Loss of α-actinin-3 during human evolution provides superior cold resilience and muscle heat generation.

The protein α-actinin-3 expressed in fast-twitch skeletal muscle fiber is absent in 1.5 billion people worldwide due to homozygosity for a nonsense polymorphism in ACTN3 (R577X). The prevalence of the 577X allele increased as modern humans moved to colder climates, suggesting a link between α-actinin-3 deficiency and improved cold tolerance. Here, we show that humans lacking α-actinin-3 (XX) are superior in maintaining core body temperature during cold-water immersion due to changes in skeletal muscle thermogenesis. Muscles of XX individuals displayed a shift toward more slow-twitch isoforms of myosin heavy chain (MyHC) and sarcoplasmic reticulum (SR) proteins, accompanied by altered neuronal muscle activation resulting in increased tone rather than overt shivering. Experiments on Actn3 knockout mice showed no alterations in brown adipose tissue (BAT) properties that could explain the improved cold tolerance in XX individuals. Thus, this study provides a mechanism for the positive selection of the ACTN3 X-allele in cold climates and supports a key thermogenic role of skeletal muscle during cold exposure in humans.

Sex differences in adipose insulin resistance are linked to obesity, lipolysis and insulin receptor substrate 1.

BACKGROUND/OBJECTIVE: Insulin resistance is more prominent in men than women. If this involves adipose tissue is unknown and was presently examined. SUBJECTS/METHODS: AdipoIR (in vivo adipose insulin resistance index) was measured in 2344 women and 787 men. In 259 of the women and 54 of the men, insulin induced inhibition of lipolysis (acylglycerol breakdown) and stimulation of lipogenesis (glucose conversion to acylglycerols) were determined in subcutaneous adipocytes; in addition, basal (spontaneous) lipolysis was also determined in the fat cells. In 234 women and 115 men, RNAseq expression of canonical insulin signal genes were measured in subcutaneous adipose tissue. Messenger RNA transcripts of the most discriminant genes were quantified in 175 women and 109 men. RESULTS: Men had higher AdipoIR values than women but only when obesity (body mass index 30 kg/m2 or more) was present (p < 0.0001). The latter sex dimorphism was found among physically active and sedentary people, in those with and without cardiometabolic disease and in people using nicotine or not (p = 0.0003 or less). In obesity, adipocyte insulin sensitivity (half maximum effective hormone concentration) and maximal antilipolytic effect were tenfold and 10% lower, respectively, in men than women (p = 0.005 or less). Basal rate of lipolysis was two times higher in men than women (p > 0.0001). Sensitivity and maximum effect of insulin on lipogenesis were similar in both sexes (p = 0.26 and p = 0.18, respectively). When corrected for multiple comparison only RNAseq expression of insulin receptor substrate 1 (IRS1) was lower in men than women (p < 0.0001). The mRNA transcript for IRS1 was 60% higher in women than men (p < 0.0001). CONCLUSIONS: In obesity, adipose tissue insulin resistance is more pronounced in men than in women. The mechanism involves less efficient insulin-mediated inhibition of adipocyte lipolysis, increased basal rate of lipolysis and decreased adipose expression of a key element of insulin signaling, IRS1.

Increased tetanic calcium in early fatigue of mammalian muscle fibers is accompanied by accelerated force development despite a decreased force.

During the initial phase of fatigue induced by repeated contractions in fast-twitch muscle fibers, tetanic force decreases despite increasing tetanic free cytosolic [Ca2+ ] ([Ca2+ ]cyt ). Here, we hypothesized that the increase in tetanic [Ca2+ ]cyt nevertheless has positive effects on force in early fatigue. Experiments on enzymatically isolated mouse flexor digitorum brevis (FDB) fibers showed that an increase in tetanic [Ca2+ ]cyt during ten 350 ms contractions required trains of electrical pulses to be elicited at short intervals (≤2 s) and at high frequencies (≥70 Hz). Mechanically dissected mouse FDB fibers showed greater decrease in tetanic force when the stimulation frequency during contractions was gradually reduced to prevent the increase in tetanic [Ca2+ ]cyt . Novel analyses of data from previous studies revealed an increased rate of force development in the tenth fatiguing contraction in mouse FDB fibers, as well as in rat FDB and human intercostal fibers. Mouse FDB fibers deficient in creatine kinase showed no increase in tetanic [Ca2+ ]cyt and slowed force development in the tenth contraction; after injection of creatine kinase to enable phosphocreatine breakdown, these fibers showed an increase in tetanic [Ca2+ ]cyt and accelerated force development. Mouse FDB fibers exposed to ten short contractions (43 ms) produced at short intervals (142 ms) showed increased tetanic [Ca2+ ]cyt accompanied by a marked (~16%) increase in the developed force. In conclusion, the increase in tetanic [Ca2+ ]cyt in early fatigue is accompanied by accelerated force development, which under some circumstances can counteract the decline in physical performance caused by the concomitant decrease in maximum force.

Adipose Insulin Resistance Associates With Dyslipidemia Independent of Liver Resistance and Involves Early Hormone Signaling.

BACKGROUND: Adipose tissue insulin resistance is linked to altered plasma levels of triglycerides and HDL (high-density lipoprotein)-cholesterol. However, its degree of independence from liver resistance and different metabolic traits (lipolysis, lipogenesis) effected is not clear and was presently investigated. METHODS: In 3290 adult subjects, plasma levels of triglycerides and HDL-cholesterol were cross-sectionally measured and related to interindividual variations in measures of insulin resistance in the liver (homeostasis mode assessment of insulin resistance index) or adipose tissue (Adipo-IR index). In subgroups, insulin-induced antilipolysis and lipogenesis in isolated subcutaneous fat cells (n=578) were determined alongside global adipose tissue gene expression (n=132). RESULTS: Using linear regression, homeostasis mode assessment of insulin resistance and Adipo-IR strongly correlated with the plasma lipids explaining 33% of the variations in triglycerides. Together with other variables (age, sex, body mass index, cardiometabolic disorders, nicotine use, ethnicity, and physical activity) in multiple regression, homeostasis mode assessment of insulin resistance, and Adipo-IR each remained an important regressor for triglycerides and HDL-cholesterol (P<0.0001). In fat cells, half-maximum effective concentration but not maximum effect of insulin on antilipolysis and lipogenesis contributed independently to variations in triglycerides and HDL-cholesterol (P=0.001 or lower). This was linked to expression of the insulin receptor, insulin receptor substrate-1, and AKT serine/threonine kinase 2 in adipose tissue. CONCLUSIONS: Markers of insulin resistance in the liver and adipose tissue each associate strongly, and independently of each other, to elevated triglycerides and decreased HDL levels. At the fat cell, early insulin receptor signaling and sensitivity, but not maximum insulin action contributes to the variations in circulating triglycerides and HDL-cholesterol.

Acyl ghrelin improves cardiac function in heart failure and increases fractional shortening in cardiomyocytes without calcium mobilization.

BACKGROUND AND AIMS: Ghrelin is an endogenous appetite-stimulating peptide hormone with potential cardiovascular benefits. Effects of acylated (activated) ghrelin were assessed in patients with heart failure and reduced ejection fraction (HFrEF) and in ex vivo mouse cardiomyocytes. METHODS AND RESULTS: In a randomized placebo-controlled double-blind trial, 31 patients with chronic HFrEF were randomized to synthetic human acyl ghrelin (0.1 µg/kg/min) or placebo intravenously over 120 min. The primary outcome was change in cardiac output (CO). Isolated mouse cardiomyocytes were treated with acyl ghrelin and fractional shortening and calcium transients were assessed. Acyl ghrelin but not placebo increased cardiac output (acyl ghrelin: 4.08 ± 1.15 to 5.23 ± 1.98 L/min; placebo: 4.26 ± 1.23 to 4.11 ± 1.99 L/min, P < 0.001). Acyl ghrelin caused a significant increase in stroke volume and nominal increases in left ventricular ejection fraction and segmental longitudinal strain and tricuspid annular plane systolic excursion. There were no effects on blood pressure, arrhythmias, or ischaemia. Heart rate decreased nominally (acyl ghrelin: 71 ± 11 to 67 ± 11 b.p.m.; placebo 69 ± 8 to 68 ± 10 b.p.m.). In cardiomyocytes, acyl ghrelin increased fractional shortening, did not affect cellular Ca2+ transients, and reduced troponin I phosphorylation. The increase in fractional shortening and reduction in troponin I phosphorylation was blocked by the acyl ghrelin antagonist D-Lys 3. CONCLUSION: In patients with HFrEF, acyl ghrelin increased cardiac output without causing hypotension, tachycardia, arrhythmia, or ischaemia. In isolated cardiomyocytes, acyl ghrelin increased contractility independently of preload and afterload and without Ca2+ mobilization, which may explain the lack of clinical side effects. Ghrelin treatment should be explored in additional randomized trials. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT05277415.

Treatment Monitoring of a Patient with Synchronous Metastatic Angiosarcoma and Breast Cancer Using ctDNA.

Angiosarcoma is a rare and aggressive type of soft-tissue sarcoma with high propensity to metastasize. For patients with metastatic angiosarcoma, prognosis is dismal and treatment options are limited. To improve the outcomes, identifying patients with poor treatment response at an earlier stage is imperative, enabling alternative therapy. Consequently, there is a need for improved methods and biomarkers for treatment monitoring. Quantification of circulating tumor-DNA (ctDNA) is a promising approach for patient-specific monitoring of treatment response. In this case report, we demonstrate that quantification of ctDNA using SiMSen-Seq was successfully utilized to monitor a patient with metastatic angiosarcoma. By quantifying ctDNA levels using 25 patient-specific mutations in blood plasma throughout surgery and palliative chemotherapy, we predicted the outcome and monitored the clinical response to treatment. This was accomplished despite the additional complexity of the patient having a synchronous breast cancer. The levels of ctDNA showed a superior correlation to the clinical outcome compared with the radiological evaluations. Our data propose a promising approach for personalized biomarker analysis to monitor treatment in angiosarcomas, with potential applicability to other cancers and for patients with synchronous malignancies.

Associations between rheumatoid arthritis, incident heart failure, and left ventricular ejection fraction.

BACKGROUND: Rheumatoid arthritis (RA) is an independent risk factor for heart failure (HF). Yet, the association between RA and left ventricular ejection fraction (LVEF) in incident HF is not well studied, nor are outcomes of HF in RA by LVEF. METHODS: We identified incident HF patients between 2003 and 2018 through the Swedish Heart Failure Registry, enriched with data from national health registers. Using logistic regression, associations between a prior diagnosis of RA and LVEF among HF patients and vs age, sex, and geographical area matched general population controls without HF were assessed. Additionally, associations between HF with vs without a prior diagnosis of RA, by LVEF, and outcomes up to 5 years after HF diagnosis were investigated using Cox regression. LVEF was primarily dichotomized at 40% and secondarily categorized as <40%, 40% to 49%, and ≥50%. Covariates included demographics and cardiovascular comorbidities. RESULTS: Among 20,916 incident HF patients, 331 (1.6%) had RA vs 1,047/103,501 (1.0%) of HF-free controls. The odds ratio (OR) for RA was 1.4 (95% CI: 1.1-1.8) in LVEF<40% vs HF-free controls and 1.6 (95% CI: 1.3-2.0) in LVEF≥40% vs HF-free controls. Among HF patients, RA was more common in HF with LVEF ≥40% (1.9%) vs LVEF<40% (1.3%), corresponding to OR 1.4 (95% CI: 1.1-1.7). No associations between RA and cardiovascular outcomes were observed across LVEF. An association between RA and all-cause mortality was observed only for patients with LVEF<40% (hazard ratio: 1.4; 95% CI: 1.1-1.8). CONCLUSIONS: RA was independently associated with incident HF, particularly HF with LVEF≥40%. RA did not associate with cardiovascular outcomes following HF diagnosis but was associated with increased risk of all-cause mortality in HF with LVEF<40%.

Adipose tissue specific CCL18 associates with cardiometabolic diseases in non-obese individuals implicating CD4+ T cells.

AIM: Obesity is linked to cardiometabolic diseases, however non-obese individuals are also at risk for type 2 diabetes (T2D) and cardiovascular disease (CVD). White adipose tissue (WAT) is known to play a role in both T2D and CVD, but the contribution of WAT inflammatory status especially in non-obese patients with cardiometabolic diseases is less understood. Therefore, we aimed to find associations between WAT inflammatory status and cardiometabolic diseases in non-obese individuals. METHODS: In a population-based cohort containing non-obese healthy (n = 17), T2D (n = 16), CVD (n = 18), T2D + CVD (n = 19) individuals, seventeen different cytokines were measured in WAT and in circulation. In addition, 13-color flow cytometry profiling was employed to phenotype the immune cells. Human T cell line (Jurkat T cells) was stimulated by rCCL18, and conditioned media (CM) was added to the in vitro cultures of human adipocytes. Lipolysis was measured by glycerol release. Blocking antibodies against IFN-γ and TGF-ß were used in vitro to prove a role for these cytokines in CCL18-T-cell-adipocyte lipolysis regulation axis. RESULTS: In CVD, T2D and CVD + T2D groups, CCL18 and CD4+ T cells were upregulated significantly compared to healthy controls. WAT CCL18 secretion correlated with the amounts of WAT CD4+ T cells, which also highly expressed CCL18 receptors suggesting that WAT CD4+ T cells are responders to this chemokine. While direct addition of rCCL18 to mature adipocytes did not alter the adipocyte lipolysis, CM from CCL18-treated T cells increased glycerol release in in vitro cultures of adipocytes. IFN-γ and TGF-ß secretion was significantly induced in CM obtained from T cells treated with CCL18. Blocking these cytokines in CM, prevented CM-induced upregulation of adipocyte lipolysis. CONCLUSION: We suggest that in T2D and CVD, increased production of CCL18 recruits and activates CD4+ T cells to secrete IFN-γ and TGF-ß. This, in turn, promotes adipocyte lipolysis - a possible risk factor for cardiometabolic diseases.

Absolute Quantification of Donor-Derived Cell-Free DNA in Pediatric and Adult Patients After Heart Transplantation: A Prospective Study.

In this prospective study we investigated a cohort after heart transplantation with a novel PCR-based approach with focus on treated rejection. Blood samples were collected coincidentally to biopsies, and both absolute levels of dd-cfDNA and donor fraction were reported using digital PCR. 52 patients (11 children and 41 adults) were enrolled (NCT03477383, clinicaltrials.gov), and 557 plasma samples were analyzed. 13 treated rejection episodes >14 days after transplantation were observed in 7 patients. Donor fraction showed a median of 0.08% in the cohort and was significantly elevated during rejection (median 0.19%, p < 0.0001), using a cut-off of 0.1%, the sensitivity/specificity were 92%/56% (AUC ROC-curve: 0.78). Absolute levels of dd-cfDNA showed a median of 8.8 copies/mL and were significantly elevated during rejection (median 23, p = 0.0001). Using a cut-off of 7.5 copies/mL, the sensitivity/specificity were 92%/43% for donor fraction (AUC ROC-curve: 0.75). The results support the feasibility of this approach in analyzing dd-cfDNA after heart transplantation. The obtained values are well aligned with results from other trials. The possibility to quantify absolute levels adds important value to the differentiation between ongoing graft damage and quiescent situations.

Lipolysis defect in people with obesity who undergo metabolic surgery.

OBJECTIVE: Cross-sectional studies demonstrate that catecholamine stimulation of fat cell lipolysis is blunted in obesity. We investigated whether this defect persists after substantial weight loss has been induced by metabolic surgery, and whether it is related to the outcome. DESIGN/METHODS: Patients with obesity not able to successfully reduce body weight by conventional means (n = 126) were investigated before and 5 years after Roux-en-Y gastric bypass surgery (RYGB). They were compared with propensity-score matched subjects selected from a control group (n = 1017), and with the entire group after adjustment for age, sex, body mass index (BMI), fat cell volume and other clinical parameters. Catecholamine-stimulated lipolysis (glycerol release) was investigated in isolated fat cells using noradrenaline (natural hormone) or isoprenaline (synthetic beta-adrenoceptor agonist). RESULTS: Following RYGB, BMI was reduced from 39.9 (37.5-43.5) (median and interquartile range) to 29.5 (26.7-31.9) kg/m2 (p < 0.0001). The post-RYGB patients had about 50% lower lipolysis rates compared with the matched and total series of controls (p < 0.0005). Nordrenaline activation of lipolysis at baseline was associated with the RYGB effect; those with high lipolysis activation (upper tertile) lost 30%-45% more in body weight, BMI or fat mass than those with low (bottom tertile) initial lipolysis activation (p < 0.0007). CONCLUSION: Patients with obesity requiring metabolic surgery have impaired ability of catecholamines to stimulate lipolysis, which remains despite long-term normalization of body weight by RYGB. Furthermore, preoperative variations in the ability of catecholamines to activate lipolysis may predict the long-term reduction in body weight and fat mass.

Subcutaneous adipose tissue expansion mechanisms are similar in early and late onset overweight/obesity.

BACKGROUND/OBJECTIVE: The development of overweight/obesity associates with alterations in white adipose tissue (WAT) cellularity (fat cell size/number) and lipid metabolism, in particular lipolysis. If these changes differ between early/juvenile (EOO < 18 years of age) or late onset overweight/obesity (LOO) is unknown and was presently examined. SUBJECTS/METHODS: We included 439 subjects with validated information on body mass index (BMI) at 18 years of age. Using this information and current BMI, subjects were divided into never overweight/obese (BMI < 25 kg/m2), EOO and LOO. Adipocyte size, number, morphology (size in relation to body fat) and lipolysis were determined in subcutaneous abdominal WAT. Body composition and WAT distribution was assessed by dual-X-ray absorptiometry. RESULTS: Compared with never overweight/obese, EOO and LOO displayed larger WAT amounts in all examined depots, which in subcutaneous WAT was explained by a combination of increased size and number of fat cells in EOO and LOO. EOO had 40% larger subcutaneous fat mass than LOO (p < 0.0001). Visceral WAT mass, WAT morphology and lipolysis did not differ between EOO and LOO except for minor differences in men between the two obesity groups. On average, the increase in BMI per year was 57% higher in subjects with EOO compared to LOO (p < 0.0001). CONCLUSION: Early onset overweight/obesity causes a more rapid and pronounced accumulation of subcutaneous WAT than adult onset. However, fat mass expansion measures including WAT cellularity, morphology and fat cell lipolysis do not differ in an important way suggesting that similar mechanisms of WAT growth operate in EOO and LOO.

Myositis-specific autoantibodies and QTc changes by ECG in idiopathic inflammatory myopathies.

OBJECTIVES: The aim of this study was to investigate cardiac involvement detected by ECG in patients with idiopathic inflammatory myopathies (IIMs) and to evaluate possible associations between the autoantibody profile and ECG changes in these patients. METHODS: In a Scandinavian cross-sectional study, patients were included from two Danish centres and one Swedish centre. Resting 12-lead ECG was investigated in 261 patients with IIM compared with 102 patients with systemic sclerosis (SSc) and 48 healthy controls (HCs). ECG changes were correlated to clinical manifestations and myositis-specific and myositis-associated autoantibodies (MSAs and MAAs, respectively). RESULTS: Patients with IIM had a longer mean corrected QT (QTc) duration and more frequently presented with prolonged QTc (≥450 ms; P = 0.038) compared with HCs. A longer QTc duration was recorded in SSc compared with IIM [433 ms (s.d. 23) vs 426 (24); P = 0.011], yet there was no significant difference in the fraction with prolonged QTc (SSc: 22%, IIM: 16%; P = 0.19). In multivariable regression analyses, anti-Mi2 (P = 0.01, P = 0.035) and anti-Pl-7 (P = 0.045, P = 0.014) were associated with QTc duration and prolonged QTc in IIM. Elevated CRP was associated with prolonged QTc (P = 0.041). CONCLUSION: The presence of QTc abnormalities was as common in patients with IIM as in patients with SSc, including prolonged QTc seen in almost one-fifth of the patients. Anti-Mi2, anti-Pl-7 and elevated CRP may serve as biomarkers for cardiac disease in IIM, but needs to be confirmed in a larger prospective study.

Larger improvements in fatigue resistance and mitochondrial function with high- than with low-intensity contractions during interval training of mouse skeletal muscle.

Interval training (IT) results in improved fatigue resistance in skeletal muscle mainly due to an increased aerobic capacity, which involves increased muscle mitochondrial content and/or improved mitochondrial function. We hypothesized that IT with high-intensity contractions is more effective in increasing mitochondrial function, and hence fatigue resistance, than low-intensity contractions. To study this hypothesis without interference from differences in muscle fiber recruitment obliged to occur during voluntary contractions, IT was performed with in situ supramaximal electrical stimulation where all muscle fibers are recruited. We compared the effect of IT with repeated low-intensity (20 Hz stimulation, IT20) and high-intensity (100 Hz stimulation, IT100) contractions on fatigue resistance and mitochondrial content and function in mouse plantar flexor muscles. Muscles were stimulated every other day for 4 weeks. The averaged peak torque during IT bouts was 4.2-fold higher with IT100 than with IT20. Both stimulation protocols markedly improved in situ fatigue resistance, although the improvement was larger with IT100. The citrate synthase activity, a biomarker of mitochondrial content, was similarly increased with IT20 and IT100. Conversely, increased expression of mitochondrial respiratory chain (MRC) complexes I, III, and IV was only observed with IT100 and this was accompanied by increases in MRC supercomplex formation and pyruvate-malate-driven state 3 respiration in isolated mitochondria. In conclusion, the IT-induced increase in fatigue resistance is larger with high-intensity than with low-intensity contractions and this is linked to improved mitochondrial function due to increased expression of MRC complexes and assembly of MRC supercomplexes.

Mitochondrial NDUFA4L2 is a novel regulator of skeletal muscle mass and force.

The hypoxia-inducible nuclear-encoded mitochondrial protein NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 4-like 2 (NDUFA4L2) has been demonstrated to decrease oxidative phosphorylation and production of reactive oxygen species in neonatal cardiomyocytes, brain tissue and hypoxic domains of cancer cells. Prolonged local hypoxia can negatively affect skeletal muscle size and tissue oxidative capacity. Although skeletal muscle is a mitochondrial rich, oxygen sensitive tissue, the role of NDUFA4L2 in skeletal muscle has not previously been investigated. Here we ectopically expressed NDUFA4L2 in mouse skeletal muscles using adenovirus-mediated expression and in vivo electroporation. Moreover, femoral artery ligation (FAL) was used as a model of peripheral vascular disease to induce hind limb ischemia and muscle damage. Ectopic NDUFA4L2 expression resulted in reduced mitochondrial respiration and reactive oxygen species followed by lowered AMP, ADP, ATP, and NAD+ levels without affecting the overall protein content of the mitochondrial electron transport chain. Furthermore, ectopically expressed NDUFA4L2 caused a ~20% reduction in muscle mass that resulted in weaker muscles. The loss of muscle mass was associated with increased gene expression of atrogenes MurF1 and Mul1, and apoptotic genes caspase 3 and Bax. Finally, we showed that NDUFA4L2 was induced by FAL and that the Ndufa4l2 mRNA expression correlated with the reduced capacity of the muscle to generate force after the ischemic insult. These results show, for the first time, that mitochondrial NDUFA4L2 is a novel regulator of skeletal muscle mass and force. Specifically, induced NDUFA4L2 reduces mitochondrial activity leading to lower levels of important intramuscular metabolites, including adenine nucleotides and NAD+ , which are hallmarks of mitochondrial dysfunction and hence shows that dysfunctional mitochondrial activity may drive muscle wasting.

A longitudinal study of the antilipolytic effect of insulin in women following bariatric surgery.

Insulin resistance of glucose utilization is fully restored following BMI normalization after bariatric surgery. We investigated if this also pertains to insulin-induced effects on fatty acid handling. Forty-three women with obesity (OB) were investigated before and 2 years after Roux-en-Y gastric by-pass when BMI was <30 kg/m2 (PO) and compared with 26 never obese women (NO). The Adipo-IR index was used as measure of insulin antilipolytic sensitivity. Changes (delta) in circulating glycerol and fatty acid levels during hyperinsulinemic euglycemic clamp represented the insulin maximum antilipolytic effect. Overall fatty acid utilization was reflected by delta fatty acids minus 3 × delta glycerol. Adipo-IR was higher in OB than in NO and PO (p < 0.0001), the latter two groups having similar values. Insulin lowered glycerol levels by about 70% in all groups, but delta glycerol was 30% larger in PO than in NO (p = 0.04). Delta adds and adds utilization were similar in all groups. We conclude that women with obesity, whose BMI is normalized after bariatric surgery, have improved maximum in vivo antilipolytic effect of insulin above expected in absolute but not relative terms as regards glycerol changes, while the handling of circulating fatty acids is changed to the normal state.

FET family fusion oncoproteins target the SWI/SNF chromatin remodeling complex.

Members of the human FET family of RNA-binding proteins, comprising FUS, EWSR1, and TAF15, are ubiquitously expressed and engage at several levels of gene regulation. Many sarcomas and leukemias are characterized by the expression of fusion oncogenes with FET genes as 5' partners and alternative transcription factor-coding genes as 3' partners. Here, we report that the N terminus of normal FET proteins and their oncogenic fusion counterparts interact with the SWI/SNF chromatin remodeling complex. In contrast to normal FET proteins, increased fractions of FET oncoproteins bind SWI/SNF, indicating a deregulated and enhanced interaction in cancer. Forced expression of FET oncogenes caused changes of global H3K27 trimethylation levels, accompanied by altered gene expression patterns suggesting a shift in the antagonistic balance between SWI/SNF and repressive polycomb group complexes. Thus, deregulation of SWI/SNF activity could provide a unifying pathogenic mechanism for the large group of tumors caused by FET fusion oncoproteins. These results may help to develop common strategies for therapy.

Usefulness of surrogate markers to determine insulin action in fat cells.

BACKGROUND: Obesity is a major factor behind insulin resistance. The validity of simple biochemical surrogate measures to estimate insulin resistance at the fat cell level is unclear. OBJECTIVE: To investigate if the surrogate measures HOMA-IR (glucose/insulin product) and Adipo-IR (fatty acids/insulin product) reflect insulin action on glucose/lipid metabolism in fat cells. DESIGN: Insulin-induced lipogenesis and lipolysis inhibition (antilipolysis) in subcutaneous fat cells were investigated for sensitivity (reflecting receptor-near events) and responsiveness (i.e., maximum action reflecting distal post-receptor events) in 363 subjects. Results were compared with log10 transformed values for HOMA-IR and Adipo-IR. RESULTS: Individually, the four measures of in vitro insulin action on fat cells correlated significantly (p < 0.0001) but weakly with each other (adjusted r2 0.05-0.23). HOMA-IR and Adipo-IR correlated strongly with each other (adjusted r2 = 0.81). Using Spearman or simple linear regression all in vitro measures except antilipolytic responsiveness expressed per lipid weight, correlated significantly with Adipo-IR or HOMA-IR (p values <0.0001). Similar relationships remained after combined correction for age, body mass index and sex. Together, the four in vitro measures explained 50% of the variability in HOMA-IR and ADIPO-IR (p < 0.0001). Receiver-operating characteristic analysis showed good sensitivity and specificity for Adipo-IR and HOMA-IR to detect combined insulin resistance of antilipolysis and lipogenesis in fat cells (area under the curve = 0.8). CONCLUSIONS: Insulin action at the receptor and post-receptor levels on lipolysis and lipogenesis in fat cells correlates significantly with Adipo-IR and HOMA-IR. Both surrogate measures give similar information about insulin resistance of glucose and lipid metabolism in fat cells.

Prospective analyses of white adipose tissue gene expression in relation to long-term body weight changes.

BACKGROUND: Transcriptome analysis of abdominal subcutaneous white adipose tissue (sWAT) has identified important obesity-associated disturbances. However, the relation between sWAT transcriptome and long-term future changes in body weight remains elusive. OBJECTIVE: To investigate sWAT transcriptome signatures before and after long-term weight changes and assess their predictive value for body weight changes. DESIGN: A total of 56 women were followed longitudinally and subdivided into weight-stable (WS, n = 25), weight-gaining (WG, n = 14) and weight-losing (WL, n = 17) groups between baseline and follow-up (13 ± 1 years). The fasting sWAT transcriptome was analyzed by gene microarray at baseline and follow-up. Key genes associated with weight changes were validated using quantitative real-time PCR. RESULTS: In total 285 transcripts exhibited difference (FDR < 30%) in expression fold change over time between WL and WS women. WL women displayed decreased pro-inflammatory (NLRP3) but increased insulin-response gene (FASN and GLUT4) expression over time. In comparison, 461 transcripts displayed difference in expression fold change over time between WG and WS women (P < 0.05). Genes involved in autophagic processes (CDK5, SQSTM1 and FBXL2) were generally upregulated in WG women. At baseline, 307 and 302 transcripts were differentially expressed (FDR < 30%) in WL and WG women, respectively, when independently compared against WS women. Baseline expression of adipogenic and lipogenic genes (PPARG, IRS2 and HACD2) was lower, while pro-fibrotic (COL6A1) was higher, in WL than WS women; whereas protein processing genes were lower expressed in WG than in WS women. CONCLUSION: In adult women, long-term body weight change associates with altered sWAT transcriptome. Expression of genes associated with inflammation, insulin response, adipogenesis and lipogenesis are linked to weight loss. However, other pathways such as autophagy not only associate but also predict future weight gain suggesting that intrinsic factors in sWAT impact tissue expansion.

Head-to-head comparison of inorganic nitrate and metformin in a mouse model of cardiometabolic disease.

BACKGROUND/PURPOSE: Unhealthy dietary habits contribute to the increasing incidence of metabolic syndrome and type 2 diabetes (T2D), which is accompanied by oxidative stress, compromised nitric oxide (NO) bioavailability and increased cardiovascular risk. Apart from lifestyle changes, biguanides such as metformin are the first-line pharmacological treatment for T2D. Favourable cardiometabolic effects have been demonstrated following dietary nitrate supplementation to boost the nitrate-nitrite-NO pathway. Here we aim to compare the therapeutic value of inorganic nitrate and metformin alone and their combination in a model of cardiometabolic disease. EXPERIMENTAL APPROACH: Mice were fed control or high fat diet (HFD) for 7 weeks in combination with the NO synthase (NOS) inhibitor l-NAME to induce metabolic syndrome. Simultaneously, the mice were treated with vehicle, inorganic nitrate, metformin or a combination of nitrate and metformin in (drinking water). Cardiometabolic functions were assessed in vivo and tissues were collected/processed for analyses. KEY RESULTS: HFD + L-NAME was associated with cardiometabolic dysfunction, compared with controls, as evident from elevated blood pressure, endothelial dysfunction, impaired insulin sensitivity and compromised glucose clearance as well as liver steatosis. Both nitrate and metformin improved insulin/glucose homeostasis, whereas only nitrate had favourable effects on cardiovascular function and steatosis. Mechanistically, metformin and nitrate improved AMPK signalling, whereas only nitrate attenuated oxidative stress. Combination of nitrate and metformin reduced HbA1c and trended to further increase AMPK activation. CONCLUSION/IMPLICATIONS: Nitrate and metformin had equipotent metabolic effects, while nitrate was superior regarding protection against cardiovascular dysfunction and liver steatosis. If reproduced in future clinical trials, these findings may have implications for novel nutrition-based strategies against metabolic syndrome, T2D and associated complications.