Associations of circulating GDF15 with combined cognitive frailty and depression in older adults of the MARK-AGE study.

Growth differentiation factor-15 (GDF15) might be involved in the development of cognitive frailty and depression. Therefore, we evaluated cross-sectional associations of plasma GDF15 with combined cognitive-frailty-and-depression in older (i.e. ≥ 55 years) and younger adults of the MARK-AGE study. In the present work, samples and data of MARK-AGE ("European study to establish bioMARKers of human AGEing") participants (N = 2736) were analyzed. Cognitive frailty was determined by the global cognitive functioning score (GCF) and depression by the Self-Rating Depression Scale (SDS score). Adults were classified into three groups: (I) neither-cognitive-frailty-nor-depression, (II) either-cognitive-frailty-or-depression or (III) both-cognitive-frailty-and-depression. Cross-sectional associations were determined by unadjusted and by age, BMI, sex, comorbidities and hsCRP-adjusted linear and logistic regression analyses. Cognitive frailty, depression, age and GDF15 were significantly related within the whole study sample. High GDF15 levels were significantly associated with both-cognitive-frailty-and-depression (adjusted ß = 0.177 [0.044 - 0.310], p = 0.009), and with low GCF scores and high SDS scores. High GDF15 concentrations and quartiles were significantly associated with higher odds to have both-cognitive-frailty-and-depression (adjusted odds ratio = 2.353 [1.267 - 4.372], p = 0.007; and adjusted odds ratio = 1.414 [1.025 - 1.951], p = 0.035, respectively) independent of age, BMI, sex, comorbidities and hsCRP. These associations remained significant when evaluating older adults. We conclude that plasma GDF15 concentrations are significantly associated with combined cognitive-frailty-and-depression status and, with cognitive frailty and depressive symptoms separately in old as well as young community-dwelling adults.

Temporal dynamics of muscle mitochondrial uncoupling-induced integrated stress response and ferroptosis defense.

Mitochondria play multifaceted roles in cellular function, and impairments across domains of mitochondrial biology are known to promote cellular integrated stress response (ISR) pathways as well as systemic metabolic adaptations. However, the temporal dynamics of specific mitochondrial ISR related to physiological variations in tissue-specific energy demands remains unknown. Here, we conducted a comprehensive 24-hour muscle and plasma profiling of male and female mice with ectopic mitochondrial respiratory uncoupling in skeletal muscle (mUcp1-transgenic, TG). TG mice are characterized by increased muscle ISR, elevated oxidative stress defense, and increased secretion of FGF21 and GDF15 as ISR-induced myokines. We observed a temporal signature of both cell-autonomous and systemic ISR in the context of endocrine myokine signaling and cellular redox balance, but not of ferroptotic signature which was also increased in TG muscle. We show a progressive increase of muscle ISR on transcriptional level during the active phase (night time), with a subsequent peak in circulating FGF21 and GDF15 in the early resting phase. Moreover, we found highest levels of muscle oxidative defense (GPX and NQO1 activity) between the late active to early resting phase, which could aim to counteract excessive iron-dependent lipid peroxidation and ferroptosis in muscle of TG mice. These findings highlight the temporal dynamics of cell-autonomous and endocrine ISR signaling under skeletal muscle mitochondrial uncoupling, emphasizing the importance of considering such dissociation in translational strategies and sample collection for diagnostic biomarker analysis.

Heptadecanoic Acid Is Not a Key Mediator in the Prevention of Diet-Induced Hepatic Steatosis and Insulin Resistance in Mice.

Epidemiological studies found that the intake of dairy products is associated with an increased amount of circulating odd-chain fatty acids (OCFA, C15:0 and C17:0) in humans and further indicate that especially C17:0 is associated with a lower incidence of type 2 diabetes. However, causal relationships are not elucidated. To provide a mechanistic link, mice were fed high-fat (HF) diets supplemented with either milk fat or C17:0 for 20 weeks. Cultured primary mouse hepatocytes were used to distinguish differential effects mediated by C15:0 or C17:0. Despite an induction of OCFA after both dietary interventions, neither long-term milk fat intake nor C17:0 supplementation improved diet-induced hepatic lipid accumulation and insulin resistance in mice. HF feeding with milk fat actually deteriorates liver inflammation. Treatment of primary hepatocytes with C15:0 and C17:0 suppressed JAK2/STAT3 signaling, but only C15:0 enhanced insulin-stimulated phosphorylation of AKT. Overall, the data indicate that the intake of milk fat and C17:0 do not mediate health benefits, whereas C15:0 might be promising in further studies.

Decreased methylglyoxal-mediated protein glycation in the healthy aging mouse model of ectopic expression of UCP1 in skeletal muscle.

Mice with ectopic expression of uncoupling protein-1 (UCP1) in skeletal muscle exhibit a healthy aging phenotype with increased longevity and resistance to impaired metabolic health. This may be achieved by decreasing protein glycation by the reactive metabolite, methylglyoxal (MG). We investigated protein glycation and oxidative damage in skeletal muscle of mice with UCP1 expression under control of the human skeletal actin promoter (HSA-mUCP1) at age 12 weeks (young) and 70 weeks (aged). We found both young and aged HSA-mUCP1 mice had decreased advanced glycation endproducts (AGEs) formed from MG, lysine-derived Nε(1-carboxyethyl)lysine (CEL) and arginine-derived hydroimidazolone, MG-H1, whereas protein glycation by glucose forming Nε-fructosyl-lysine (FL) was increased ca. 2-fold, compared to wildtype controls. There were related increases in FL-linked AGEs, Nε-carboxymethyl-lysine (CML) and 3-deoxylglucosone-derived hydroimidazolone 3DG-H, and minor changes in protein oxidative and nitration adducts. In aged HSA-mUCP1 mice, urinary MG-derived AGEs/FL ratio was decreased ca. 60% whereas there was no change in CML/FL ratio - a marker of oxidative damage. This suggests that, normalized for glycemic status, aged HSA-mUCP1 mice had a lower flux of whole body MG-derived AGE exposure compared to wildtype controls. Proteomics analysis of skeletal muscle revealed a shift to increased heat shock proteins and mechanoprotection and repair in HSA-mUCP1 mice. Decreased MG-derived AGE protein content in skeletal muscle of aged HSA-mUCP1 mice is therefore likely produced by increased proteolysis of MG-modified proteins and increased proteostasis surveillance of the skeletal muscle proteome. From this and previous transcriptomic studies, signaling involved in enhanced removal of MG-modified protein is likely increased HSPB1-directed HUWE1 ubiquitination through eIF2α-mediated, ATF5-induced increased expression of HSPB1. Decreased whole body exposure to MG-derived AGEs may be linked to increased weight specific physical activity of HSA-mUCP1 mice. Decreased formation and increased clearance of MG-derived AGEs may be associated with healthy aging in the HSA-mUCP1 mouse.

The Effect of Dextrose or Protein Ingestion on Circulating Growth Differentiation Factor 15 and Appetite in Older Compared to Younger Women.

Growth differentiation factor 15 (GDF15) is a stress signal that can be induced by protein restriction and is associated with reduced food intake. Anorexia of aging, insufficient protein intake as well as high GDF15 concentrations often occur in older age, but it is unknown whether GDF15 concentrations change acutely after meal ingestion and affect appetite in older individuals. After an overnight fast, appetite was assessed in older (n = 20; 73.7 ± 6.30 years) and younger (n = 20; 25.7 ± 4.39 years) women with visual analogue scales, and concentrations of circulating GDF15 and glucagon-like peptide-1 (GLP-1) were quantified before and at 1, 2 and 4 h after ingestion of either dextrose (182 kcal) or a mixed protein-rich meal (450 kcal). In response to dextrose ingestion, appetite increased in both older and younger women, whereas GDF15 concentrations increased only in the older group. In older women, appetite response was negatively correlated with the GDF15 response (rho = -0.802, p = 0.005). Following high-protein ingestion, appetite increased in younger women, but remained low in the old, while GDF15 concentrations did not change significantly in either age group. GLP-1 concentrations did not differ between age groups or test meals. In summary, acute GDF15 response differed between older and younger women. Associations of postprandial appetite and GDF15 following dextrose ingestion in older women suggest a reduced appetite response when the GDF15 response is high, thus supporting the proposed anorectic effects of high GDF15 concentrations.

Mitochondrial stress-induced GFRAL signaling controls diurnal food intake and anxiety-like behavior.

Growth differentiation factor 15 (GDF15) is a mitochondrial stress-induced cytokine that modulates energy balance in an endocrine manner. However, the importance of its brainstem-restricted receptor GDNF family receptor alpha-like (GFRAL) to mediate endocrine GDF15 signaling to the brain upon mitochondrial dysfunction is still unknown. Using a mouse model with muscle-specific mitochondrial dysfunction, we here show that GFRAL is required for activation of systemic energy metabolism via daytime-restricted anorexia but not responsible for muscle wasting. We further find that muscle mitochondrial stress response involves a GFRAL-dependent induction of hypothalamic corticotropin-releasing hormone, without elevated corticosterone levels. Finally, we identify that GFRAL signaling governs an anxiety-like behavior in male mice with muscle mitochondrial dysfunction, with females showing a less robust GFRAL-dependent anxiety-like phenotype. Together, we here provide novel evidence of a mitochondrial stress-induced muscle-brain crosstalk via the GDF15-GFRAL axis to modulate food intake and anxiogenic behavior.

Seed- and leaf-based expression of FGF21-transferrin fusion proteins for oral delivery and treatment of non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis (NASH) is a global disease with no effective medication. The fibroblast growth factor 21 (FGF21) can reverse this liver dysfunction, but requires targeted delivery to the liver, which can be achieved via oral administration. Therefore, we fused FGF21 to transferrin (Tf) via a furin cleavage site (F), to promote uptake from the intestine into the portal vein, yielding FGF21-F-Tf, and established its production in both seeds and leaves of commercial Nicotiana tabacum cultivars, compared their expression profile and tested the bioavailability and bioactivity in feeding studies. Since biopharmaceuticals need to be produced in a contained environment, e.g., greenhouses in case of plants, the seed production was increased in this setting from 239 to 380 g m-2 a-1 seed mass with costs of 1.64 € g-1 by side branch induction, whereas leaves yielded 8,193 g m-2 a-1 leave mass at 0.19 € g-1. FGF21-F-Tf expression in transgenic seeds and leaves yielded 6.7 and 5.6 mg kg-1 intact fusion protein, but also 4.5 and 2.3 mg kg-1 additional Tf degradation products. Removing the furin site and introducing the liver-targeting peptide PLUS doubled accumulation of intact FGF21-transferrin fusion protein when transiently expressed in Nicotiana benthamiana from 0.8 to 1.6 mg kg-1, whereas truncation of transferrin (nTf338) and reversing the order of FGF21 and nTf338 increased the accumulation to 2.1 mg kg-1 and decreased the degradation products to 7% for nTf338-FGF21-PLUS. Application of partially purified nTf338-FGF21-PLUS to FGF21-/- mice by oral gavage proved its transfer from the intestine into the blood circulation and acutely affected hepatic mRNA expression. Hence, the medication of NASH via oral delivery of nTf338-FGF21-PLUS containing plants seems possible.

Detrimental effects of branched-chain amino acids in glucose tolerance can be attributed to valine induced glucotoxicity in skeletal muscle.

OBJECTIVE: Current data regarding the roles of branched-chain amino acids (BCAA) in metabolic health are rather conflicting, as positive and negative effects have been attributed to their intake. METHODS: To address this, individual effects of leucine and valine were elucidated in vivo (C57BL/6JRj mice) with a detailed phenotyping of these supplementations in high-fat (HF) diets and further characterization with in vitro approaches (C2C12 myocytes). RESULTS: Here, we demonstrate that under HF conditions, leucine mediates beneficial effects on adiposity and insulin sensitivity, in part due to increasing energy expenditure-likely contributing partially to the beneficial effects of a higher milk protein intake. On the other hand, valine feeding leads to a worsening of HF-induced health impairments, specifically reducing glucose tolerance/insulin sensitivity. These negative effects are driven by an accumulation of the valine-derived metabolite 3-hydroxyisobutyrate (3-HIB). Higher plasma 3-HIB levels increase basal skeletal muscle glucose uptake which drives glucotoxicity and impairs myocyte insulin signaling. CONCLUSION: These data demonstrate the detrimental role of valine in an HF context and elucidate additional targetable pathways in the etiology of BCAA-induced obesity and insulin resistance.

The Role of GDF15 as a Myomitokine.

Growth differentiation factor 15 (GDF15) is a cytokine best known for affecting systemic energy metabolism through its anorectic action. GDF15 expression and secretion from various organs and tissues is induced in different physiological and pathophysiological states, often linked to mitochondrial stress, leading to highly variable circulating GDF15 levels. In skeletal muscle and the heart, the basal expression of GDF15 is very low compared to other organs, but GDF15 expression and secretion can be induced in various stress conditions, such as intense exercise and acute myocardial infarction, respectively. GDF15 is thus considered as a myokine and cardiokine. GFRAL, the exclusive receptor for GDF15, is expressed in hindbrain neurons and activation of the GDF15-GFRAL pathway is linked to an increased sympathetic outflow and possibly an activation of the hypothalamic-pituitary-adrenal (HPA) stress axis. There is also evidence for peripheral, direct effects of GDF15 on adipose tissue lipolysis and possible autocrine cardiac effects. Metabolic and behavioral outcomes of GDF15 signaling can be beneficial or detrimental, likely depending on the magnitude and duration of the GDF15 signal. This is especially apparent for GDF15 production in muscle, which can be induced both by exercise and by muscle disease states such as sarcopenia and mitochondrial myopathy.

Effect of Microbial Status on Hepatic Odd-Chain Fatty Acids Is Diet-Dependent.

Odd-chain fatty acids (OCFA) are inversely associated with type-2-diabetes in epidemiological studies. They are considered as a biomarker for dairy intake because fermentation in ruminants yields high amounts of propionate, which is used as the primer for lipogenesis. Recently, we demonstrated endogenous OCFA synthesis from propionate in humans and mice, but how this is affected by microbial colonization is still unexplored. Here, we investigated the effect of increasing microbiota complexity on hepatic lipid metabolism and OCFA levels in different dietary settings. Germ-free (GF), gnotobiotic (SIH, simplified human microbiota) or conventional (CONV) C3H/HeOuJ-mice were fed a CHOW or high-fat diet with inulin (HFI) to induce microbial fermentation. We found that hepatic lipogenesis was increased with increasing microbiota complexity, independently of diet. In contrast, OCFA formation was affected by diet as well as microbiota. On CHOW, hepatic OCFA and intestinal gluconeogenesis decreased with increasing microbiota complexity (GF > SIH > CONV), while cecal propionate showed a negative correlation with hepatic OCFA. On HFI, OCFA levels were highest in SIH and positively correlated with cecal propionate. The propionate content in the CHOW diet was 10 times higher than that of HFI. We conclude that bacterial propionate production affects hepatic OCFA formation, unless this effect is masked by dietary propionate intake.

Postprandial dynamics and response of fibroblast growth factor 21 in older adults.

BACKGROUND & AIMS: Fibroblast growth factor 21 (FGF21) plays a pivotal role in glucose and lipid metabolism and has been proposed as a longevity hormone. However, elevated plasma FGF21 concentrations are paradoxically associated with mortality in higher age and little is known about the postprandial regulation of FGF21 in older adults. In this parallel group study, we investigated postprandial FGF21 dynamics and response in older (65-85 years) compared to younger (18-35 years) adults following test meals with varying macronutrient composition. METHODS: Participants (n = 60 older; n = 60 younger) were randomized to one of four test meals: dextrose, high carbohydrate (HC), high fat (HF) or high protein (HP). Blood was drawn before and 15, 30, 60, 120, 240 min after meal ingestion. Postprandial dynamics were evaluated using repeated measures ANCOVA. FGF21 response was assessed by incremental area under the curve. RESULTS: Fasting FGF21 concentrations were significantly higher in older adults. FGF21 dynamics were affected by test meal (p < 0.001) and age (p = 0.013), when adjusted for BMI and fasting FGF21. Postprandial FGF21 concentrations steadily declined over 240 min in both age groups after HF and HP, but not after dextrose or HC ingestion. At 240 min, FGF21 concentrations were significantly higher in older than in younger adults following dextrose (133 pg/mL, 95%CI: 103, 172 versus 91.2 pg/mL, 95%CI: 70.4, 118; p = 0.044), HC (109 pg/mL, 95%CI: 85.1, 141 versus 70.3 pg/mL, 95%CI: 55.2, 89.6; p = 0.014) and HP ingestion (45.4 pg/mL, 95%CI: 34.4, 59.9 versus 27.9 pg/mL 95%CI: 20.9, 37.1; p = 0.018). FGF21 dynamics and response to HF were similar for both age groups. CONCLUSIONS: The age-specific differences in postprandial FGF21 dynamics and response in healthy adults, potentially explain higher FGF21 concentrations in older age. Furthermore, there appears to be a significant impact of acute and recent protein intake on FGF21 secretion.

Pseudo-Starvation Driven Energy Expenditure Negatively Affects Ovarian Follicle Development.

In the present investigation, we examined whether a change in whole body energy fluxes could affect ovarian follicular development, employing mice ectopically expressing uncoupling protein 1 in skeletal muscle (UCP1-TG). Female UCP1-TG and wild-type (WT) mice were dissected at the age of 12 weeks. Energy intake and expenditure, activity, body weight and length, and body composition were measured. Plasma insulin, glucose, leptin, plasma fibroblast growth factor 21 (FGF21) and plasma insulin-like growth factor 1 (IGF1) levels were analyzed and ovarian follicle and corpus luteum numbers were counted. IGF1 signaling was analyzed by immunohistochemical staining for the activation of insulin receptor substrate 1/2 (IRS1/2) and AKT. UCP1-TG female mice had increased energy expenditure, reduced body size, maintained adiposity, and decreased IGF1 concentrations compared to their WT littermates, while preantral and antral follicle numbers were reduced by 40% and 60%, respectively. Corpora lutea were absent in 40% of the ovaries of UCP1-TG mice. Phospho-IRS1, phospho-AKT -Ser473 and -Thr308 immunostaining was present in the granulosa cells of antral follicles in WT ovaries, but faint to absent in the antral follicles of UCP1-TG mice. In conclusion, the reduction in circulating IGF1 levels due to the ectopic expression of UCP1 is associated with reduced immunostaining of the IRS1-PI3/AKT pathway, which may negatively affect ovarian follicle development and ovulation.

Regulation of diurnal energy balance by mitokines.

The mammalian system of energy balance regulation is intrinsically rhythmic with diurnal oscillations of behavioral and metabolic traits according to the 24 h day/night cycle, driven by cellular circadian clocks and synchronized by environmental or internal cues such as metabolites and hormones associated with feeding rhythms. Mitochondria are crucial organelles for cellular energy generation and their biology is largely under the control of the circadian system. Whether mitochondrial status might also feed-back on the circadian system, possibly via mitokines that are induced by mitochondrial stress as endocrine-acting molecules, remains poorly understood. Here, we describe our current understanding of the diurnal regulation of systemic energy balance, with focus on fibroblast growth factor 21 (FGF21) and growth differentiation factor 15 (GDF15), two well-known endocrine-acting metabolic mediators. FGF21 shows a diurnal oscillation and directly affects the output of the brain master clock. Moreover, recent data demonstrated that mitochondrial stress-induced GDF15 promotes a day-time restricted anorexia and systemic metabolic remodeling as shown in UCP1-transgenic mice, where both FGF21 and GDF15 are induced as myomitokines. In this mouse model of slightly uncoupled skeletal muscle mitochondria GDF15 proved responsible for an increased metabolic flexibility and a number of beneficial metabolic adaptations. However, the molecular mechanisms underlying energy balance regulation by mitokines are just starting to emerge, and more data on diurnal patterns in mouse and man are required. This will open new perspectives into the diurnal nature of mitokines and action both in health and disease.

Associations Between Serum GDF15 Concentrations, Muscle Mass, and Strength Show Sex-Specific Differences in Older Hospital Patients.

Aging is accompanied by a progressive decline of muscle mass and strength and also higher levels of circulating cytokines such as growth differentiation factor 15 (GDF15). Studies evaluating the association of GDF15 with muscle mass and strength are rare. In this analysis, we investigated GDF15 concentrations and their relationship with muscle mass and strength in older men compared with women. GDF15 serum concentrations were measured in 103 (60 years and older) hospital patients and an age-matched control group with an immunosorbent assay. Skeletal muscle mass was determined with the bioelectrical impedance analysis. Grip strength and knee extension strength were assessed and normalized for height. Associations between GDF15 concentrations and muscle mass and strength were evaluated with general linear models. Male patients showed higher levels of GDF15 compared with female patients (p = 0.021). Elevated GDF15 concentrations were associated with lower measures of muscle mass, exclusively in men, after adjustment for age and number of drugs per day. Our results indicate sex differences between associations of GDF15 with muscle mass and strength parameters in a cohort of older hospital patients.

Age-related fatigue is associated with reduced mitochondrial function in peripheral blood mononuclear cells.

BACKGROUND: Fatigue is a complex syndrome associated with exhaustion not relieved by sleep. It occurs frequently in older adults in the context of chronic disease and is associated with decreased physical capacity. Whether a mitochondrial dysfunction and therefore an impaired energy production might contribute to the development of fatigue during aging is yet unknown. The aim of this study was to evaluate mitochondrial respiration of peripheral blood mononuclear cells (PBMCs) in older patients with and without fatigue. METHOD: Fatigue was determined according to the Brief Fatigue Inventory. Mitochondrial respiration of freshly isolated PBMCs was investigated by high-resolution respirometry using the Oroboros Oxygraph-O2k. Functional impairment and depressive symptoms were assessed using questionnaires. RESULTS: 23 geriatric patients (77.8 ± 4.9 years; 43.5% female) with fatigue and 22 without fatigue (75.4 ± 5.4 years; 45.5% female) were analyzed. Patients with fatigue exhibited more functional limitations and more depressive symptoms. High-resolution respirometry of intact PBMCs revealed a lower routine (4.82 ± 1.14 pmol/s versus 5.89 ± 1.90 pmol/s, p = 0.041) and maximum (6.55 ± 1.51 pmol/s versus 8.43 ± 3.67 pmol/s, p = 0.013) oxygen consumption rate, resulting in a reduced ATP-linked respiration (4.26 ± 1.00 pmol/s versus 5.09 ± 1.53 pmol/s, p = 0.035) of PBMCs from geriatric patients with fatigue compared to controls without. CONCLUSIONS: This short report shows that in this group of older patients, fatigue is associated with lower PBMC mitochondrial respiration. Whether the impaired mitochondrial respiration is accompanied by a reduced mitochondrial activity in other organs (e.g. muscle) remains to be elucidated.

N-Acetylcysteine as Modulator of the Essential Trace Elements Copper and Zinc.

N-acetylcysteine (NAC) is a frequently prescribed drug and known for its metal chelating capability. However, to date it is not well characterized whether NAC intake affects the homeostasis of essential trace elements. As a precursor of glutathione (GSH), NAC also has the potential to modulate the cellular redox homeostasis. Thus, we aimed to analyze effects of acute and chronic NAC treatment on the homeostasis of copper (Cu) and zinc (Zn) and on the activity of the redox-sensitive transcription factor Nrf2. Cells were exposed to 1 mM NAC and were co-treated with 50 µM Cu or Zn. We showed that NAC treatment reduced the cellular concentration of Zn and Cu. In addition, NAC inhibited the Zn-induced Nrf2 activation and limited the concomitant upregulation of cellular GSH concentrations. In contrast, mice chronically received NAC via drinking water (1 g NAC/100 mL). Cu and Zn concentrations were decreased in liver and spleen. In the duodenum, NQO1, TXNRD, and SOD activities were upregulated by NAC. All of them can be induced by Nrf2, thus indicating a putative Nrf2 activation. Overall, NAC modulates the homeostasis of Cu and Zn both in vitro and in vivo and accordingly affects the cellular redox balance.

Cell autonomous requirement of neurofibromin (Nf1) for postnatal muscle hypertrophic growth and metabolic homeostasis.

BACKGROUND: Neurofibromatosis type 1 (NF1) is a multi-organ disease caused by mutations in neurofibromin 1 (NF1). Amongst other features, NF1 patients frequently show reduced muscle mass and strength, impairing patients' mobility and increasing the risk of fall. The role of Nf1 in muscle and the cause for the NF1-associated myopathy are mostly unknown. METHODS: To dissect the function of Nf1 in muscle, we created muscle-specific knockout mouse models for NF1, inactivating Nf1 in the prenatal myogenic lineage either under the Lbx1 promoter or under the Myf5 promoter. Mice were analysed during prenatal and postnatal myogenesis and muscle growth. RESULTS: Nf1Lbx1 and Nf1Myf5 animals showed only mild defects in prenatal myogenesis. Nf1Lbx1 animals were perinatally lethal, while Nf1Myf5 animals survived only up to approximately 25 weeks. A comprehensive phenotypic characterization of Nf1Myf5 animals showed decreased postnatal growth, reduced muscle size, and fast fibre atrophy. Proteome and transcriptome analyses of muscle tissue indicated decreased protein synthesis and increased proteasomal degradation, and decreased glycolytic and increased oxidative activity in muscle tissue. High-resolution respirometry confirmed enhanced oxidative metabolism in Nf1Myf5 muscles, which was concomitant to a fibre type shift from type 2B to type 2A and type 1. Moreover, Nf1Myf5 muscles showed hallmarks of decreased activation of mTORC1 and increased expression of atrogenes. Remarkably, loss of Nf1 promoted a robust activation of AMPK with a gene expression profile indicative of increased fatty acid catabolism. Additionally, we observed a strong induction of genes encoding catabolic cytokines in muscle Nf1Myf5 animals, in line with a drastic reduction of white, but not brown adipose tissue. CONCLUSIONS: Our results demonstrate a cell autonomous role for Nf1 in myogenic cells during postnatal muscle growth required for metabolic and proteostatic homeostasis. Furthermore, Nf1 deficiency in muscle drives cross-tissue communication and mobilization of lipid reserves.

High-protein diet more effectively reduces hepatic fat than low-protein diet despite lower autophagy and FGF21 levels.

BACKGROUND AND AIMS: Non-alcoholic fatty liver disease (NAFLD) is becoming increasingly prevalent and nutrition intervention remains the most important therapeutic approach for NAFLD. Our aim was to investigate whether low- (LP) or high-protein (HP) diets are more effective in reducing liver fat and reversing NAFLD and which mechanisms are involved. METHODS: 19 participants with morbid obesity undergoing bariatric surgery were randomized into two hypocaloric (1500-1600 kcal/day) diet groups, a low protein (10E% protein) and a high protein (30E% protein), for three weeks prior to surgery. Intrahepatic lipid levels (IHL) and serum fibroblast growth factor 21 (FGF21) were measured before and after the dietary intervention. Autophagy flux, histology, mitochondrial activity and gene expression analyses were performed in liver samples collected during surgery. RESULTS: IHL levels decreased by 42.6% in the HP group, but were not significantly changed in the LP group despite similar weight loss. Hepatic autophagy flux and serum FGF21 increased by 66.7% and 42.2%, respectively, after 3 weeks in the LP group only. Expression levels of fat uptake and lipid biosynthesis genes were lower in the HP group compared with those in the LP group. RNA-seq analysis revealed lower activity of inflammatory pathways upon HP diet. Hepatic mitochondrial activity and expression of ß-oxidation genes did not increase in the HP group. CONCLUSIONS: HP diet more effectively reduces hepatic fat than LP diet despite of lower autophagy and FGF21. Our data suggest that liver fat reduction upon HP diets result primarily from suppression of fat uptake and lipid biosynthesis.

The branched-chain amino acids valine and leucine have differential effects on hepatic lipid metabolism.

Dairy intake, as a source of branched-chain amino acids (BCAA), has been linked to a lower incidence of type-2-diabetes and increased circulating odd-chain fatty acids (OCFA). To understand this connection, we aimed to investigate differences in BCAA metabolism of leucine and valine, a possible source of OCFA, and their role in hepatic metabolism. Male mice were fed a high-fat diet supplemented with leucine and valine for 1 week and phenotypically characterized with a focus on lipid metabolism. Mouse primary hepatocytes were treated with the BCAA or a Pparα activator WY-14643 to systematically examine direct hepatic effects and their mechanisms. Here, we show that only valine supplementation was able to increase hepatic and circulating OCFA levels via two pathways; a PPARα-dependent induction of α-oxidation and an increased supply of propionyl-CoA for de novo lipogenesis. Meanwhile, we were able to confirm leucine-mediated effects on the inhibition of food intake and transport of fatty acids, as well as induction of S6 ribosomal protein phosphorylation. Taken together, these data illustrate differential roles of the BCAA in lipid metabolism and provide preliminary evidence that exclusively valine contributes to the endogenous formation of OCFA which is important for a better understanding of these metabolites in metabolic health.

Muscle-derived GDF15 drives diurnal anorexia and systemic metabolic remodeling during mitochondrial stress.

Mitochondrial dysfunction promotes metabolic stress responses in a cell-autonomous as well as organismal manner. The wasting hormone growth differentiation factor 15 (GDF15) is recognized as a biomarker of mitochondrial disorders, but its pathophysiological function remains elusive. To test the hypothesis that GDF15 is fundamental to the metabolic stress response during mitochondrial dysfunction, we investigated transgenic mice (Ucp1-TG) with compromised muscle-specific mitochondrial OXPHOS capacity via respiratory uncoupling. Ucp1-TG mice show a skeletal muscle-specific induction and diurnal variation of GDF15 as a myokine. Remarkably, genetic loss of GDF15 in Ucp1-TG mice does not affect muscle wasting or transcriptional cell-autonomous stress response but promotes a progressive increase in body fat mass. Furthermore, muscle mitochondrial stress-induced systemic metabolic flexibility, insulin sensitivity, and white adipose tissue browning are fully abolished in the absence of GDF15. Mechanistically, we uncovered a GDF15-dependent daytime-restricted anorexia, whereas GDF15 is unable to suppress food intake at night. Altogether, our evidence suggests a novel diurnal action and key pathophysiological role of mitochondrial stress-induced GDF15 in the regulation of systemic energy metabolism.