabolic dysfunction. This article reviews available methods for the assessment of hepatic steatosis. RECENT FINDINGS: Apart from liver biopsy, there are several noninvasive radiologic modalities for evaluating nonalcoholic fatty liver disease. Ultrasonography, computed tomography, and traditional MRI remain largely qualitative methods for detecting mild to severe degrees of steatosis rather than quantitative methods for measuring liver fat content, even though novel attempts to collect objective quantitative information have recently been developed. Still, their sensitivity at mild degrees of steatosis is poor. Undoubtedly, most methodological advances have occurred in the field of MRI and magnetic resonance spectroscopy, which currently enable the accurate quantification of intrahepatic fat even at normal or near normal levels. Xenon computed tomography was also recently shown to offer another objective tool for the quantitative assessment of steatosis, although more validation studies are required. SUMMARY: Several modalities can be used for measuring intrahepatic fat and assessing steatosis; the choice will ultimately depend on the intended use and available resources.