Mitophagy suppression by miquelianin-rich lotus leaf extract induces 'beiging' of white fat via AMPK/DRP1-PINK1/PARKIN signaling axis.

BACKGROUND: Lotus (Nelumbo nucifera) leaf has been described to have anti-obesity activity, but the role of white fat 'browning' or 'beiging' in its beneficial metabolic actions remains unclear. Here, 3T3-L1 cells and high-fat-diet (HFD)-fed mice were used to evaluate the effects of miquelianin-rich lotus leaf extract (LLE) on white-to-beige fat conversion and its regulatory mechanisms. RESULTS: Treatment with LLE increased mitochondrial abundance, mitochondrial membrane potential and NAD+ /NADH ratio in 3T3-L1 cells, suggesting its potential in promoting mitochondrial activity. qPCR and/or western blotting analysis confirmed that LLE induced the expression of beige fat-enriched gene signatures (e.g. Sirt1, Cidea, Dio2, Prdm16, Ucp1, Cd40, Cd137, Cited1) and mitochondrial biogenesis-related markers (e.g. Nrf1, Cox2, Cox7a, Tfam) in 3T3-L1 cells and inguinal white adipose tissue of HFD-fed mice. Furthermore, we found that LLE treatment inhibited mitochondrial fission protein DRP1 and blocked mitophagy markers such as PINK1, PARKIN, BECLIN1 and LC-3B. Chemical inhibition experiments revealed that AMPK/DRP1 signaling was required for LLE-induced beige fat formation via suppressing PINK1/PARKIN/mitophagy. CONCLUSION: Our data reveal a novel mechanism underlying the anti-obesity effect of LLE, namely the induction of white fat beiging via AMPK/DRP1/mitophagy signaling. © 2023 Society of Chemical Industry.

Metal Poly(heptazine imides) as Multifunctional Photocatalysts for Solar Fuel Production.

Solar-driven photocatalysis employing particulate semiconductors represents a promising approach for sustainable production of valuable chemical feedstock. Metal poly(heptazine imide) (MPHI), a novel 2D ionic carbon nitride, has been recognized as an emerging photocatalyst with distinctive properties. In this minireview, we first delineate the forefront innovations of MPHI photocatalysts, spanning from synthetic strategies and solving structures to the exploration of novel properties. We place special emphasis on the structural design principles aimed at developing high-performance MPHI systems toward photocatalytic solar fuel production such as H2 evolution, H2O oxidation, H2O2 production and CO2 reduction. Finally, we discuss crucial insights and challenges in leveraging highly active MPHIs for efficient solar-to-chemical energy conversion.

The impact of metabolic endotoxaemia on the browning process in human adipocytes.

BACKGROUND: Dysfunctional adipose tissue (AT) is known to contribute to the pathophysiology of metabolic disease, including type 2 diabetes mellitus (T2DM). This dysfunction may occur, in part, as a consequence of gut-derived endotoxaemia inducing changes in adipocyte mitochondrial function and reducing the proportion of BRITE (brown-in-white) adipocytes. Therefore, the present study investigated whether endotoxin (lipopolysaccharide; LPS) directly contributes to impaired human adipocyte mitochondrial function and browning in human adipocytes, and the relevant impact of obesity status pre and post bariatric surgery. METHODS: Human differentiated abdominal subcutaneous (AbdSc) adipocytes from participants with obesity and normal-weight participants were treated with endotoxin to assess in vitro changes in mitochondrial function and BRITE phenotype. Ex vivo human AbdSc AT from different groups of participants (normal-weight, obesity, pre- and 6 months post-bariatric surgery) were assessed for similar analyses including circulating endotoxin levels. RESULTS: Ex vivo AT analysis (lean & obese, weight loss post-bariatric surgery) identified that systemic endotoxin negatively correlated with BAT gene expression (p < 0.05). In vitro endotoxin treatment of AbdSc adipocytes (lean & obese) reduced mitochondrial dynamics (74.6% reduction; p < 0.0001), biogenesis (81.2% reduction; p < 0.0001) and the BRITE phenotype (93.8% reduction; p < 0.0001). Lean AbdSc adipocytes were more responsive to adrenergic signalling than obese AbdSc adipocytes; although endotoxin mitigated this response (92.6% reduction; p < 0.0001). CONCLUSIONS: Taken together, these data suggest that systemic gut-derived endotoxaemia contributes to both individual adipocyte dysfunction and reduced browning capacity of the adipocyte cell population, exacerbating metabolic consequences. As bariatric surgery reduces endotoxin levels and is associated with improving adipocyte functionality, this may provide further evidence regarding the metabolic benefits of such surgical interventions.

Food phenolics stimulate adipocyte browning via regulating gut microecology.

Fat browning has piqued the interest of researchers as a potential target for treating obesity and related metabolic disorders. Recruitment of brown adipocytes leads to enhanced energy dissipation and reduced adiposity, thus facilitating the maintenance of metabolic homeostasis. Evidence is increasing to support the crucial roles of polyphenols and gut microecology in turning fat "brown". However, it is not clear whether the intestinal microecology is involved in polyphenol-mediated regulation of adipose browning, so this concept is worthy of exploration. In this review, we summarize the current knowledge, mostly from studies with murine models, supporting the concept that the effects of food phenolics on brown fat activation and white fat browning can be attributed to their regulatory actions on gut microecology, including microbial community profile, gut metabolites, and gut-derived hormones. Furthermore, the potential underlying pathways involved are also discussed. Basically, understanding gut microecology paves the way to determine the underlying roles and mechanisms of food phenolics in adipose browning.

Alkaloids from lotus (Nelumbo nucifera): recent advances in biosynthesis, pharmacokinetics, bioactivity, safety, and industrial applications.

Different parts of lotus (Nelumbo nucifera Gaertn.) including the seeds, rhizomes, leaves, and flowers, are used for medicinal purposes with health promoting and illness preventing benefits. The presence of active chemicals such as alkaloids, phenolic acids, flavonoids, and terpenoids (particularly alkaloids) may account for this plant's pharmacological effects. In this review, we provide a comprehensive overview and summarize up-to-date research on the biosynthesis, pharmacokinetics, and bioactivity of lotus alkaloids as well as their safety. Moreover, the potential uses of lotus alkaloids in the food, pharmaceutical, and cosmetic sectors are explored. Current evidence shows that alkaloids, mainly consisting of aporphines, 1-benzylisoquinolines, and bisbenzylisoquinolines, are present in different parts of lotus. The bioavailability of these alkaloids is relatively low in vivo but can be enhanced by technological modification using nanoliposomes, liposomes, microcapsules, and emulsions. Available data highlights their therapeutic and preventive effects on obesity, diabetes, neurodegeneration, cancer, cardiovascular disease, etc. Additionally, industrial applications of lotus alkaloids include their use as food, medical, and cosmetic ingredients in tea, other beverages, and healthcare products; as lipid-lowering, anticancer, and antipsychotic drugs; and in facial masks, toothpastes, and shower gels. However, their clinical efficacy and safety remains unclear; hence, larger and longer human trials are needed to achieve their safe and effective use with minimal side effects.

Metabolic responses of light and taste receptors - unexpected actions of GPCRs in adipocytes.

The G-protein-coupled receptor (GPCR) superfamily includes sensory receptors that can detect and respond to taste and light. Recent investigations have identified key metabolic roles for such receptors in tissues considered 'non-sensory' such as adipose tissue. The major functions of white and brown adipose tissues include energy storage/release and thermogenesis, respectively. These processes are tightly controlled by GPCR pathways that serve to maintain energy homeostasis. Opsins 3 and 4 are GPCRs activated by blue light and in adipocytes control lipolysis as well as affect brown adipocyte activity. Furthermore, Opsin 3 signals to regulate the conversion of white to thermogenic beige/BRITE (Brown-in-white) adipocytes. Taste receptors that respond to fatty acids, sweet and bitter are expressed in adipocytes as well as in taste buds. Ffar2 and the long chain fatty acid receptor GPR120 are highly expressed in white adipocytes and the human tongue. In adipose tissue Ffar2 mediates the metabolic effects of butyrate and propionate produced by the gut microbiome. GPR120 is highly expressed in brown adipose tissue and regulates fatty acid oxidation and mitochondrial function. The type I taste receptor Tas1r3 senses sweet and umami, is expressed in adipocytes and on obesogenic diets Tas1r3 global gene knockout protects from metabolic dysfunction. Type II taste receptors that sense bitter are expressed by adipocytes and bitter agonists have been found to modulate adipocyte differentiation and lipid storage levels. This review explores recent unexpected findings of light and taste receptors in adipocytes and examines effects of their signaling in the control of adipose tissue biology.

Smad4 promotes diabetic nephropathy by modulating glycolysis and OXPHOS.

Diabetic nephropathy (DN) is the leading cause of end-stage kidney disease. TGF-ß1/Smad3 signalling plays a major pathological role in DN; however, the contribution of Smad4 has not been examined. Smad4 depletion in the kidney using anti-Smad4 locked nucleic acid halted progressive podocyte damage and glomerulosclerosis in mouse type 2 DN, suggesting a pathogenic role of Smad4 in podocytes. Smad4 is upregulated in human and mouse podocytes during DN. Conditional Smad4 deletion in podocytes protects mice from type 2 DN, independent of obesity. Mechanistically, hyperglycaemia induces Smad4 localization to mitochondria in podocytes, resulting in reduced glycolysis and oxidative phosphorylation and increased production of reactive oxygen species. This operates, in part, via direct binding of Smad4 to the glycolytic enzyme PKM2 and reducing the active tetrameric form of PKM2. In addition, Smad4 interacts with ATPIF1, causing a reduction in ATPIF1 degradation. In conclusion, we have discovered a mitochondrial mechanism by which Smad4 causes diabetic podocyte injury.

A Four Carbon Organonitrate as a Significant Product of Secondary Isoprene Chemistry.

Oxidation of isoprene by nitrate radicals (NO3) or by hydroxyl radicals (OH) under high NOx conditions forms a substantial amount of organonitrates (ONs). ONs impact NOx concentrations and consequently ozone formation while also contributing to secondary organic aerosol. Here we show that the ONs with the chemical formula C4H7NO5 are a significant fraction of isoprene-derived ONs, based on chamber experiments and ambient measurements from different sites around the globe. From chamber experiments we found that C4H7NO5 isomers contribute 5%-17% of all measured ONs formed during nighttime and constitute more than 40% of the measured ONs after further daytime oxidation. In ambient measurements C4H7NO5 isomers usually dominate both nighttime and daytime, implying a long residence time compared to C5 ONs which are removed more rapidly. We propose potential nighttime sources and secondary formation pathways, and test them using a box model with an updated isoprene oxidation scheme.

Fibrotic Phenotype of Peritumour Mesenteric Adipose Tissue in Human Colon Cancer: A Potential Hallmark of Metastatic Properties.

The impact of tumour associated stroma on cancer metastasis is an emerging field. However, cancer associated genes in peritumoral adipose tissue (pAT) in human colon cancer have not been explored. The aim of this study was to identify differentially expressed genes (DEGs) associated with cancer pathways in mesenteric pAT compared with adjacent adipose tissue. In total, nine patients with colon cancer pathological stage T2/T4 were employed in this study. DEGs were identified in 6 patients employing Nanostring PanCancer Pathway Panel and pathway enrichment analyses were performed. Differential expression of the 5 most up-regulated and 2 down regulated genes was validated with qRT-PCR. Results showed collagen type I alpha 1 chain (COL1A1) p = 0.007; secreted frizzled related protein (SFRP2) p = 0.057; fibroblast growth factor 7 (FGF7) not significant (ns); phospholipase A2, group IIA (PLA2G2A) ns; nerve growth factor receptor (NGFR) ns; lymphoid enhancer binding factor 1 (LEF1) p = 0.03; cadherin 1, Type 1, E-cadherin (epithelial) (CDH1) 0.09. Results have highlighted down-regulation of the Wingless/Integrated (Wnt) pathway in mesenteric pAT compared to distal adipose tissue. Highly upregulated genes in mesenteric pAT were involved in extracellular matrix (ECM)-receptor interactions and focal adhesion. Highly down regulated genes were involved in the cell cycle. Immunohistochemistry revealed differential distribution of COL1A1 showing maximum levels in tumour tissue and gradually decreasing in distant adipose tissue. COL1A1 and down regulation of Wnt pathway may have a role in local invasion and distant metastasis. COL1A1 may represent a stromal prognostic biomarker and therapeutic target in colon cancer.

Elucidation of the roles of brown and brite fat genes: GPR120 is a modulator of brown adipose tissue function.

NEW FINDINGS: What is the topic of this review? Activation of brown adipose tissue with G protein-coupled receptors as key druggable targets as a strategy to increase energy consumption and reduce fat mass. What advances does it highlight? GPR120 is a fatty acid receptor highly expressed in brown adipose tissue. Its activation by selective ligands increases brown adipose tissue activity. This is mediated by changes in mitochondrial dynamics resulting in increased O2 consumption leading to enhanced nutrient uptake and a reduction in fat mass. ABSTRACT: The identification of druggable targets to stimulate brown adipose tissue (BAT) is a strategy to combat obesity due to this highly metabolically active tissue utilising thermogenesis to burn fat. Upon cold exposure BAT is activated by the sympathetic nervous system via ß3 -adrenergic receptors. Determination of additional receptors expressed by brown, white and brite (brown-in-white) fat can lead to new pharmacological treatments to activate BAT. GPR120 is a G protein-coupled fatty acid receptor that is highly expressed in BAT and further increases in response to cold. Activation of this receptor with the selective agonist TUG-891 acutely increases fat oxidation and reduces fat mass in mice. The effects are coincident with increased BAT activity and enhanced nutrient uptake. TUG-891 stimulation of brown adipocytes induces intracellular Ca2+ release which results in elevated O2 consumption as well as mitochondrial depolarisation and fission. Thus, activation of GPR120 in BAT with ligands such as TUG-891 is a promising strategy to increase fat consumption.

Androgen Reduces Mitochondrial Respiration in Mouse Brown Adipocytes: A Model for Disordered Energy Balance in Polycystic Ovary Syndrome.

Polycystic ovary syndrome (PCOS) is a common endocrinopathy that is associated with an adverse metabolic profile including reduced postprandial thermogenesis. Although abnormalities in adipose tissue function have been widely reported in women with PCOS, less is known about direct effects of androgen on white and, particularly, brown adipocytes. The purpose of this study was to investigate the effect of the nonaromatizable androgen dihydrotestosterone (DHT) on (1) lipid accumulation and expression of adipogenic markers in immortalized mouse brown adipose cell lines (IMBATs), (2) mitochondrial respiration in IMBATs, (3) mitochondrial DNA content and gene expression, (4) expression of brown adipose tissue (BAT) markers and thermogenic activation. In addition, we profiled the relative levels of 38 adipokines secreted from BAT explants and looked at androgen effects on adipokine gene expression in both IMBATs and immortalized mouse white adipose (IMWATs) cell lines. Androgen treatment inhibited IMBAT differentiation in a dose-dependent manner, reduced markers of adipogenesis, and attenuated the ß-adrenoceptor-stimulated increase in uncoupling protein-1 (UCP1) expression. In explants of mouse interscapular BAT, androgen reduced expression of UCP1, peroxisome proliferator-activated receptor-γ coactivator-1 (PCG-1) and Cidea. Significantly, as well as affecting genes involved in thermogenesis in BAT, androgen treatment reduced mitochondrial respiration in IMBATs, as measured by the Seahorse XF method. The results of this study suggest a role for excess androgen in inhibiting brown adipogenesis, attenuating the activation of thermogenesis and reducing mitochondrial respiration in BAT. Together, these data provide a plausible molecular mechanism that may contribute to reduced postprandial thermogenesis and the tendency to obesity in women with PCOS.

Indoor ozone/human chemistry and ventilation strategies.

This study aimed to better understand and quantify the influence of ventilation strategies on occupant-related indoor air chemistry. The oxidation of human skin oil constituents was studied in a continuously ventilated climate chamber at two air exchange rates (1 h-1 and 3 h-1 ) and two initial ozone mixing ratios (30 and 60 ppb). Additional measurements were performed to investigate the effect of intermittent ventilation ("off" followed by "on"). Soiled t-shirts were used to simulate the presence of occupants. A time-of-flight-chemical ionization mass spectrometer (ToF-CIMS) in positive mode using protonated water clusters was used to measure the oxygenated reaction products geranyl acetone, 6-methyl-5-hepten-2-one (6-MHO) and 4-oxopentanal (4-OPA). The measurement data were used in a series of mass balance models accounting for formation and removal processes. Reactions of ozone with squalene occurring on the surface of the t-shirts are mass transport limited; ventilation rate has only a small effect on this surface chemistry. Ozone-squalene reactions on the t-shirts produced gas-phase geranyl acetone, which was subsequently removed almost equally by ventilation and further reaction with ozone. About 70% of gas-phase 6-MHO was produced in surface reactions on the t-shirts, the remainder in secondary gas-phase reactions of ozone with geranyl acetone. 6-MHO was primarily removed by ventilation, while further reaction with ozone was responsible for about a third of its removal. 4-OPA was formed primarily on the surfaces of the shirts (~60%); gas-phase reactions of ozone with geranyl acetone and 6-MHO accounted for ~30% and ~10%, respectively. 4-OPA was removed entirely by ventilation. The results from the intermittent ventilation scenarios showed delayed formation of the reaction products and lower product concentrations compared to continuous ventilation.

In Vitro Models for Study of Brown Adipocyte Biology.

Brown adipocytes are the key cell type in brown adipose tissue (BAT) that express the genes required for heat production through the process of thermogenesis. Brown adipocyte cell culture models are important for researching the molecular pathways that control cell autonomous processes. In vitro tools for the study of brown adipocytes include BAT explant cultures and BAT primary cultures that are first proliferated and then differentiated. A number of stable brown preadipocyte cell lines have been generated by the expression transforming factors such as SV40 T antigen. The application of these cell lines reduces the requirement for animal tissue which is needed for primary culture and explants. Furthermore, brown adipocyte cell lines that effectively recapitulate the properties of brown adipocytes permit large-scale experimental procedures that are generally unfeasible with primary cultures that undergo a restricted number of cell divisions. Cell lines are valuable for applications such as large-scale endogenous protein expression, ChIP assay, and procedures requiring antibiotic selection over several cell divisions including stable exogenous gene expression and CRISR/Cas9 gene editing.

Cdkn1c Boosts the Development of Brown Adipose Tissue in a Murine Model of Silver Russell Syndrome.

The accurate diagnosis and clinical management of the growth restriction disorder Silver Russell Syndrome (SRS) has confounded researchers and clinicians for many years due to the myriad of genetic and epigenetic alterations reported in these patients and the lack of suitable animal models to test the contribution of specific gene alterations. Some genetic alterations suggest a role for increased dosage of the imprinted CYCLIN DEPENDENT KINASE INHIBITOR 1C (CDKN1C) gene, often mutated in IMAGe Syndrome and Beckwith-Wiedemann Syndrome (BWS). Cdkn1c encodes a potent negative regulator of fetal growth that also regulates placental development, consistent with a proposed role for CDKN1C in these complex childhood growth disorders. Here, we report that a mouse modelling the rare microduplications present in some SRS patients exhibited phenotypes including low birth weight with relative head sparing, neonatal hypoglycemia, absence of catch-up growth and significantly reduced adiposity as adults, all defining features of SRS. Further investigation revealed the presence of substantially more brown adipose tissue in very young mice, of both the classical or canonical type exemplified by interscapular-type brown fat depot in mice (iBAT) and a second type of non-classic BAT that develops postnatally within white adipose tissue (WAT), genetically attributable to a double dose of Cdkn1c in vivo and ex-vivo. Conversely, loss-of-function of Cdkn1c resulted in the complete developmental failure of the brown adipocyte lineage with a loss of markers of both brown adipose fate and function. We further show that Cdkn1c is required for post-transcriptional accumulation of the brown fat determinant PR domain containing 16 (PRDM16) and that CDKN1C and PRDM16 co-localise to the nucleus of rare label-retaining cell within iBAT. This study reveals a key requirement for Cdkn1c in the early development of the brown adipose lineages. Importantly, active BAT consumes high amounts of energy to generate body heat, providing a valid explanation for the persistence of thinness in our model and supporting a major role for elevated CDKN1C in SRS.

Hydroxysteroid dehydrogenase family proteins on lipid droplets through bacteria, C. elegans, and mammals.

Lipid droplets (LDs) are the main fat storing sites in almost all species from bacteria to humans. The perilipin family has been found as LD proteins in mammals, Drosophila, and a couple of slime molds, but no bacterial LD proteins containing sequence conservation were identified. In this study, we reported that the hydroxysteroid dehydrogenase (HSD) family was found on LDs across all organisms by LD proteomic analysis. Imaging experiments confirmed LD targeting of three representative HSD proteins including ro01416 in RHA1, DHS-3 in C. elegans, and 17ß-HSD11 in human cells. In C. elegans, 17ß-HSD11 family proteins (DHS-3, DHS-4 and DHS-19) were localized on LDs in distinct tissues. In intestinal cells of C. elegans, DHS-3 targeted to cytoplasmic LDs, while DHS-9 labeled nuclear LDs. Furthermore, the N-terminal hydrophobic domains of 17ß-HSD11 family were necessary for their targeting to LDs. Last, 17ß-HSD11 family proteins induced LD aggregation, and deletion of DHS-3 in C. elegans caused lipid decrease. Independent of their presumptive catalytic sites, 17ß-HSD11 family proteins regulated LD dynamics and lipid metabolism through affecting the LD-associated ATGL, which was conserved between C. elegans and humans. Together, these findings for HSDs provide a new insight not only into the mechanistic studies of the dynamics and functions of LDs in multiple organisms, but also into understanding the evolutionary history of the organelle.

Diacylglycerol acyltransferase 2 links glucose utilization to fatty acid oxidation in the brown adipocytes.

Brown adipose tissue uptake of glucose and fatty acids is very high during nonshivering thermogenesis. Adrenergic stimulation markedly increases glucose uptake, de novo lipogenesis, and FA oxidation simultaneously. The mechanism that enables this concerted response has hitherto been unknown. Here, we find that in primary brown adipocytes and brown adipocyte-derived cell line (IMBAT-1), acute inhibition and longer-term knockdown of DGAT2 links the increased de novo synthesis of fatty acids from glucose to a pool of TAG that is simultaneously hydrolyzed, providing FA for mitochondrial oxidation. DGAT1 does not contribute to this pathway, but uses exogenous FA and glycerol to synthesize a functionally distinct pool of TAG to which DGAT2 also contributes. The DGAT2-dependent channelling of 14C from glucose into TAG and CO2 was reproduced in ß3-agonist-stimulated primary brown adipocytes. Knockdown of DGAT2 in IMBAT-1 affected the mRNA levels of UCP1 and genes important in FA activation and esterification. Therefore, in ß3-agonist activated brown adipocytes, DGAT2 specifically enables channelling of de novo synthesized FA into a rapidly mobilized pool of TAG, which is simultaneously hydrolyzed to provide substrates for mitochondrial fatty acid oxidation.

The K+ channel TASK1 modulates ß-adrenergic response in brown adipose tissue through the mineralocorticoid receptor pathway.

Brown adipose tissue (BAT) is essential for adaptive thermogenesis and dissipation of caloric excess through the activity of uncoupling protein (UCP)-1. BAT in humans is of great interest for the treatment of obesity and related diseases. In this study, the expression of Twik-related acid-sensitive K(+) channel (TASK)-1 [a pH-sensitive potassium channel encoded by the potassium channel, 2-pore domain, subfamily K, member 3 (Kcnk3) gene] correlated highly with Ucp1 expression in obese and cold-exposed mice. In addition, Task1-null mice, compared with their controls, became overweight, mainly because of an increase in white adipose tissue mass and BAT whitening. Task1(-/-)-mouse-derived brown adipocytes, compared with wild-type mouse-derived brown adipocytes, displayed an impaired ß3-adrenergic receptor response that was characterized by a decrease in oxygen consumption, Ucp1 expression, and lipolysis. This phenotype was thought to be caused by an exacerbation of mineralocorticoid receptor (MR) signaling, given that it was mimicked by corticoids and reversed by an MR inhibitor. We concluded that the K(+) channel TASK1 controls the thermogenic activity in brown adipocytes through modulation of ß-adrenergic receptor signaling.

Lipid droplet remodeling and interaction with mitochondria in mouse brown adipose tissue during cold treatment.

Brown adipose tissue (BAT) maintains animal body temperature by non-shivering thermogenesis, which is through uncoupling protein 1 (UCP1) that uncouples oxidative phosphorylation and utilizes ß-oxidation of fatty acids released from triacylglycerol (TAG) in lipid droplets (LDs). Increasing BAT activity and "browning" other tissues such as white adipose tissue (WAT) can enhance the expenditure of excess stored energy, and in turn reduce prevalence of metabolic diseases. Although many studies have characterized the biology of BAT and brown adipocytes, BAT LDs especially their activation induced by cold exposure remain to be explored. We have isolated LDs from mouse interscapular BAT and characterized the full proteome using mass spectrometry. Both morphological and biochemical experiments showed that the LDs could tightly associate with mitochondria. Under cold treatment mouse BAT started expressing LD structure protein PLIN-2/ADRP and increased expression of PLIN1. Both hormone sensitive lipase (HSL) and adipose TAG lipase (ATGL) were increased in LDs. In addition, isolated BAT LDs showed increased levels of the mitochondrial protein UCP1, and prolonged cold exposure could stimulate BAT mitochondrial cristae biogenesis. These changes were in agreement with the data from transcriptional analysis. Our results provide the BAT LD proteome for the first time and show that BAT LDs facilitate heat production by coupling increasing TAG hydrolysis through recruitment of ATGL and HSL to the organelle and expression of another LD resident protein PLIN2/ADRP, as well as by tightly associating with activated mitochondria. These findings will benefit the study of BAT activation and the interaction between LDs and mitochondria.

Activation of the Wnt/ß-catenin pathway represses the transcription of the ß-amyloid precursor protein cleaving enzyme (BACE1) via binding of T-cell factor-4 to BACE1 promoter.

Alterations in the Wnt signaling pathway have been implicated in Alzheimer's disease; however, its role in the processing of the amyloid precursor protein remains unknown. In this study, activation of the Wnt pathway by overexpression of the agonist Wnt3a or ß-catenin or by inhibition of glycogen kinase synthase-3 in N2a cells resulted in a reduction in Aß levels and in the activity and expression of BACE1 (ß-APP cleaving enzyme). Conversely, inhibition of the pathway by transfection of the antagonists secreted frizzled receptor protein-1 or dickkopf-1 produced the opposite effects. Chromatin immunoprecipitation analysis demonstrated that ß-catenin binds specifically to regions within the promoter of BACE1 containing putative T-cell factor/lymphoid enhancer binding factor-1 (TCF/LEF) motifs, consistent with canonical Wnt target regulation. Furthermore, cells transfected with ß-catenin mutants incapable of binding to TCF/LEF increased BACE1 gene promoter activity. Interestingly, TCF4 knockdown reversed the effects of Wnt3a activation on BACE1 transcription. We found that TCF4 binds to the same region on BACE1 promoter following Wnt3a stimulation, indicating that TCF4 functions as a transcriptional repressor of BACE1 gene. In conclusion, Wnt/ß-catenin stimulation may repress BACE1 transcription via binding of TCF4 to BACE1 gene, and therefore, activation of the Wnt pathway may hold the key to new treatments of Alzheimer disease.-Parr, C., Mirzaei, N., Christian, M., and Sastre, M. Activation of the Wnt/ß-catenin pathway represses the transcription of the ß-amyloid precursor protein cleaving enzyme (BACE1) via binding of T-cell factor-4 to BACE1 promoter.

Arachidonic acid-dependent gene regulation during preadipocyte differentiation controls adipocyte potential.

Arachidonic acid (AA) is a major PUFA that has been implicated in the regulation of adipogenesis. We examined the effect of a short exposure to AA at different stages of 3T3-L1 adipocyte differentiation. AA caused the upregulation of fatty acid binding protein 4 (FABP4/aP2) following 24 h of differentiation. This was mediated by the prostaglandin F(2α) (PGF(2α)), as inhibition of cyclooxygenases or PGF(2α) receptor signaling counteracted the AA-mediated aP2 induction. In addition, calcium, protein kinase C, and ERK are all key elements of the pathway through which AA induces the expression of aP2. We also show that treatment with AA during the first 24 h of differentiation upregulates the expression of the transcription factor Fos-related antigen 1 (Fra-1) via the same pathway. Finally, treatment with AA for 24 h at the beginning of the adipocyte differentiation is sufficient to inhibit the late stages of adipogenesis through a Fra-1-dependent pathway, as Fra-1 knockdown rescued adipogenesis. Our data show that AA is able to program the differentiation potential of preadipocytes by regulating gene expression at the early stages of adipogenesis.