UCP1 Content Research Articles

Polyelectrolyte nanofilms on cell surface can induce brown adipogenic differentiation of DFATs

Obesity and its related health issues significantly burden public health systems. Brown adipose tissue holds promise for addressing metabolic disorders and balancing the body's energy, making it a key research focus. Stimulating brown adipogenesis from stem cells could advance regenerative medicine and healthcare. In our previous research, we discovered that poly-l-lysine (PLL) significantly stimulates brown adipogenesis in three-dimensional differentiation of dedifferentiated fat cells (DFATs) within fibrin gels. In this study, we evaluated polyelectrolyte (PE) nanofilms made of PLL and dextran sulfate, applied directly to DFAT surfaces to improve brown adipogenic differentiation through an innovative approach. This approach involved coating the DFAT surfaces with PE nanofilms, forming a multilayer structure that not only provided a supportive matrix but also facilitated the adsorption of essential molecules like T3 and insulin for brown adipogenesis. DFATs coated with three PE layers and encapsulated in fibrin gel showed a significant increase in the adipogenic marker UCP1 gene expression and content. This PLL-based PE nanofilm coating on DFAT surfaces can be a novel and crucial technology for promoting brown adipogenesis in regenerative medicine and healthcare.

Walnut supplementation increases levels of UCP1 and CD36 in brown adipose tissue independently of diet type.

Dietary interventions that modulate the brown adipose tissue (BAT) thermogenic activity could represent a promising therapy for metabolic disorders. In order to examine if dietary walnuts intake regulates the expression of BAT thermogenic markers levels in healthy and metabolically challenged (fructose fed) animals, rats were initially divided into the control and fructose-fed groups. After nine weeks, these groups were subdivided into the one kept on the original regimens and the other supplemented with walnuts. High-fructose diet resulted in an increased relative BAT mass and no change in UCP1 content, while the walnut supplementation increased the amount of UCP1 in BAT, but did not affect 5-HT, NA, DHPG content and DHPG/NA ratio regardless of the diet. Moreover, the CD36 levels were increased following the walnut consumption, unlike FATP1, GLUT1, GLUT4, and glycogen content which remained unchanged. Additionally, the BAT levels of activated IR and Akt were not affected by walnut consumption, while ERK signaling was decreased. Overall, we found that walnut consumption increased UCP1 and CD36 content in the BAT of both control and metabolically challenged rats, suggesting that FFAs represent the BAT preferred substrate under the previously described circumstances. This further implies that incorporating walnuts into the everyday diet may help to alleviate some symptoms of the metabolic disorder.

FoxO1 regulates adipose transdifferentiation and iron influx by mediating Tgfβ1 signaling pathway

Adipose plasticity is critical for metabolic homeostasis. Adipocyte transdifferentiation plays an important role in adipose plasticity, but the molecular mechanism of transdifferentiation remains incompletely understood. Here we show that the transcription factor FoxO1 regulates adipose transdifferentiation by mediating Tgfβ1 signaling pathway. Tgfβ1 treatment induced whitening phenotype in beige adipocytes, reducing UCP1 and mitochondrial capacity and enlarging lipid droplets. Deletion of adipose FoxO1 (adO1KO) dampened Tgfβ1 signaling by downregulating Tgfbr2 and Smad3 and induced browning of adipose tissue in mice, increasing UCP1 and mitochondrial content and activating metabolic pathways. Silencing FoxO1 also abolished the whitening effect of Tgfβ1 on beige adipocytes. The adO1KO mice exhibited a significantly higher energy expenditure, lower fat mass, and smaller adipocytes than the control mice. The browning phenotype in adO1KO mice was associated with an increased iron content in adipose tissue, concurrent with upregulation of proteins that facilitate iron uptake (DMT1 and TfR1) and iron import into mitochondria (Mfrn1). Analysis of hepatic and serum iron along with hepatic iron-regulatory proteins (ferritin and ferroportin) in the adO1KO mice revealed an adipose tissue-liver crosstalk that meets the increased iron requirement for adipose browning. The FoxO1-Tgfβ1 signaling cascade also underlay adipose browning induced by β3-AR agonist CL316243. Our study provides the first evidence of a FoxO1-Tgfβ1 axis in the regulation of adipose browning-whitening transdifferentiation and iron influx, which sheds light on the compromised adipose plasticity in conditions of dysregulated FoxO1 and Tgfβ1 signaling.

Fenofibrate mildly stimulates browning-associated expression in white adipose tissues of young but not old male rats.

The aim of this study was to examine the effects of the hypolipemic drug fenofibrate (FF) and aging on the expression of factors/enzymes involved in brown adipose tissue (BAT) function and browning of white adipose tissue epididymal (eWAT) and subcutaneous (sWAT) depots. Young-adult and old male Wistar rats were fed standard chow (control) or supplemented with 0.1% or 0.5% FF for 30 days. Tissue samples were analysed for gene expression and protein content, and stained with Oil Red O or hematoxylin and eosin. In BAT of young rats, 0.5% FF increased only Cbp/p300 interacting transactivator with Glu/Asp rich carboxy-terminal domain 1 (CITED1) protein content and Fgf21 and Gpr109A mRNA expression. The expression of oxidative metabolism related genes (Pgc1α, Cpt1b, Mcad) decreased after 0.5% FF. In BAT of old rats, FF did not affect UCP1 and CITED1 content and had little effect on gene expression. Oil Red O staining of BAT revealed no changes in lipid droplet area upon treatment in either age group. In eWAT of young rats, 0.1FF elevated UCP1 protein content and Ucp1, Pgc-1α, and Mcad expression, whereas 0.5% FF increased PPARα content and Pgc-1α, Cpt1b, Mcad, and Gpr109A levels. In eWAT of old rats, only 0.1FF increased Pgc1α and Mcad expression. In both age groups median cell area of eWAT adipocytes was reduced after 0.5% FF. In sWAT Ucp1 gene expression was very low and UCP1 protein was undetectable. FF upregulated Ucp1, Cited1, Eva1, and Cpt1b expression in sWAT of young rats, with diminished effects in old rats. In both age groups 0.5% FF increased Fgf21 expression in sWAT. Median cell area of sWAT adipocytes decreased only in young rats treated with 0.5% FF. Our results reveal that fenofibrate differentially affects gene expression in BAT, with diminished effects in old compared to young rats. In WAT of young rats FF modestly stimulates the expression of factors/enzymes involved in lipid oxidative metabolism and browning. Aging reduces both these effects. Gpr109A may present a novel gene target upregulated by FF in BAT and eWAT.

Yerba mate consumption by ovariectomized rats alters white adipose tissue

IntroductionMenopause and post-menopause are characterized by low levels of estrogen that can be associated with the emergence of metabolic diseases. While hormone replacement therapy can alleviate many symptoms, it can also exacerbate other diseases such as breast cancer. In the search for natural alternatives, Ilex paraguariensis (Yerba Mate) has been identified as a potential therapy for the onset of obesity. Here, the effect of MATE consumption on white adipose tissue (WAT) was studied in ovariectomized rats, an animal model for post-menopause hormone loss. MethodsFour groups of animals were used: ovariectomy with MATE (OVX MATE) and without MATE (OVX), as well as sham surgery with MATE (Sham MATE) and without MATE (Sham). MATE was provided by gavage at 1 g/kg of body weight for eight weeks before measuring biochemical parameters in plasma and characterizing WAT morphology. ResultsThe consumption of Yerba MATE significantly decreased weight gain in ovariectomized rats and presented near control levels of triglycerides, total cholesterol, and LDL. A morphometric analysis of WAT showed a significant decrease in the area occupied by adipocytes in the group that consumed MATE. Finally, MATE consumption increased the UCP1 content in the WAT of the ovariectomized group. Yerba MATE treatment was also associated with higher levels of SIRT1 protein. ConclusionMATE consumption has a preventive effect on the weight gain observed in ovariectomized rats and potential benefits in naturally avoiding the onset of obesity post menopause.

Autophagy-mediated NCOR1 degradation is required for brown fat maturation and thermogenesis

Brown adipose tissue (BAT) thermogenesis affects energy balance, and thereby it has the potential to induce weight loss and to prevent obesity. Here we document a macroautophagic/autophagic-dependent mechanism of PPARG activity regulation that induces brown adipose differentiation and thermogenesis, and that is mediated by TP53INP2. Disruption of TP53INP2-dependent autophagy reduced brown adipogenesis in cultured cells. In vivo specific-tp53inp2 ablation in brown precursor cells or in adult mice decreased the expression of thermogenic and mature adipocytes genes in BAT. As a result, TP53INP2-deficient mice had reduced UCP1 content in BAT and impaired maximal thermogenic capacity, leading to lipid accumulation and to positive energy balance. Mechanistically, TP53INP2 stimulates PPARG activity and adipogenesis in brown adipose cells by promoting the autophagic degradation of NCOR1, a PPARG co-repressor. Moreover, the modulation of TP53INP2 expression in BAT and in human brown adipocytes suggest that this protein increases PPARG activity during metabolic activation of brown fat. In all, we have identified a novel molecular explanation to the contribution of autophagy to BAT energy metabolism that could facilitate the design of therapeutic strategies against obesity and its metabolic complications.

Maternal L-carnitine supplementation promotes brown adipose tissue thermogenesis of newborn goats after cold exposure.

Brown adipose tissue (BAT) is an important component of energy expenditure and necessary to maintain body temperature for newborn mammals. In the previous study, we found that L-carnitine was enriched in BAT and promoted BAT adipogenesis and thermogenesis in goat brown adipocytes. However, whether dietary L-carnitine regulates BAT heat production and energy expenditure in lambs remains unclear. In this study, maternal L-carnitine supplementation elevated the rectal temperature, as well as the expression of UCP1 and mitochondrial DNA content to promote BAT thermogenesis in newborn goats. Moreover, maternal L-carnitine supplementation increased the levels of triglycerides (TG), non-esterified fatty acids (NEFA), and lactate in plasma, as well as the content of lipid droplet and glycogen in BAT of newborn goats. Lipidomic analysis showed that maternal L-carnitine supplementation remodeled the lipid composition of BAT in newborn goats. L-carnitine significantly increased the levels of TG and diglyceride (DG) and decreased the levels of glycerophospholipids and sphingolipids in BAT. Further studies showed that L-carnitine promoted TG and glycogen deposition in brown adipocytes through AMPKα. Our results indicate that maternal L-carnitine supplementation promotes BAT development and thermogenesis in newborn goats and provides new evidence for newborn goats to maintain body temperature in response to cold exposure.

Obligatory homeothermy of mesic habitat-adapted African striped mice, Rhabdomys pumilio, is governed by seasonal basal metabolism and year-round 'thermogenic readiness' of brown adipose tissue.

Small mammals undergo thermoregulatory adjustments in response to changing environmental conditions. Whereas small heterothermic mammals can employ torpor to save energy in the cold, homeothermic species must increase heat production to defend normothermia through the recruitment of brown adipose tissue (BAT). Here, we studied thermoregulatory adaptation in an obligate homeotherm, the African striped mouse (Rhabdomys pumilio), captured from a subpopulation living in a mesic, temperate climate with marked seasonal differences. Basal metabolic rate (BMR), non-shivering thermogenesis (NST) and summit metabolic rate (Msum) increased from summer to winter, with NST and Msum already reaching maximal rates in autumn, suggesting seasonal preparation for the cold. Typical of rodents, cold-induced metabolic rates were positively correlated with BAT mass. Analysis of cytochrome c oxidase (COX) activity and UCP1 content, however, demonstrated that thermogenic capacity declined with BAT mass. This resulted in seasonal differences in NST being driven by changes in BMR. The increase in BMR was supported by a comprehensive anatomical analysis of metabolically active organs, revealing increased mass proportions in the cold season. The thermoregulatory response of R. pumilio was associated with the maintenance of body mass throughout the year (48.3±1.4 g), contrasting large summer-winter mass reductions often observed in Holarctic rodents. Collectively, bioenergetic adaptation of this Afrotropical rodent involves seasonal organ adjustments influencing BMR, combined with a constant thermogenic capacity dictated by trade-offs in the thermogenic properties of BAT. Arguably, this high degree of plasticity was a response to unpredictable cold spells throughout the year. Consequently, the reliance on such a resource-intensive thermoregulatory strategy may expose more energetic vulnerability in changing environments of food scarcity and extreme weather conditions due to climate change, with major ramifications for survival of the species.

Mitophagy Mediates the Beige to White Transition of Human Primary Subcutaneous Adipocytes Ex Vivo.

Brown and beige adipocytes have multilocular lipid droplets, express uncoupling protein (UCP) 1, and promote energy expenditure. In rodents, when the stimulus of browning subsides, parkin-dependent mitophagy is activated and dormant beige adipocytes persist. In humans, however, the molecular events during the beige to white transition have not been studied in detail. In this study, human primary subcutaneous abdominal preadipocytes were differentiated to beige for 14 days, then either the beige culture conditions were applied for an additional 14 days or it was replaced by a white medium. Control white adipocytes were differentiated by their specific cocktail for 28 days. Peroxisome proliferator-activated receptor γ-driven beige differentiation resulted in increased mitochondrial biogenesis, UCP1 expression, fragmentation, and respiration as compared to white. Morphology, UCP1 content, mitochondrial fragmentation, and basal respiration of the adipocytes that underwent transition, along with the induction of mitophagy, were similar to control white adipocytes. However, white converted beige adipocytes had a stronger responsiveness to dibutyril-cAMP, which mimics adrenergic stimulus, than the control white ones. Gene expression patterns showed that the removal of mitochondria in transitioning adipocytes may involve both parkin-dependent and -independent pathways. Preventing the entry of beige adipocytes into white transition can be a feasible way to maintain elevated thermogenesis and energy expenditure.

Lean in one way, in obesity another: effects of moderate exercise in brown adipose tissue of early overfed male Wistar rats.

Early postnatal overfeeding (PO) induces long-term overweight and reduces brown adipose tissue (BAT) thermogenesis. Exercise has been suggested as a possible intervention to increase BAT function. In this study, we investigated chronical effects of moderate-intensity exercise in BAT function in postnatal overfed male Wistar rats METHODS: Litters' delivery was on postnatal-day 0 - PN0. At PN2, litters were adjusted to nine (normal litter - NL) or three pups (small litter - SL) per dam. Animals were weaned on PN21 and in PN30 randomly divided into sedentary (NL-Sed and SL-Sed) or exercised (NL-Exe and SL-Exe), N of 14 litters per group. Exercise protocol started (PN30) with an effort test; training sessions were performed three times weekly at 60% of the VO2max achieved in effort test, until PN80. On PN81, a temperature transponder was implanted beneath the interscapular BAT, whose temperature was assessed in periods of lights-on and -off from PN87 to PN90. Sympathetic nerve activation of BAT was registered at PN90. Animals were euthanized at PN91 and tissues collected RESULTS: PO impaired BAT thermogenesis in lights-on (pPO < 0.0001) and -off (pPO < 0.01). Exercise increased BAT temperature in lights-on (pExe < 0.0001). In NL-Exe, increased BAT activity was associated with higher sympathetic activity (pExe < 0.05), β3-AR (pExe < 0.001), and UCP1 (pExe < 0.001) content. In SL-Exe, increasing BAT thermogenesis is driven by a combination of tissue morphology remodeling (pExe < 0.0001) with greater effect in increasing UCP1 (pExe < 0.001) and increased β3-AR (pExe < 0.001) content. Moderate exercise chronically increased BAT thermogenesis in both, NL and SL groups. In NL-Exe by increasing Sympathetic activity, and in SL-Exe by a combination of increased β3-AR and UCP1 content with morphologic remodeling of BAT. Chronically increasing BAT thermogenesis in obese subjects may lead to higher overall energy expenditure, favoring the reduction of obesity and related comorbidities.

Adipocyte-specific mTORC2 deficiency impairs BAT and iWAT thermogenic capacity without affecting glucose uptake and energy expenditure in cold-acclimated mice.

Deletion of mechanistic target of rapamycin complex 2 (mTORC2) essential component rapamycin insensitive companion of mTOR (Rictor) by a Cre recombinase under control of the broad, nonadipocyte-specific aP2/FABP4 promoter impairs thermoregulation and brown adipose tissue (BAT) glucose uptake on acute cold exposure. We investigated herein whether adipocyte-specific mTORC2 deficiency affects BAT and inguinal white adipose tissue (iWAT) signaling, metabolism, and thermogenesis in cold-acclimated mice. For this, 8-wk-old male mice bearing Rictor deletion and therefore mTORC2 deficiency in adipocytes (adiponectin-Cre) and littermates controls were either kept at thermoneutrality (30 ± 1°C) or cold-acclimated (10 ± 1°C) for 14 days and evaluated for BAT and iWAT signaling, metabolism, and thermogenesis. Cold acclimation inhibited mTORC2 in BAT and iWAT, but its residual activity is still required for the cold-induced increases in BAT adipocyte number, total UCP-1 content and mRNA levels of proliferation markers Ki67 and cyclin 1 D, and de novo lipogenesis enzymes ATP-citrate lyase and acetyl-CoA carboxylase. In iWAT, mTORC2 residual activity is partially required for the cold-induced increases in multilocular adipocytes, mitochondrial mass, and uncoupling protein 1 (UCP-1) content. Conversely, BAT mTORC1 activity and BAT and iWAT glucose uptake were upregulated by cold independently of mTORC2. Noteworthy, the impairment in BAT and iWAT total UCP-1 content and thermogenic capacity induced by adipocyte mTORC2 deficiency had no major impact on whole body energy expenditure in cold-acclimated mice due to a compensatory activation of muscle shivering. In conclusion, adipocyte mTORC2 deficiency impairs, through different mechanisms, BAT and iWAT total UCP-1 content and thermogenic capacity in cold-acclimated mice, without affecting glucose uptake and whole body energy expenditure.NEW & NOTEWORTHY BAT and iWAT mTORC2 is inhibited by cold acclimation, but its residual activity is required for cold-induced increases in total UCP-1 content and thermogenic capacity, but not glucose uptake and mTORC1 activity. The impaired BAT and iWAT total UCP-1 content and thermogenic capacity induced by adipocyte mTORC2 deficiency are compensated by activation of muscle shivering in cold-acclimated mice.

Cold Exposure Distinctively Modulates Parathyroid and Thyroid Hormones in Cold-Acclimatized and Non-Acclimatized Humans.

Cold-induced activation of thermogenesis modulates energy metabolism, but the role of humoral mediators is not completely understood. We aimed to investigate the role of parathyroid and thyroid hormones in acute and adaptive response to cold in humans. Examinations were performed before/after 15 minutes of ice-water swimming (n = 15) or 120 to 150 minutes of cold-induced nonshivering thermogenesis (NST) applied to cold-acclimatized (n = 6) or non-acclimatized (n = 11) individuals. Deep-neck brown adipose tissue (BAT) was collected from non-acclimatized patients undergoing elective neck surgery (n = 36). Seasonal variations in metabolic/hormonal parameters of ice-water swimmers were evaluated. We found that in ice-water swimmers, PTH and TSH increased and free T3, T4 decreased after a 15-minute winter swim, whereas NST-inducing cold exposure failed to regulate PTH and free T4 and lowered TSH and free T3. Ice-water swimming-induced increase in PTH correlated negatively with systemic calcium and positively with phosphorus. In non-acclimatized men, NST-inducing cold decreased PTH and TSH. Positive correlation between systemic levels of PTH and whole-body metabolic preference for lipids as well as BAT volume was found across the 2 populations. Moreover, NST-cooling protocol-induced changes in metabolic preference for lipids correlated positively with changes in PTH. Finally, variability in circulating PTH correlated positively with UCP1/UCP1, PPARGC1A, and DIO2 in BAT from neck surgery patients. Our data suggest that regulation of PTH and thyroid hormones during cold exposure in humans varies by cold acclimatization level and/or cold stimulus intensity. Possible role of PTH in NST is indicated by its positive relationships with whole-body metabolic preference for lipids, BAT volume, and UCP1 content.

Chronic serotonin reuptake inhibition uncouples brown fat mitochondria and induces beiging/browning process of white fat in overfed rats

AimTo investigate whether a chronic 5-HT reuptake inhibitor (i.e. Fluoxetine-FLX) exposure in young adult rats overfed during suckling period would modulate interscapular brown adipose tissue (iBAT) mitochondria and browning agents in white adipose tissue (WAT). MethodsMale Wistar rats were assigned into either a normofed group (n = 9 per group) or an overfed group (n = 3 per group) induced by litter size reduction at postnatal day 3 (PND3). Pharmacological manipulation was carried out between PND39 and PND59 and groups were assigned accordingly: Normofed + vehicle solution – NaCl 0.9% (NV group), normofed + FLX solution – 10 mg/kg b.w. (NF group), overfed + vehicle (OV group) and overfed + FLX (OF group). We evaluated mitochondrial oxygen consumption and reactive species (RS) production, oxidative stress analyses (MDA concentration, carbonyl content, REDOX state [GSH/GSSG], global oxy score) in the iBAT, gene (leptin, Ucp1, Sirt1, Pgc1α and Prdm16) and protein (UCP1) expression in the iBAT and epididymal WAT (eWAT). Key findingsOV group increased body weight gain, Lee index and oxidative stress in the iBAT. Both FLX-treated groups showed less weight gain compared to their controls. OF group showed different leptin expression in the WAT and iBAT; increased functional UCP1 content and mitochondrial activity with less oxidative stress in the iBAT and upregulation of browning genes in eWAT (Pgc1α, Prdm16 and Ucp1). ConclusionAltogether our findings indicated that FLX treatment in young adult overfed animals improved the iBAT mitochondrial function, reduced oxidative stress and induced transcriptional activation of browning agents in white adipose tissue.

UCP2 ameliorates mitochondrial dysfunction, inflammation, and oxidative stress in lipopolysaccharide-induced acute kidney injury

ObjectiveUCP2 is involved in the maintenance of mitochondrial function, immune response and regulation of oxidative stress under physiological or pathological conditions. The aim of this study was to investigate the effects of UCP2 on mitochondrial dysfunction, inflammation, and oxidative stress in septic acute kidney injury (AKI). MethodsWe established LPS-induced AKI model in mice and HK-2 cells. In vivo, the UCP2 inhibitor genipin was used to downregulate UCP2 in mouse kidneys. In vitro, UCP2 overexpression or knockdown was achieved by LV5-UCP2 or si-UCP2 transfection, respectively, to characterize the mechanisms of UCP2 in septic AKI. Indicators of renal injury, cell apoptosis, inflammation, oxidative stress, and mitochondrial dysfunction were assessed. ResultsCompared to the control group, LPS treatment increased UCP2 expression in vitro and in vivo. In vitro, UCP2 overexpression protected HK-2 cells from LPS-induced injury by suppression of apoptosis, inflammation, oxidative stress, MMP loss and ROS production, increase of ATP production and mtDNA content, and amelioration of damage to the mitochondrial ultrastructure. Additionally, inhibition of UCP2 expression by si-UCP2 resulted in decreased HK-2 cell resistance to LPS toxicity, as shown by increased apoptosis, inflammation, mitochondrial dysfunction and oxidative stress. In vivo, UCP2 downregulation aggravated the LPS-induced renal injury, inflammation, macrophages infiltration, mitochondrial dysfunction, and oxidative stress. ConclusionUCP2 may protect LPS-induced AKI by ameliorating mitochondrial dysfunction, anti-inflammation, and antioxidative activities, ultimately inhibiting tubule epithelial cell apoptosis, and that increasing the UCP2 content in mitochondria constitutes a new therapeutic approach for septic AKI.

Age, Sex, and Depot Differences in Adipose Tissue from Obese Subjects

PurposeAdipose tissue (AT) distribution and metabolism are largely affected by the sex hormone, estrogen (E2), presumably through E2 receptors (ER) a and b. In both males and females, E2 biosynthesis in AT occurs through aromatase enzyme, which is responsible for converting androstenedione to estrone. Menopause, loss of ovarian E2, associates with an abrupt increase in metabolic disease risk for reasons that are not fully understood, implicating E2 signaling within AT as playing a role in metabolic health. The purpose of this study was to determine the individual and combined effects of menopause and age on AT depot‐specific immunometabolic characteristics, in morbidly obese, age matched, men and women.Methods34 women (20 Pre‐menopausal (PRE), 14 Post‐menopausal (POST)) and 27 age‐matched men (M) (14 M‐PRE, 13 M‐POST) undergoing bariatric surgery consented to collection of omental (OM) AT; 26 of which consented to collection of abdominal subcutaneous (SQ) AT. Blood samples were collected prior to surgery. AT samples were immediately processed for histological, gene, and protein analyses.Statistics2‐way ANOVA for age group (A), sex (S), and A*S interactions were performed; p&lt;0.05 was considered significant.ResultsBlood glucose, cholesterol, triglycerides, and HDL‐c were similar between groups; whereas, free fatty acids were significantly higher in females than males (1.07±0.04 vs. 0.95±0.05 mmol/L, respectively, p=0.049). OMAT cell size was larger in males than females (6382.0±521.1 vs. 4610.4±521.1 mm2, respectively, p=0.028); SQAT cell size was not different by S or A. SQAT gene expression (but not protein content) of ERb was significantly higher in females than males (1.21±0.22 vs. 0.60 ± 0.12, p=0.049). ERa gene expression was similar between A and S in OM and SQAT but there was a trend toward A*S interaction (p=0.052) in ERa protein in OMAT such that POST &gt; than PRE, M‐PRE, and M‐POST, all p values &lt;0.01. There was an A*S interaction for OMAT gene expression of aromatase (p=0.014); M‐Post &gt; POST (p=0.023) and M‐POST &gt; M‐PRE (p=0.044). The older age group had increased expression of SQAT TNFa (p=0.009). In SQAT, an A*S interaction (p=0.025) occurred in NFkB protein content, such that PRE &lt; POST (p=0.092) and PRE &lt; M‐PRE (p=0.076). Mitochondrial protein of electron transport chain (ETC) complexes (C) in SQAT had significant A*S interactions for C 2 and 3 (p=0.040 and 0.027, respectively). Post hoc differences in SQAT C2 PRE &lt; POST (p=0.043) and C3 POST &gt; PRE (p=0.060) and POST &gt; M‐POST (p=0.065). C1 tended to follow similar patterns between A and S as C 2 and 3, A*S interaction p=0.055. OMAT ETC protein content was similar between A and S. An A*S trend (p=0.063) in SQ UCP1 content was observed such that POST &gt; PRE (p=0.051) and POST &gt; M‐POST (p=0.062); OMAT UCP1 was similar by A and S.ConclusionIn morbid obesity, loss of E2 and aging may differentially affect OMAT E2 signaling pathways in men and women. Conversely, loss of E2 and aging affect SQAT mitochondrial and inflammatory pathways.Support or Funding InformationResearch supported by Dr. Kanaley RO1DK101513This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

ACE I/D gene variant predicts ACE enzyme content in blood but not the ACE, UCP2, and UCP3 protein content in human skeletal muscle in the Gene SMART study.

Angiotensin-converting enzyme (ACE) is expressed in human skeletal muscle. The ACE I/D polymorphism has been associated with athletic performance in some studies. Studies have suggested that the ACE I/D gene variant is associated with ACE enzyme content in serum, and there is an interaction between ACE and uncoupling proteins 2 and 3 (UCP2 and UCP3). However, no studies have explored the effect of ACE I/D on ACE, UCP2, and UCP3 protein content in human skeletal muscle. Utilizing the Gene SMART cohort ( n = 81), we investigated whether the ACE I/D gene variant is associated with ACE enzyme content in blood and ACE, UCP2, and UCP3 protein content in skeletal muscle at baseline and following a session of high-intensity interval exercise (HIIE). Using a stringent and robust statistical analyses, we found that the ACE I/D gene variant was associated with ACE enzyme content in blood ( P < 0.005) at baseline but not the ACE, UCP2, and UCP3 protein content in muscle at baseline. A single session of HIIE tended (0.005 < P < 0.05) to increase blood ACE content immediately postexercise, whereas muscle ACE protein content was lower 3 h after a single session of HIIE ( P < 0.005). Muscle UCP3 protein content decreased immediately after a single session of HIIE ( P < 0.005) and remained low 3 h postexercise. However, those changes in the muscle were not genotype dependent. In conclusion, The ACE I/D gene variant predicts ACE enzyme content in blood but not the ACE, UCP2, and UCP3 protein content of human skeletal muscle. NEW & NOTEWORTHY This paper describes the association between ACE I/D gene variant and ACE protein content in blood and ACE, UCP2, and UCP3 protein content in skeletal muscle at baseline and after exercise in a large cohort of healthy males. Our data suggest that ACE I/D is a strong predictor of blood ACE content but not muscle ACE content.

The nuclear retinoid-related orphan receptor RORα controls circadian thermogenic programming in white fat depots.

The ROR α‐deficient staggerer (sg/sg) mouse is lean and resistant to diet‐induced obesity. Its thermogenic activity was shown to be increased not only in brown adipose tissue (BAT), but also in subcutaneous white adipose tissue (WAT) where UCP1 content was enhanced, however, without Prdm16 coexpression. Our observation of partial multilocular lipid morphology of WAT in sg/sg mice both in the inguinal and perigonadal sites led us to focus on the phenotype of both fat depots. Because ROR α is a nuclear factor acting in the clock machinery, we looked at the circadian expression profile of genes involved in thermogenesis and browning in WAT and BAT depots of sg/sg and WT mice, through real‐time quantitative PCR and western blotting. This 24‐h period approach revealed both a rhythmic expression of thermogenic genes in WAT and an increased browning of all the WAT depots tested in sg/sg mice that indeed involved the canonical browning process (through induction of Pgc‐1α and Prdm16). This was associated with an enhanced isoproterenol‐induced oxygen consumption rate of WAT explants from sg/sg mice, which was reproducible in WT explants by treatment with a ROR α inverse agonist SR 3335, that induced a parallel increase in the UCP1 protein. Inhibitors of browning differentiation, such as TLE3 and RIP140, could be new targets of ROR α that would be rather implicated in the whitening of adipocytes. Our study showed the pivotal role of ROR α as an inhibitor of the thermogenic program in WAT, the role that could be counteracted in vivo with the ROR α antagonists currently in development.

Replacing maize silage with red clover silage in total mixed rations for dairy cows: In vitro ruminal fermentation characteristics and associative effects

The objective of this study was to evaluate fermentation characteristics and associative effects by replacing maize silage (MS) with red clover silage (RCS) using in vitro techniques. Treatments included total mixed rations (TMR) containing forage (MS and RCS) and concentrates (0.75:0.25) with targeted proportions of RCS in TMR of 0.15 (RCS15), 0.30 (RCS30), 0.45 (RCS45), and 0.60 (RCS60) on a dry matter (DM) basis. The Hohenheim gas test (HGT) was used to describe the kinetic of gas production. The modified HGT was used to estimate the utilizable crude protein at the duodenum (uCP). Effective uCP was calculated according to passage rates of 2, 5 and 8%/h (uCP2, uCP5 and uCP8, respectively). Associative effects were defined as the differences (%) between the observed values of TMR and the sum of the observed values of the individual components of the TMR. The gas production from incubation time 8 to 72h, the potential gas production (A), organic matter digestibility (OMD), and metabolizable energy (ME) decreased with increasing levels of RCS. The effective uCP2 content was similar between TMR (average 93.7g/kg DM). However, uCP5 and uCP8 were reduced with increasing level of RCS (from 145 to 136g/kg DM, and from 172 to 157g/kg DM). Positive associative effect was found for cumulative gas production. But this effect was reduced when increasing incubation time and when increasing levels of RCS. Associative effects of gas production at 24h of incubation averaged 1.35%. Consequently, those of ME (−0.09 to 1.01%) and OMD (1.60 to 2.76%) were of minor relevance as well. Associative effects on uCP depended on rate of passage and on level of RCS. Within TMR, associative effects on uCP increased with increasing rate of passage (e.g. for RCS15; 3.67 and 5.39% for 2 and 8%/h, respectively). Within rate of passage, associative effect was reduced with increasing level of RCS (e.g. for uCP8; 5.39 and −0.95% for RCS15 and RCS60, respectively). Based on associative effects of fermentation characteristics and estimated uCP content, this study suggests that incrementing levels of RCS may lead to a reduction on the feed value of diets, and that the optimal proportion of RCS:MS in TMR to achieve beneficial effects was 0.15:0.60 on DM basis. Therefore, caution should be taken before introducing high levels of RCS in rations of high-yielding dairy cows. However, these results should not be directly extrapolated to in vivo conditions.

Protective effects of PGC-1α via the mitochondrial pathway in rat brains after intracerebral hemorrhage

Peroxisome-proliferator-activated receptor co-activator-1α (PGC-1α) is a transcriptional co-activator that coordinately regulates genes required for mitochondrial biogenesis, which stimulates mitochondrial activity. It is also a major factor in the up-regulation of antioxidant activities that are a response to oxidative stress. However, the role of PGC-1α after intracerebral hemorrhage (ICH) has not been studied. The purpose of the present work was to investigate the effects and mechanism of PGC-1α after ICH in the brain.Brain injury was induced by injecting autologous arterial blood (50μL) into the rat brain. PGC-1α siRNAs were injected into rat brains 24h prior to ICH. Then, 72h after ICH, brains were collected for investigation. Post-assessment included western blot analysis, RT-PCR assay, neurobehavioral function testing, measurement of brain water content, high-performance liquid chromatography (HPLC), and projection electron microscopy on ICH rat models. The concentration of PGC-1α was higher in the ipsilateral striatum after ICH, peaking around 72h after ICH. The expression of NRF-1, TFAM, SOD2, UCP2, mitochondrial DNA, ATP concentration, mitochondrial quantity, and brain water content were increased 72h after ICH. However, the neurological score was decreased 72h after ICH. Treatment with PGC-1α siRNAs significantly decreased the neurological score, ATP concentration, number of mitochondria, expression of NRF-1, TFAM, SOD2, UCP2, and mitochondrial DNA, and increased brain water content and formation of mitochondrial myelin layer structures. In conclusion, our data suggest that PGC-1α protects rat brains via a mitochondrial pathway following ICH. Key words: PGC-1α intracerebral hemorrhage(ICH); mitochondrial; neuroprotection.

Constitutive adipocyte mTORC1 activation enhances mitochondrial activity and reduces visceral adiposity in mice

Mechanistic target of rapamycin complex 1 (mTORC1) loss of function reduces adiposity whereas partial mTORC1 inhibition enhances fat deposition. Herein we evaluated how constitutive mTORC1 activation in adipocytes modulates adiposity in vivo. Mice with constitutive mTORC1 activation in adipocytes induced by tuberous sclerosis complex (Tsc)1 deletion and littermate controls were evaluated for body mass, energy expenditure, glucose and fatty acid metabolism, mitochondrial function, mRNA and protein contents. Adipocyte-specific Tsc1 deletion reduced visceral, but not subcutaneous, fat mass, as well as adipocyte number and diameter, phenotypes that were associated with increased lipolysis, UCP-1 content (browning) and mRNA levels of pro-browning transcriptional factors C/EBPβ and ERRα. Adipocyte Tsc1 deletion enhanced mitochondrial oxidative activity, fatty acid oxidation and the expression of PGC-1α and PPARα in both visceral and subcutaneous fat. In brown adipocytes, however, Tsc1 deletion did not affect UCP-1 content and basal respiration. Adipocyte Tsc1 deletion also reduced visceral adiposity and enhanced glucose tolerance, liver and muscle insulin signaling and adiponectin secretion in mice fed with purified low- or high-fat diet. In conclusion, adipocyte-specific Tsc1 deletion enhances mitochondrial activity, induces browning and reduces visceral adiposity in mice.