Prolonged FGF21 treatment increases energy expenditure and induces weight loss in obese mice independently of UCP1 and adrenergic signaling.

Fibroblast growth factor 21 (FGF21) reduces body weight, which was attributed to induced energy expenditure (EE). Conflicting data have been published on the role of uncoupling protein 1 (UCP1) in this effect. Therefore, we aimed to revisit the thermoregulatory effects of FGF21 and their implications for body weight regulation. We found that an 8-day treatment with FGF21 lowers body weight to similar extent in both wildtype (WT) and UCP1-deficient (KO) mice fed high-fat diet. In WT mice, this effect is solely due to increased EE, associated with a strong activation of UCP1 and with excess heat dissipated through the tail. This thermogenesis takes place in the interscapular region and can be attenuated by a ß-adrenergic inhibitor propranolol. In KO mice, FGF21-induced weight loss correlates with a modest increase in EE, which is independent of adrenergic signaling, and with a reduced energy intake. Interestingly, the gene expression profile of interscapular brown adipose tissue (but not subcutaneous white adipose tissue) of KO mice is massively affected by FGF21, as shown by increased expression of genes encoding triacylglycerol/free fatty acid cycle enzymes. Thus, FGF21 elicits central thermogenic and pyretic effects followed by a concomitant increase in EE and body temperature, respectively. The associated weight loss is strongly dependent on UCP1-based thermogenesis. However, in the absence of UCP1, alternative mechanisms of energy dissipation may contribute, possibly based on futile triacylglycerol/free fatty acid cycling in brown adipose tissue and reduced food intake.

Study protocol: Identification and validation of integrative biomarkers of physical activity level and health in children and adolescents (INTEGRActiv).

Background: Physical activity (PA) provides health benefits across the lifespan and improves many established cardiovascular risk factors that have a significant impact on overall mortality. However, discrepancies between self-reported and device-based measures of PA make it difficult to obtain consistent results regarding PA and its health effects. Moreover, PA may produce different health effects depending on the type, intensity, duration, and frequency of activities and individual factors such as age, sex, body weight, early life conditions/exposures, etc. Appropriate biomarkers relating the degree of PA level with its effects on health, especially in children and adolescents, are required and missing. The main objective of the INTEGRActiv study is to identify novel useful integrative biomarkers of PA and its effects on the body health in children and adolescents, who represent an important target population to address personalized interventions to improve future metabolic health. Methods/design: The study is structured in two phases. First, biomarkers of PA and health will be identified at baseline in a core cohort of 180 volunteers, distributed into two age groups: prepubertal (n = 90), and postpubertal adolescents (n = 90). Each group will include three subgroups (n = 30) with subjects of normal weight, overweight, and obesity, respectively. Identification of new biomarkers will be achieved by combining physical measures (PA and cardiorespiratory and muscular fitness, anthropometry) and molecular measures (cardiovascular risk factors, endocrine markers, cytokines and circulating miRNA in plasma, gene expression profile in blood cells, and metabolomics profiling in plasma). In the second phase, an educational intervention and its follow-up will be carried out in a subgroup of these subjects (60 volunteers), as a first validation step of the identified biomarkers. Discussion: The INTEGRActiv study is expected to provide the definition of PA and health-related biomarkers (PA-health biomarkers) in childhood and adolescence. It will allow us to relate biomarkers to factors such as age, sex, body weight, sleep behavior, dietary factors, and pubertal status and to identify how these factors quantitatively affect the biomarkers' responses. Taken together, the INTEGRActiv study approach is expected to help monitor the efficacy of interventions aimed to improve the quality of life of children/adolescents through physical activity. Clinical Trial Registration: ClinicalTrials.gov, Identifier NCT05907785.

Highly recruited brown adipose tissue does not in itself protect against obesity.

OBJECTIVE: The possibility to counteract the development of obesity in humans by recruiting brown or brite/beige adipose tissue (and thus UCP1) has attracted much attention. Here we examine if a diet that can activate diet-induced thermogenesis can exploit pre-enhanced amounts of UCP1 to counteract the development of diet-induced obesity. METHODS: To investigate the anti-obesity significance of highly augmented amounts of UCP1 for control of body energy reserves, we physiologically increased total UCP1 amounts by recruitment of brown and brite/beige tissues in mice. We then examined the influence of the augmented UCP1 levels on metabolic parameters when the mice were exposed to a high-fat/high-sucrose diet under thermoneutral conditions. RESULTS: The total UCP1 levels achieved were about 50-fold higher in recruited than in non-recruited mice. Contrary to underlying expectations, in the mice with highly recruited UCP1 and exposed to a high-fat/high-sucrose diet the thermogenic capacity of this UCP1 was completely inactivate. The mice even transiently (in an adipostat-like manner) demonstrated a higher metabolic efficiency and fat gain than did non-recruited mice. This was accomplished without altering energy expenditure or food absorption efficiency. The metabolic efficiency here was indistinguishable from that of mice totally devoid of UCP1. CONCLUSIONS: Although UCP1 protein may be available, it is not inevitably utilized for diet-induced thermogenesis. Thus, although attempts to recruit UCP1 in humans may become successful as such, it is only if constant activation of the UCP1 is also achieved that amelioration of obesity development could be attained.

Impairment of adrenergically-regulated thermogenesis in brown fat of obesity-resistant mice is compensated by non-shivering thermogenesis in skeletal muscle.

OBJECTIVE: Non-shivering thermogenesis (NST) mediated by uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) can be activated via the adrenergic system in response to cold or diet, contributing to both thermal and energy homeostasis. Other mechanisms, including metabolism of skeletal muscle, may also be involved in NST. However, relative contribution of these energy dissipating pathways and their adaptability remain a matter of long-standing controversy. METHODS: We used warm-acclimated (30 °C) mice to characterize the effect of an up to 7-day cold acclimation (6 °C; CA) on thermoregulatory thermogenesis, comparing inbred mice with a genetic background conferring resistance (A/J) or susceptibility (C57BL/6 J) to obesity. RESULTS: Both warm-acclimated C57BL/6 J and A/J mice exhibited similar cold endurance, assessed as a capability to maintain core body temperature during acute exposure to cold, which improved in response to CA, resulting in comparable cold endurance and similar induction of UCP1 protein in BAT of mice of both genotypes. Despite this, adrenergic NST in BAT was induced only in C57BL/6 J, not in A/J mice subjected to CA. Cold tolerance phenotype of A/J mice subjected to CA was not based on increased shivering, improved insulation, or changes in physical activity. On the contrary, lipidomic, proteomic and gene expression analyses along with palmitoyl carnitine oxidation and cytochrome c oxidase activity revealed induction of lipid oxidation exclusively in skeletal muscle of A/J mice subjected to CA. These changes appear to be related to skeletal muscle NST, mediated by sarcolipin-induced uncoupling of sarco(endo)plasmic reticulum calcium ATPase pump activity and accentuated by changes in mitochondrial respiratory chain supercomplexes assembly. CONCLUSIONS: Our results suggest that NST in skeletal muscle could be adaptively augmented in the face of insufficient adrenergic NST in BAT, depending on the genetic background of the mice. It may provide both protection from cold and resistance to obesity, more effectively than BAT.

Loss of UCP1 function augments recruitment of futile lipid cycling for thermogenesis in murine brown fat.

OBJECTIVE: Classical ATP-independent non-shivering thermogenesis enabled by uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) is activated, but not essential for survival, in the cold. It has long been suspected that futile ATP-consuming substrate cycles also contribute to thermogenesis and can partially compensate for the genetic ablation of UCP1 in mouse models. Futile ATP-dependent thermogenesis could thereby enable survival in the cold even when brown fat is less abundant or missing. METHODS: In this study, we explore different potential sources of UCP1-independent thermogenesis and identify a futile ATP-consuming triglyceride/fatty acid cycle as the main contributor to cellular heat production in brown adipocytes lacking UCP1. We uncover the mechanism on a molecular level and pinpoint the key enzymes involved using pharmacological and genetic interference. RESULTS: ATGL is the most important lipase in terms of releasing fatty acids from lipid droplets, while DGAT1 accounts for the majority of fatty acid re-esterification in UCP1-ablated brown adipocytes. Furthermore, we demonstrate that chronic cold exposure causes a pronounced remodeling of adipose tissues and leads to the recruitment of lipid cycling capacity specifically in BAT of UCP1-knockout mice, possibly fueled by fatty acids from white fat. Quantification of triglyceride/fatty acid cycling clearly shows that UCP1-ablated animals significantly increase turnover rates at room temperature and below. CONCLUSION: Our results suggest an important role for futile lipid cycling in adaptive thermogenesis and total energy expenditure.

A pyrexic effect of FGF21 independent of energy expenditure and UCP1.

OBJECTIVE: Administration of FGF21 to mice reduces body weight and increases body temperature. The increase in body temperature is generally interpreted as hyperthermia, i.e. a condition secondary to the increase in energy expenditure (heat production). Here, we examine an alternative hypothesis: that FGF21 has a direct pyrexic effect, i.e. FGF21 increases body temperature independently of any effect on energy expenditure. METHODS: We studied the effects of FGF21 treatment on body temperature and energy expenditure in high-fat-diet-fed and chow-fed mice exposed acutely to various ambient temperatures, in high-fat diet-fed mice housed at 30 °C (i.e. at thermoneutrality), and in mice lacking uncoupling protein 1 (UCP1). RESULTS: In every model studied, FGF21 increased body temperature, but energy expenditure was increased only in some models. The effect of FGF21 on body temperature was more (not less, as expected in hyperthermia) pronounced at lower ambient temperatures. Effects on body temperature and energy expenditure were temporally distinct (daytime versus nighttime). FGF21 enhanced UCP1 protein content in brown adipose tissue (BAT); there was no measurable UCP1 protein in inguinal brite/beige adipose tissue. FGF21 increased energy expenditure through adrenergic stimulation of BAT. In mice lacking UCP1, FGF21 did not increase energy expenditure but increased body temperature by reducing heat loss, e.g. a reduced tail surface temperature. CONCLUSION: The effect of FGF21 on body temperature is independent of UCP1 and can be achieved in the absence of any change in energy expenditure. Since elevated body temperature is a primary effect of FGF21 and can be achieved without increasing energy expenditure, only limited body weight-lowering effects of FGF21 may be expected.

Additive Effects of Omega-3 Fatty Acids and Thiazolidinediones in Mice Fed a High-Fat Diet: Triacylglycerol/Fatty Acid Cycling in Adipose Tissue.

Long-chain n-3 polyunsaturated fatty acids (Omega-3) and anti-diabetic drugs thiazolidinediones (TZDs) exhibit additive effects in counteraction of dietary obesity and associated metabolic dysfunctions in mice. The underlying mechanisms need to be clarified. Here, we aimed to learn whether the futile cycle based on the hydrolysis of triacylglycerol and re-esterification of fatty acids (TAG/FA cycling) in white adipose tissue (WAT) could be involved. We compared Omega-3 (30 mg/g diet) and two different TZDs-pioglitazone (50 mg/g diet) and a second-generation TZD, MSDC-0602K (330 mg/g diet)-regarding their effects in C57BL/6N mice fed an obesogenic high-fat (HF) diet for 8 weeks. The diet was supplemented or not by the tested compound alone or with the two TZDs combined individually with Omega-3. Activity of TAG/FA cycle in WAT was suppressed by the obesogenic HF diet. Additive effects in partial rescue of TAG/FA cycling in WAT were observed with both combined interventions, with a stronger effect of Omega-3 and MSDC-0602K. Our results (i) supported the role of TAG/FA cycling in WAT in the beneficial additive effects of Omega-3 and TZDs on metabolism of diet-induced obese mice, and (ii) showed differential modulation of WAT gene expression and metabolism by the two TZDs, depending also on Omega-3.

UCP1-independent glucose-lowering effect of leptin in type 1 diabetes: only in conditions of hypoleptinemia.

The possibility to use leptin therapeutically for lowering glucose levels in patients with type 1 diabetes has attracted interest. However, earlier animal models of type 1 diabetes are severely catabolic with very low endogenous leptin levels, unlike most patients with diabetes. Here, we aim to test glucose-lowering effects of leptin in novel, more human-like murine models. We examined the glucose-lowering potential of leptin in diabetic models of two types: streptozotocin-treated mice and mice treated with the insulin receptor antagonist S961. To prevent hypoleptinemia, we used combinations of thermoneutral temperature and high-fat feeding. Leptin fully normalized hyperglycemia in standard chow-fed streptozotocin-treated diabetic mice. However, more humanized physiological conditions (high-fat diets or thermoneutral temperatures) that increased adiposity - and thus also leptin levels - in the diabetic mice abrogated the effects of leptin, i.e., the mice developed leptin resistance also in this respect. The glucose-lowering effect of leptin was not dependent on the presence of the uncoupling protein-1 and was not associated with alterations in plasma insulin, insulin-like growth factor 1, food intake or corticosterone but fully correlated with decreased plasma glucagon levels and gluconeogenesis. An important implication of these observations is that the therapeutic potential of leptin as an additional treatment in patients with type 1 diabetes is probably limited. This is because such patients are treated with insulin and do not display low leptin levels. Thus, the potential for a glucose-lowering effect of leptin would already have been attained with standard insulin therapy, and further effects on blood glucose level through additional leptin cannot be anticipated.