Neonatal Brown Adipose Tissue Research Articles

Meteorin-like levels are associated with active brown adipose tissue in early infancy.

Meteorin-like (METRNL) is a hormonal factor released by several tissues, including thermogenically active brown and beige adipose tissues. It exerts multiple beneficial effects on metabolic and cardiovascular systems in experimental models. However, the potential role of METRNL as brown adipokine in humans has not been investigated previously, particularly in relation to the metabolic adaptations taking place in early life, when brown adipose tissue (BAT) is particularly abundant. METRNL levels, as well as body composition (DXA) and circulating endocrine-metabolic variables, were assessed longitudinally in a cohort of infants at birth, and at ages 4 and 12 months. BAT activity was measured by infrared thermography at age 12 months. METRNL levels were also determined cross-sectionally in adults; METRNL gene expression (qRT-PCR) was assessed in BAT and liver samples from neonates, and in adipose tissue and liver samples form adults. Simpson-Golabi-Behmel Syndrome (SGBS) adipose cells were thermogenically activated using cAMP, and METRNL gene expression and METRNL protein released were analysed. Serum METRNL levels were high at birth and declined across the first year of life albeit remaining higher than in adulthood. At age 4 and 12 months, METRNL levels correlated positively with circulating C-X-C motif chemokine ligand 14 (CXCL14), a chemokine released by thermogenically active BAT, but not with parameters of adiposity or metabolic status. METRNL levels also correlated positively with infrared thermography-estimated posterior-cervical BAT activity in girls aged 12 months. Gene expression analysis indicated high levels of METRNL mRNA in neonatal BAT. Thermogenic stimulus of brown/beige adipocytes led to a significant increase of METRNL gene expression and METRN protein release to the cell culture medium. Circulating METRNL levels are high in the first year of life and correlate with indices of BAT activity and with levels of an established brown adipokine such as CXCL14. These data, in addition with the high expression of METRNL in neonatal BAT and in thermogenically-stimulated brown/beige adipocytes, suggest that METRNL is actively secreted by BAT and may be a circulating biomarker of BAT activity in early life.

BMP8 and activated brown adipose tissue in human newborns

The classical dogma states that brown adipose tissue (BAT) plays a major role in the regulation of temperature in neonates. However, although BAT has been studied in infants for more than a century, the knowledge about its physiological features at this stage of life is rather limited. This has been mainly due to the lack of appropriate investigation methods, ethically suitable for neonates. Here, we have applied non-invasive infrared thermography (IRT) to investigate neonatal BAT activity. Our data show that BAT temperature correlates with body temperature and that mild cold stimulus promotes BAT activation in newborns. Notably, a single short-term cold stimulus during the first day of life improves the body temperature adaption to a subsequent cold event. Finally, we identify that bone morphogenic protein 8B (BMP8B) is associated with the BAT thermogenic response in neonates. Overall, our data uncover key features of the setup of BAT thermogenesis in newborns.

Embryonic exposure to hyper glucocorticoids suppresses brown fat development and thermogenesis via REDD1

Maternal stress during pregnancy is prevailing worldwide, which exposes fetuses to intrauterine hyper glucocorticoids (GC), programming offspring to obesity and metabolic diseases. Despite the importance of brown adipose tissue (BAT) in maintaining long-term metabolic health, impacts of prenatal hyper GC on postnatal BAT thermogenesis and underlying regulations remain poorly defined. Pregnant mice were administrated with synthetic GC dexamethasone (DEX) at levels comparable to fetal GC exposure of stressed mothers. Prenatal GC exposure dose-dependently reduced BAT thermogenic activity, contributing to lower body temperature and higher mortality of neonates; such difference was abolished under thermoneutrality, underscoring BAT deficiency was the major contributor to adverse changes in postnatal thermogenesis due to excessive GC. Prenatal GC exposure highly activated Redd1 expression and reduced Ppargc1a transcription from the alternative promoter (Ppargc1a-AP) in neonatal BAT. During brown adipocyte differentiation, ectopic Redd1 expression reduced Ppargc1a-AP expression and mitochondrial biogenesis; and the inhibitory effects of GC on mitochondrial biogenesis and Ppargc1a-AP expression were blocked by Redd1 ablation. Redd1 reduced protein kinase A phosphorylation and suppressed cyclic adenosine monophosphate (cAMP) -responsive element-binding protein (CREB) binding to the cAMP regulatory element (CRE) in Ppargc1a-AP promoter, leading to Ppargc1a-AP inactivation. In summary, excessive maternal GC exposure during pregnancy dysregulates Redd1-Ppargc1a-AP axis, which impairs fetal BAT development, hampering postnatal thermogenic adaptation and metabolic health of offspring.

1710-P: Maternal Supplementation of Tetrahydrobiopterin Regulates Differentiation of Fetal Brown Adipose Tissue and Contributes to Offspring Metabolic Health

Brown adipose tissue (BAT) is a key organ that produces heat and dissipates energy, and may be clinically relevant to the treatment of obesity and diabetes. Tetrahydrobiopterin (BH4) is an essential co-factor of tyrosine hydroxylase, a rate-limiting enzyme of catecholamine biosynthesis including that of norepinephrine (NA) and nitric oxide (NO) synthase. Although both NA and NO are well-known factors in BAT differentiation, the role of BH4 in the differentiation of BAT is poorly understood. We investigated the role of BH4 in differentiation of BAT using a mouse model of BH4 deficiency, the Hph-1 mouse. Hph-1 neonatal mice exhibit dysplasia of BAT as well as attenuated thermogenesis-related gene expressions (UCP1, Dio2, Pgc1a) compared with control mice. As differentiation capacity is considered to be at maximum during late gestation, we administered BH4 to fetal Hph-1 mice via placental transfer using intraperitoneal injection to pregnant Hph-1 mice for 6 days just before birth. Maternal BH4 supplementation ameliorated dysplasia of neonatal BAT and restored thermogenesis-related genes expression. Intriguingly, offspring of the maternal BH4-supplementation group showed less weight gain, ameliorated glucose intolerance, and improved cold tolerance after growth under high fat chow diet. We then evaluated the direct effect of BH4 on brown adipocyte differentiation using brown pre-adipocytes isolated from control and Hph-1 mice. Compared with control mice, brown pre-adipocytes from Hph-1 mice showed reduced differentiation capacity, while BH4 supplementation in brown pre-adipocytes from Hph-1 mice ameliorated differentiation capacity nitric oxide-dependently. Taking these findings together, BH4 plays an integral role in the differentiation of BAT, especially in the fetal period, and may represent a novel target for prevention of metabolic disorders such as obesity and diabetes. Disclosure H. Minamino: None. Y. Fujita: None. Y. Oguri: None. T. Goto: None. A. Ohashi: None. F. Furuya: None. N. Isomura: None. T. Kohei: None. Y. Li: None. S. Kawarasaki: None. T. Kawada: None. H. Hasegawa: None. N. Inagaki: Research Support; Self; Astellas Pharma Inc., Drawbridge Health, Japan Tobacco Inc., Kissei Pharmaceutical Co., Ltd., Kyowa Hakko Kirin Co., Ltd., Life Scan Japan, Nippon Boehringer Ingelheim Co. Ltd., Novartis Pharma K.K., Novo Nordisk Pharma Ltd.,, Sanofi K.K., Sanwa Kagaku Kenkyusho, Terumo Medical Corporation. Speaker’s Bureau; Self; Astellas Pharma Inc., Merck & Co., Inc., Mitsubishi Tanabe Pharma Corporation, Nippon Boehringer Ingelheim Co. Ltd., Novo Nordisk Inc., Ono Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited.

Excessive Glucocorticoids During Pregnancy Impair Fetal Brown Fat Development and Predispose Offspring to Metabolic Dysfunctions.

Maternal stress during pregnancy exposes fetuses to hyperglucocorticoids, which increases the risk of metabolic dysfunctions in offspring. Despite being a key tissue for maintaining metabolic health, the impacts of maternal excessive glucocorticoids (GC) on fetal brown adipose tissue (BAT) development and its long-term thermogenesis and energy expenditure remain unexamined. For testing, pregnant mice were administered dexamethasone (DEX), a synthetic GC, in the last trimester of gestation, when BAT development is the most active. DEX offspring had glucose, insulin resistance, and adiposity and also displayed cold sensitivity following cold exposure. In BAT of DEX offspring, Ppargc1a expression was suppressed, together with reduced mitochondrial density, and the brown progenitor cells sorted from offspring BAT demonstrated attenuated brown adipogenic capacity. Increased DNA methylation in Ppargc1a promoter had a fetal origin; elevated DNA methylation was also detected in neonatal BAT and brown progenitors. Mechanistically, fetal GC exposure increased GC receptor/DNMT3b complex in binding to the Ppargc1a promoter, potentially driving its de novo DNA methylation and transcriptional silencing, which impaired fetal BAT development. In summary, maternal GC exposure during pregnancy increases DNA methylation in the Ppargc1a promoter, which epigenetically impairs BAT thermogenesis and energy expenditure, predisposing offspring to metabolic dysfunctions.

Comprehensive Analysis of the Characteristics and Differences in Adult and Newborn Brown Adipose Tissue (BAT): Newborn BAT Is a More Active/Dynamic BAT.

Brown adipose tissue (BAT) plays an essential role in maintaining body temperature and in treating obesity and diabetes. The adult BAT (aBAT) and neonatal BAT (neBAT) vary greatly in capacity, but the characteristics and differences between them on the molecular level, as well as the related features of BAT as it develops post-delivery, have not yet been fully determined. In this study, we examined the morphological features of aBAT and neBAT of mice by using hematoxylin-eosin (H&E) staining, transmission electron microscopy (TEM), and scanning electron microscopy (SEM). We found that neBAT contains a smaller number and size of lipid droplets, as well as more abundant mitochondria, compared with aBAT. The dynamic morphological changes revealed that the number and size of lipid droplets increase, but the number of mitochondria gradually decrease during the post-delivery development, which consisted of some differences in RNA or protein expression levels, such as gradually decreased uncoupling protein 1 (UCP1) expression levels and mitochondrial genes, such as mitochondrial transcription factor A (Tfam). The adipocyte differentiation-related genes, such as transcription factor CCAAT enhancer-binding protein β (CEBPβ), were also continuously upregulated. Additionally, the different features of aBAT and neBAT were analyzed from the global transcription (RNA-Seq) level, which included messenger RNA (mRNA), microRNA, long non-coding RNA (lncRNA), circRNA, and DNA methylation, as well as proteins (proteomics). Differentially methylated region (DMR) analysis identified 383 hyper- and 503 hypo-methylated genes, as well as 1221 new circRNA in ne-BAT and 1991 new circRNA in a-BAT, with significantly higher expression of circRNA in aBAT compared with neBAT. Gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that mitochondrial activity, protein synthesis, and cell life activity levels were higher in neBAT, and pathways related to ribosomes, spliceosomes, and metabolism were significantly activated in neBAT compared to aBAT. Collectively, this study describes the dynamic changes occurring throughout post-delivery development from the morphological, molecular and omics perspectives. Our study provides information that may be utilized in improving BAT functional activity through gene regulation and/or epigenetic regulation.

GPR120 controls neonatal brown adipose tissue thermogenic induction.

Adaptive induction of thermogenesis in brown adipose tissue (BAT) is essential for the survival of mammals after birth. We show here that G protein-coupled receptor protein 120 (GPR120) expression is dramatically induced after birth in mouse BAT. GPR120 expression in neonatal BAT is the highest among GPR120-expressing tissues in the mouse at any developmental stage tested. The induction of GPR120 in neonatal BAT is caused by postnatal thermal stress rather than by the initiation of suckling. GPR120-null neonates were found to be relatively intolerant to cold: close to one-third did not survive at 21°C, but all such pups survived at 25°C. Heat production in BAT was significantly impaired in GPR120-null pups. Deficiency in GPR120 did not modify brown adipocyte morphology or the anatomical architecture of BAT, as assessed by electron microscopy, but instead impaired the expression of uncoupling protein-1 and the fatty acid oxidation capacity of neonatal BAT. Moreover, GPR120 deficiency impaired fibroblast growth factor 21 (FGF21) gene expression in BAT and reduced plasma FGF21 levels. These results indicate that GPR120 is essential for neonatal adaptive thermogenesis.

Glycerol-3-phosphate Acyltransferase Isoform-4 (GPAT4) Limits Oxidation of Exogenous Fatty Acids in Brown Adipocytes

Glycerol-3-phosphate acyltransferase-4 (GPAT4) null pups grew poorly during the suckling period and, as adults, were protected from high fat diet-induced obesity. To determine why Gpat4(-/-) mice failed to gain weight during these two periods of high fat feeding, we examined energy metabolism. Compared with controls, the metabolic rate of Gpat4(-/-) mice fed a 45% fat diet was 12% higher. Core body temperature was 1 ºC higher after high fat feeding. Food intake, fat absorption, and activity were similar in both genotypes. Impaired weight gain in Gpat4(-/-) mice did not result from increased heat loss, because both cold tolerance and response to a β3-adrenergic agonist were similar in both genotypes. Because GPAT4 comprises 65% of the total GPAT activity in brown adipose tissue (BAT), we characterized BAT function. A 45% fat diet increased the Gpat4(-/-) BAT expression of peroxisome proliferator-activated receptor α (PPAR) target genes, Cpt1α, Pgc1α, and Ucp1, and BAT mitochondria oxidized oleate and pyruvate at higher rates than controls, suggesting that fatty acid signaling and flux through the TCA cycle were enhanced. To assess the role of GPAT4 directly, neonatal BAT preadipocytes were differentiated to adipocytes. Compared with controls, Gpat4(-/-) brown adipocytes incorporated 33% less fatty acid into triacylglycerol and 46% more into the pathway of β-oxidation. The increased oxidation rate was due solely to an increase in the oxidation of exogenous fatty acids. These data suggest that in the absence of cold exposure, GPAT4 limits excessive fatty acid oxidation and the detrimental induction of a hypermetabolic state.

Maternal low protein diet leads to dysregulation of placental iNKT cells and M1/M2 macrophage ratio, body weight loss in male, neonate Sprague‐Dawley rats and increased UCP‐1 mediated thermogenesis

Placental immune cells provide cytokines and growth factors that are necessary for placenta development and function. Invariant natural killer T (iNKT) cells are innate cells specific for glycolipid antigens presented by the CD1d molecule and secrete Th1 cytokines in the placenta, suggesting an immunoregulatory role. Self‐lipid antigens at the maternal‐fetal interface may activate iNKT cells, leading to placental inflammation and dysfunction, which can increase brown adipose tissue (BAT) thermogenesis and reduce fetal growth. Maternal low protein (LP) diets reduce offspring birth weight (BW) perhaps via a BAT thermogenesis‐mediated effect. However, whether maternal LP nutrition alters placental immune cell populations and offspring thermogenesis in not yet known. Obese‐prone Sprague‐Dawley dams were fed 8% LP or 20% normal protein (NP) diets for 3 wks prior to breeding and through pregnancy. BW of neonatal LP males was lower than NP males. Vα14 mRNA marker for iNKT cells was 11‐fold higher in LP placenta, while there was also an increase in proinflammatory M1/M2 macrophage mRNA marker ratio. UCP‐1 gene and protein expression and transcription factors PRDM16 and PPARα in BAT were 2‐ to 6‐fold greater in LP than NP neonatal males. FNDC5, an irisin precursor and thermogenesis activator, was 2‐fold greater in neonatal LP BAT than NP males. These results suggest that prenatal protein restriction increases proinflammatory iNKT and M1 macrophages in the placenta, leading to placental dysfunction, increased offspring BAT thermogenesis, and low BW.

Neonatal brown adipose tissue, UCP1 and the novel uncoupling proteins.

Neonatal brown adipose tissue, UCP1 and the novel uncoupling proteins.

Prepartum Protein Restriction Does Not Alter Norepinephrine-Induced Thermogenesis or Brown Adipose Tissue Function in Newborn Calves , ,

We examined the effect of prepartum protein restriction on thermogenesis and several aspects of perirenal (brown) adipose tissue (BAT) in newborn calves. Lipid synthesis and morphology also were compared between BAT and sternum (white) adipose tissue. During the last 140 d of gestation, heifers were fed isocaloric diets containing adequate (10.4%) or restricted (average of 6.8%, dry matter basis) levels of protein. Body condition scores and weight gain during gestation were significantly lower in heifers fed the restricted-protein diet. However, newborn calf birth weight, calf BAT weight and composition, and calf thermoneutral metabolic rates were not affected by prepartum protein restriction. Similarly, visceral organ weights, except for lung plus trachea, were not affected (P > 0.10) by prepartum protein treatment. Peak metabolic rates were not affected (P > 0.10) by prepartum protein treatment and on average were twice the thermoneutral metabolic rates. Consistent with this, BAT of calves from heifers fed adequate- or restricted-protein diets did not differ in lipid synthesis, cellularity, or uncoupling protein mRNA:28S rRNA ratios. Although both perirenal and sternum adipocytes were mostly unilocular, perirenal adipocytes contained numerous large mitochondria with well-differentiated cristae; sternum adipocytes contained a small number of mitochondria with poorly developed cristae. Fatty acid biosynthesis from acetate was high in BAT (55–57 nmol acetate incorporated⋅100 mg−1⋅h−1) but barely detectable in sternum adipose tissue. Conversely, fatty acid biosynthesis from glucose was 80–110% higher in sternum adipose tissue than in BAT (4.5 vs 2.1–2.5 nmol glucose incorporated⋅100 mg−1⋅h−1). Thus maternal protein restriction severely affected heifers but had no effect on neonatal calf thermogenesis or BAT function.

Perinatal expression of adenylyl cyclase subtypes in rat brown adipose tissue.

The ability of norepinephrine to stimulate adenylyl cyclase (AC) activity increases during the perinatal period in rat brown adipose tissue (BAT), and this increase is associated with changes in the activities of both GS alpha and AC. The purpose of this study was to determine which AC subtypes are present in neonatal BAT and to examine whether the perinatal increase in AC activity corresponds to an increase in the expression of a particular AC subtype. Analysis of AC mRNAs by nuclease protection assay demonstrated the presence of mRNAs encoding AC-III, AC-IV, AC-VI, and AC-IX in embryonic and postnatal BAT. Of the subtypes detected, only AC-III mRNA levels increased substantially during the perinatal period. The increase in AC-III expression was paralleled by an increase in isoproterenol-stimulated AC activity. Treatment of neonates was the sympathetic neurotoxin 6-hydroxydopamine abolished the perinatal increase in both AC activity and AC-III mRNA levels but had no effect on the expression of other AC subtypes. These results strongly indicate that the increase in AC activity during the perinatal period is due to an increase in the expression of AC-III.

Brown adipose tissue development in the ovine foetus during the final month of gestation

In the ovine foetus brown adipose tissue (BAT) is mainly found in the perirenal region and grows rapidly relative to body weight between 70 to 120 days of gestation (Alexander, 1978). After this stage only a small amount of BAT growth occurs in comparison with that of the whole foetus, and in the case of undernutrition may decline (Alexander, 1978). Maternal cold stress, induced by winter shearing twin-bearing pregnant ewes 8 weeks before parturition improves lamb birth weight and lamb growth rate independently of effects on maternal food intake (Symonds, Bryant and Lomax, 1986 and 1990). At the same time this can stimulate the in vivo capacity for non-shivering thermogenesis in newborn lambs (Stott and Slee, 1985). The following study extends these findings by investigating the extent to which changing the maternal metabolic environment influences BAT development over the final month of gestation.Thirty-two Bluefaced Leicester × Swaledale ewes were housed individually at ambient temperature (−6 to 19°C) 6 weeks prior to lambing and 2 weeks later 15 ewes were shorn. Ewes were offered daily a diet comprising 200 g barley concentrate and 1 kg chopped hay. Between 116 and 145 days of gestation and within 2 h of birth ewes were humanely slaughtered with an overdose of barbiturate and foetal or neonatal perirenal BAT sampled, born from shorn or unshorn ewes. The thermogenic capacity of BAT was assessed by guanosine-5′-diphosphate (GDP) binding to uncoupling protein in mitochondrial preparations (Cooper, Dascombe, Rothwell and Vale, 1989) and the amount of mitochondrial protein measured from cytochrome Coxidase activity.

Two Pathways for Thyroxine 5’-Monodeiodination in Brown Adipose Tissue in Fetal Sheep: Ontogenesis and Divergent Responses to Hypothyroidism and 3,5,3’- Triiodothyronine Replacement*

Thermogenesis in rat brown adipose tissue (BAT) is thyroid hormone responsive. Rat BAT expresses a type II 5'-iodothyronine monodeiodinase (5'MDI) which mediates local T3 production from T4. Earlier studies show that BAT from fetal and newborn sheep contains a high Km type I, instead of type II, 5'MDI. To better characterize the 5'MDI of ovine fetal BAT, we studied the in vitro monodeiodination of [125I]T4 at a low substrate concentration (2 nM) and in the presence of 1 mM propylthiouracil in BAT homogenates of control and thyroidectomized fetuses at different gestational ages as well as in newborn lambs. Thyroidectomies were performed at three gestational ages: 99-107 days (group 1), 129-132 days (group 2), and 115-117 days (group 3A). Animals were studied 8-13 days after surgery. A significant increase in the activity of a low Km T4 5'MDI was noted in BAT from hypothyroid fetuses at all three gestational ages. This low Km activity was similar to the type II enzyme in rat BAT and brain in that the activity was also T3 resistant. A gradual rise in BAT type II 5'MDI activity was measured between 99 days gestation and term (150 days). These results indicate that ovine BAT contains two distinct iodothyronine 5'-monodeiodinating activities, one with a high Km and another with a low Km. The latter, resembling the type II 5'MDI in rat brain and BAT, is increased in ovine hypothyroid BAT. The former predominates in euthyroid tissue and is similar to the type I 5'MDI characterized in rat liver, kidney, and thyroid. We speculate that BAT type II 5'MDI may be important for neonatal BAT thermogenesis, while the type I enzyme may play a significant role in the increase in serum T3 concentration that occurs at birth.

MEASUREMENT OF THERMOGENIN SYNTHESIS IN BROWN ADIPOSE TISSUE (BAT)

Thermogenin is a mitochondrial inner membrane protein which mediates non-shivering thermogenesis in BAT. It has a molecular mass of 32,000 Da and acts by uncoupling mitochondria to produce heat. We report a method of measuring its synthesis. Polysomes were isolated from BAT of developing rabbits and translated in a reticulocyte protein synthesizing system. Newly synthesized thermogenin was isolated with monospecific antibodies. This technique has been applied in developing rabbits to demonstrate a 5-fold increase in thermogenin synthesis before birth, and a gradual decrease after birth. The technique provides a method to study other perinatal influences on thermogenin synthesis such as the effect of postnatal environmental temperature. In preliminary experiments thermogenin has been purified from human neonatal BAT.

Altered Brown Adipose Tissue Mitochondrial Function in Neonates Born to Rats Overfed Foods of Various Protein Content

On day 3 of pregnancy, 20 female Sprague-Dawley rats were randomly divided into four groups and fed either a powdered, control diet containing 27% casein (control) or “cafeteria” diets of low (7%, LP), normal (26%, NP) or high (35%, HP) protein content throughout gestation. Maternal gross energy intake was significantly increased 35% in each of the cafeteria-fed groups compared to control. At birth, mean body weight of the offspring in each diet group was not significantly different, whereas brown adipose tissue (BAT) weight and mitochondrial function (unrestricted minus GDP-inhibited anion conductance) were significantly decreased in LP offspring and increased in NP and HP offspring when compared to controls. Maternal diet composition, therefore, can alter neonatal BAT mitochondrial function and, perhaps, thermogenic capacity independent of total energy intake during pregnancy.

The development of carnitine synthesis from gamma-butyrobetaine in the rat.

Abstract The ability of the rat liver to synthesize camitine from γ-butyrobetaine increases from low values in the fetus to adult values on the 8th day after birth. The rate of synthesis of camitine is greater when determined in the high-speed supernatant than in the low-speed supernatant of the liver. No synthesis could be shown to occur in neonatal rat kidney or neonatal brown adipose tissue.