Capsaicin induces ATP-dependent thermogenesis via the activation of TRPV1/ß3-AR/α1-AR in 3T3-L1 adipocytes and mouse model.

Capsaicin (CAP) is a natural bioactive compound in chili pepper that activates the transient receptor potential vanilloid subfamily 1 (TRPV1) and is known to stimulate uncoupling protein 1 (UCP1)-dependent thermogenesis. However, its effect on ATP-dependent thermogenesis remains unknown. In this study, we employed qRT-PCR, immunoblot, staining method, and assay kit to investigate the role of CAP on ATP-dependent thermogenesis and its modulatory roles on the TRPV1, ß3-adrenergic receptor (ß3-AR), and α1-AR using in vitro and in vivo models. The studies showed that CAP treatment in high-fat diet-induced obese mice resulted in lower body weight gain and elevated ATP-dependent thermogenic effectors' protein and gene expression through ATP-consuming calcium and creatine futile cycles. In both in vitro and in vivo experiments, CAP treatment elevated the protein and gene expressions of sarcoendoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA2), ryanodine receptor 2 (RYR2), creatine kinase B (CKB), and creatine kinase mitochondrial 2 (CKMT2) mediated by the activation of ß3-AR, α1-AR, and TRPV1. Our study showed that CAP increased intracellular Ca2+ levels and the expression of voltage-dependent anion channel (VDAC) and mitochondrial calcium uniporter (MCU) which indicates that increased mitochondrial Ca2+ levels lead to increased expression of oxidative phosphorylation protein complexes as a result of ATP-futile cycle activation. A mechanistic study in 3T3-L1 adipocytes revealed that CAP induces UCP1- and ATP-dependent thermogenesis mediated by the ß3-AR/PKA/p38MAPK/ERK as well as calcium-dependent α1-AR/TRPV1/CaMKII/AMPK/SIRT1 pathway. Taken together, we identified CAP's novel functional and modulatory roles in UCP1- and ATP-dependent thermogenesis, which is important for developing therapeutic strategies for combating obesity and metabolic diseases.

Echinacoside stimulates myogenesis and ATP-dependent thermogenesis in the skeletal muscle via the activation of D1-like dopaminergic receptors.

Recent studies have shown that some natural compounds from plants prevent obesity and related disorders, including the loss of skeletal muscle mass and strength. In this study, we investigated the effect of echinacoside (ECH), a caffeic acid glycoside from the phenylpropanoid class, on myogenesis and ATP-dependent thermogenesis in the skeletal muscle and its interaction with the dopaminergic receptors 1 and 5 (DRD1 and DRD5). We applied RT-PCR, immunoblot analysis, a staining method, and an assay kit to determine the effects of ECH on diverse target genes and proteins involved in skeletal muscle myogenesis and ATP-consuming futile processes. Our study demonstrated that ECH enhanced myogenic differentiation, glucose, and fatty acid uptake, as well as lipid catabolism, and induced ATP-dependent thermogenesis in vitro and in vivo. Moreover, ECH upregulated mitochondrial biogenesis proteins, mitochondrial oxidative phosphorylation (OXPHOS) complexes, and intracellular Ca2+ signaling as well as thermogenic proteins. These findings were further elucidated by mechanistic studies which showed that ECH mediates myogenesis via the DRD1/5 in C2C12 muscle cells. In addition, ECH stimulates α1-AR-mediated ATP-dependent thermogenesis via the DRD1/5/cAMP/SLN/SERCA1a pathway in C2C12 muscle cells. To the best of our knowledge, this is the first report that demonstrates the myogenic and thermogenic potential of ECH activity through the dopaminergic receptors. Understanding the novel functions of ECH in terms of its ability to prevent skeletal muscle loss and energy expenditure via ATP-consuming futile processes could help to develop potential alternative strategies to address muscle-related diseases, including combating obesity.

Echinacoside Induces UCP1- and ATP-Dependent Thermogenesis in Beige Adipocytes via the Activation of Dopaminergic Receptors.

Echinacoside (ECH) is a naturally occurring phenylethanoid glycoside, isolated from Echinacea angustifolia, and this study aimed to analyze its effect on thermogenesis and its interaction with dopaminergic receptors 1 and 5 (DRD1 and DRD5) in 3T3-L1 white adipocytes and mice models. We employed RT-PCR, immunoblot, immunofluorescence, a staining method, and an assay kit to determine its impact. ECH showed a substantial increase in browning signals in vitro and a decrease in adipogenic signals in vivo. Additionally, analysis of the iWAT showed that the key genes involved in beiging, mitochondrial biogenesis, and ATP-dependent thermogenesis were upregulated while adipogenesis and lipogenesis genes were downregulated. OXPHOS complexes, Ca2+ signaling proteins as well as intracellular Ca2+ levels were also upregulated in 3T3-L1 adipocytes following ECH treatment. This was collectively explained by mechanistic studies which showed that ECH mediated the beiging process via the DRD1/5-cAMP-PKA and subsequent downstream molecules, whereas it co-mediated the α1-AR-signaling thermogenesis via the DRD1/5/SERCA2b/RyR2/CKmt pathway in 3T3-L1 adipocytes. Animal experiments revealed that there was a 12.28% reduction in body weight gain after the ECH treatment for six weeks. The effects of ECH treatment on adipose tissue can offer more insights into the treatment of obesity and metabolic syndrome.

Silencing of dopamine receptor D5 inhibits the browning of 3T3-L1 adipocytes and ATP-consuming futile cycles in C2C12 muscle cells.

BACKGROUND: As a part of the catecholamines, dopamine receptors (DRs) have not been extensively studied like ß3-AR in the thermogenesis process. The present study investigates the effect of DRD5 in browning events and ATP-consuming futile cycles. METHODS: siRNA technology, qPCR, immunoblot analysis, immunofluorescence, and staining methods were used to investigate the effect of DRD5 on 3T3-L1 and C2C12 cells. RESULTS: siDdr5 increased lipogenesis-associated effectors, and adipogenesis markers while reducing the expression of beige fat effectors. ATP-consuming futile cycle markers were also reduced following the siDrd5. On the contrary, pharmacological activation of DRD5 stimulated these effectors. Our mechanistic studies elucidated that DRD5 mediates fat browning via the cAMP-PKA-p38 MAPK signalling pathway in 3T3-L1 cells as well as the cAMP-SERCA-RyR pathway for the ATP-consuming futile cycles in both cells. CONCLUSIONS: siDrd5 positively regulates browning and ATP-consuming futile cycles, and understanding its functions will provide insights into novel strategies to treat obesity.

Dopamine receptor D4 (DRD4) negatively regulates UCP1- and ATP-dependent thermogenesis in 3T3-L1 adipocytes and C2C12 muscle cells.

The activation of beige fat and muscle tissues is an interesting and encouraging target for therapeutic intervention in obesity owing to their remarkable lipolytic activity and energy-consuming futile cycles. This study examined the effect of dopamine receptor D4 (DRD4) on lipid metabolisms as well as UCP1- and ATP-dependent thermogenesis in Drd4-silenced 3T3-L1 adipocytes and C2C12 muscle cells. Silencing of Drd4, followed by quantitative real-time PCR, immunoblot analysis, immunofluorescence, and staining methods, were applied to evaluate the effects of DRD4 on diverse target genes and proteins of both cells. The findings showed that DRD4 was expressed in the adipose and muscle tissues of normal and obese mice. Furthermore, the knockdown of Drd4 upregulated the expression of brown adipocyte-specific genes and proteins while downregulating lipogenesis and the adipogenesis marker proteins. Drd4 silencing also upregulated the expression of key signaling molecules involved in ATP-dependent thermogenesis in both cells. This was further elucidated by mechanistic studies showing that a Drd4 knockdown mediates UCP1-dependent thermogenesis via the cAMP/PKA/p38MAPK pathway in 3T3-L1 adipocytes and UCP1-independent thermogenesis via the cAMP/SLN/SERCA2a pathway in C2C12 muscle cells. In addition, siDrd4 also mediates myogenesis via the cAMP/PKA/ERK1/2/Cyclin D3 pathway in C2C12 muscle cells. Silencing of Drd4 promotes ß3-AR-dependent browning in 3T3-L1 adipocytes and α1-AR/SERCA-based thermogenesis through an ATP-consuming futile process in C2C12 muscle cells. Understanding the novel functions of DRD4 on adipose and muscle tissues in terms of its ability to enhance energy expenditure and regulate whole-body energy metabolism will aid in developing novel obesity intervention techniques.

ß-Carotene induces UCP1-independent thermogenesis via ATP-consuming futile cycles in 3T3-L1 white adipocytes.

The activation of brown fat and induction of beige adipocytes, so-called non-shivering thermogenesis, is emerging as a promising target for therapeutic intervention in obesity management. Our previous report demonstrated that ß-carotene (BC) induces beige adipocytes to increase UCP1-dependent thermogenic activity. However, the UCP1-independent thermogenic effect of BC on adipose tissues remains unexplored. In this study, we examined the effects of BC on UCP1-independent thermogenic activity with a focus on the ATP-consuming futile cycles in 3T3-L1 adipocytes. BC increased intracellular calcium levels and stimulated the expression of calcium cycling-related proteins, including sarcoendoplasmic reticulum Ca2+-ATPase (SERCA) 2b, ryanodine receptor 2 (RyR2), voltage-dependent anion channel (VDAC), mitochondrial calcium uniporter (MCU), and Ca2+/calmodulin-dependent protein kinase 2 (CaMK2) in 3T3-L1 white adipocytes. In addition, BC stimulated thermogenesis by activating the creatine metabolism-related thermogenic pathway. Moreover, BC activated ß-carotene oxygenase 1 (BCO1), which efficiently cleaved BC to retinal and consequently converted to its transcriptionally active form retinoic acid. These BC conversion products also exhibited thermogenic effects comparable to a similar level of BC. The mechanistic study revealed that retinal exhibited thermogenic activity independently of retinoic acid and retinoic acid-mediated thermogenesis was resulted partly from conversion of retinal. Moreover, BC activated α1-AR and UCP1-independent thermogenic effectors independently of UCP1 expression. In conclusion, the thermogenic response to BC and its conversion products in 3T3-L1 white adipocytes involves two interacting pathways, one mediated via ß3-adrenergic receptors (ß3-AR) and cyclic adenosine monophosphate (cAMP) and the other via α1-AR and increases in cytosolic Ca2+ levels activated by calcium regulatory proteins.

Dopaminergic and adrenergic receptors synergistically stimulate browning in 3T3-L1 white adipocytes.

The browning of white adipose tissue (WAT) has attracted considerable attention in the scientific community as a popular strategy for enhancing energy expenditure to combat obesity. As a part of this strategy, ß3-adrenergic receptor (ß3-AR) is the most widely studied receptor that mediates thermogenesis. Parenthetically, further studies in search for additional receptors expressed in adipocytes that can mediate thermogenesis has been appearing, and this paper reports that dopaminergic receptor 1 (DRD1) and ß3-AR synergistically stimulate browning in 3T3-L1 white adipocytes. qRT-PCR and immunoblot analysis methods were applied to evaluate the effects of DRD1 on the target proteins downstream of ß3-AR and other markers involved in lipid metabolism, mitochondrial biogenesis, and browning events. These results show that DRD1 is expressed in epididymal WAT (eWAT), brown adipose tissue (BAT), and inguinal WAT (iWAT) of normal and high-fat-fed mice, and a deficiency of DRD1 downregulates the expression of brown adipocyte-specific proteins. Silencing of DRD1 affected lipid metabolic activity in 3T3-L1 adipocytes by reducing mitochondrial biogenesis as well as levels of lipolytic and fat oxidative marker proteins in a similar pattern to ß3-AR. Moreover, mechanistic studies showed that the depletion of DRD1 downregulates ß3-AR and its downstream molecules, suggesting both receptors might synergistically stimulate browning. Parallel to the UCP1-dependent thermogenesis, the depletion of DRD1 also downregulates the expression of core proteins responsible for UCP1-independent thermogenesis. Overall, DRD1 mediates ß3-AR-dependent 3T3-L1 browning and UCP1-independent thermogenesis.

Colchicine stimulates browning via antagonism of GABA receptor B and agonism of ß3-adrenergic receptor in 3T3-L1 white adipocytes.

Colchicine has been used for therapeutic purposes and has attracted considerable attention because of its association with tubulin and the inhibition of small tubular polymerization. Although several studies have examined the possible preventive role of colchicine in metabolic diseases, its role in adipocytes is largely unknown. This study examined the novel functional role of colchicine in adipocytes demonstrating that colchicine stimulates browning in cultured white adipocytes. Colchicine stimulates browning by increasing the brown- and beige fat-specific markers in 3T3-L1 white adipocytes. Interestingly, colchicine decreased the expression of the main lipolytic proteins (ATGL, p-HSL) while it activated Ces3, suggesting a possibility for supplying essential fatty acids for inducing thermogenesis. Molecular docking analysis showed that colchicine has a strong affinity against GABA-BR and ß3-AR, and its binding activity with GABA-BR (-26.52 kJ/mol) was stronger than ß3-AR (-20.71 kJ/mol). Mechanistic studies were conducted by treating the cells separately with agonists and antagonists of GABA-BR and ß3-AR to understand the molecular mechanism underlying the browning effect of colchicine. The results showed that colchicine stimulates browning via the antagonism of GABA-BR and the agonism of ß3-AR in 3T3-L1 white adipocytes. The colchicine-mediated activation of ß3-AR stimulated the PKA/p38 MAPK signaling pathway, where consequently ATF2 acted as a positive regulator, but AFT4 was a negative regulator for the induction of browning.

Loss of cytoplasmic FMR1-interacting protein 2 (CYFIP2) induces browning in 3T3-L1 adipocytes via repression of GABA-BR and activation of mTORC1.

Obesity and related metabolic disorders are epidemic diseases. Promoting thermogenesis and a functional increase in the browning of white adipocytes may counteract obesity. On the other hand, the molecular mechanism that regulates brown and beige fat-mediated thermogenesis is unclear. This article reports a molecular network led by cytoplasmic FMR1-interacting protein 2 (CYFIP2) that negatively regulates adipocyte browning in white adipocytes. Although the function of CYFIP2 in Fragile X Syndrome (FXS) and autism have been reported, its physiological roles in adipocytes remain elusive. Therefore, this study examined the physiological consequences of its deprivation in cultured 3T3-L1 white adipocytes using loss-of-function studies. Combined real-time quantitative reverse-transcription polymerase chain reaction and immunoblot analysis showed that the loss of CYFIP2 induces fat browning, as evidenced by the gene and protein expression levels of the brown fat-associated markers. A deficiency of CYFIP2 promoted mitochondrial biogenesis and significantly enhanced the expression of the core set beige fat-specific genes (Cd137, Cidea, Cited1, Tbx1, and Tmem26) and proteins (PGC-1α, PRDM16, and UCP1). In addition, a CYFIP2 deficiency promoted lipid catabolism and suppressed adipogenesis, lipogenesis, and autophagy. A mechanistic study showed that the loss of CYFIP2 induces browning in white adipocytes, independently via the activation of mTORC1 and suppression of the GABA-BR signaling pathway. The present data revealed a previously unidentified mechanism of CYFIP2 in the browning of white adipocytes and emphasized the potential of CYFIP2 as a pharmacotherapeutic target for treating obesity and other metabolic disorders.

Prednisone stimulates white adipocyte browning via ß3-AR/p38 MAPK/ERK signaling pathway.

AIMS: Prednisone is a corticosteroid-derived drug which is widely used for its role in immunosuppression and treatment of lung disorders. The current study reports, for the first time, the critical role of prednisone in the induction of white fat browning, thereby promoting thermogenic effect in cultured white adipocytes. MAIN METHODS: The fat-browning activity of prednisone was evaluated in 3T3-L1 cells by quantitative real-time PCR, immunoblot analysis, immunofluorescence, and molecular docking techniques. KEY FINDINGS: Exposure to prednisone stimulated browning in 3T3-L1 white adipocytes by increasing the expressions of core fat browning marker proteins (UCP1, PGC-1α and PRDM16) as well as beige-specific genes (Cd137, Cidea, Cited1, and Tbx1) via ATF2 and CREB activation mediated by p38 MAPK and ERK signaling, respectively. Prednisone exposure also resulted in the robust activation of lipolytic and fatty acid oxidation marker proteins, thereby increasing mitochondrial biogenesis. In addition, prednisone treatment resulted in reduced expression levels of adipogenic transcription factors while elevating SIRT1, as well as attenuation of lipogenesis and lipid droplets formation. Furthermore, molecular docking and mechanistic studies demonstrated the recruitment of beige fat by prednisone via the ß3-AR/p38 MAPK/ERK signaling pathway. SIGNIFICANCE: Taken together, these results indicate the unique role of prednisone as a fat-browning stimulant, and demonstrate its therapeutic potential in the treatment of obesity by enhancing thermogenesis.

ß-Carotene stimulates browning of 3T3-L1 white adipocytes by enhancing thermogenesis via the ß3-AR/p38 MAPK/SIRT signaling pathway.

BACKGROUND: Natural compounds with medicinal properties are part of a strategic trend in the treatment of obesity. The vitamin A agent, ß-carotene, is a well-known carotenoid, and its numerous functions in metabolism have been widely studied. The activation of thermogenesis by stimulating white fat browning (beiging) has been identified as a treatment for obese individuals. PURPOSE: The current study was undertaken to unveil the browning activity of ß-carotene in 3T3-L1 white adipocytes. METHODS: The effects of ß-carotene were evaluated in 3T3-L1 white adipocytes, and gene/protein expressions were determined by performing quantitative real-time PCR, immunoblot analysis, immunofluorescence assessment, and molecular docking techniques. RESULTS: ß-carotene strikingly increased the expression levels of brown-fat-specific marker proteins (UCP1, PRDM16, and PGC-1α) and beige-fat-specific genes (Cd137, Cidea, Cited1, andTbx1) in 3T3-L1 cells. Exposure to ß-carotene also elevated the expressions of key adipogenic transcription factors C/EBPα and PPARγ in white adipocytes but decreased the expressions of lipogenic marker proteins ACC and FAS. Moreover, lipolysis and fat oxidation were regulated by ß-carotene via upregulation of ATGL, pHSL, ACOX, and CPT1. In addition, molecular docking studies revealed ß-carotene activation of the adenosine A2A receptor and ß3-AR. ß-Carotene increased the expressions of mitochondrial biogenic markers, stimulated the ß3-AR and p38 MAPK signaling pathways and its downstream signaling molecules (SIRTs and ATF2), thereby inducing browning. CONCLUSIONS: Taken together, our results indicate the potential of ß-carotene as a natural-source therapeutic anti-obesity agent.

Anthocyanin oligomers stimulate browning in 3T3-L1 white adipocytes via activation of the ß3-adrenergic receptor and ERK signaling pathway.

Microbial fermentation of grape-skin extracts is found to synthesize anthocyanin oligomers (AO), which are more active than the monomeric anthocyanins that are effective for some metabolic diseases such as diabetes and obesity. This study investigated the functional role of AO in 3T3-L1 white adipocyte metabolism, with a focus on inducing browning. To achieve this, we determined the expressions of core genes and protein markers responsible for browning and lipid metabolism in response to AO treatment of 3T3-L1 white adipocytes. AO exposure significantly increases the expressions of beige-specific genes (Cidea, Cited1, Ppargc1α, Prdm16, Tbx1, Tmem26, and Ucp1) and brown-fat signature proteins (UCP1, PRDM16, and PGC-1α), and suppresses the expressions of lipogenic marker proteins while enhancing the protein levels of lipolysis in white adipocytes. The mechanistic study revealed stimulation of white fat browning via activation of the ß3-AR/PKA/p38 axis and ERK/CREB signaling pathway subsequent to AO treatment. In conclusion, our current findings indicate the beneficial effects of AO for the treatment of obesity with interesting properties such as regulating the browning of adipocytes and increasing thermogenic activity. Although further research based on animal models or clinical trials remains, AO treatment can bring more insights into the treatment of obesity and metabolic syndrome.

Loss of lymphocyte cytosolic protein 1 (LCP1) induces browning in 3T3-L1 adipocytes via ß3-AR and the ERK-independent signaling pathway.

Increased browning of white adipocytes (beiging) is considered a promising therapeutic strategy to fight obesity and its associated metabolic complications. However, the molecular mechanism modulating brown and beige fat-mediated thermogenesis is not fully elucidated. Here, we identified the lymphocyte cytosolic protein 1 (LCP1) as a factor that obstructs fat browning in white adipocytes. LCP1 plays a vital role in non-hematopoietic malignancies, and is also a well-known tumor biomarker; however, evidence regarding its function in adipocytes remains to be elucidated. The current study explores the physiological role of LCP1 in cultured 3T3-L1 white adipocytes, by applying the loss-of-function study using siRNA. Induction of fat browning by LCP1 depletion was evidenced by evaluating the gene and protein expression levels of brown fat-associated markers through real-time qRT-PCR and immunoblot analysis, respectively. We observed that deficiency of LCP1 promotes mitochondrial biogenesis, and significantly enhances expressions of the core brown fat-specific genes (Cd137, Cidea, Cited1, Tbx1, and Tmem26) and proteins (PGC-1α, PRDM16, and UCP1). In addition, deficiency of LCP1 promotes lipid catabolism as well as suppresses adipogenesis and lipogenesis. Loss of LCP1 also ameliorates cellular stress by downregulating JNK and c-JUN in adipocytes, and stimulates apoptosis. A mechanistic study revealed that deficiency of LCP1 induces browning in white adipocytes, independently via ß3-AR and the ERK signaling pathway. The current data reveals a previously unknown mechanism of LCP1 in browning of white adipocytes, and highlights the potential of LCP1 as a pharmacotherapeutic target for treating obesity and other metabolic disorders.

Cytochrome P450 2F2 (CYP2F2) negatively regulates browning in 3T3-L1 white adipocytes.

Cytochromes P450 (CYPs) are a multigene superfamily of constitutively expressed and inducible enzymes responsible for the detoxification of many endogenous and exogenous compounds and for the metabolism of numerous medications. The cytochrome P450 2F2 (CYP2F2) subfamily is preferentially expressed in the respiratory tract, but its functional role in adipocytes has never been explored. We found that CYP2F2 was highly expressed during the differentiation of the C3H10T1/2 murine mesenchymal stem cells to adipocytes and here we have explored its functional role in adipocytes. The expression of thermogenic marker proteins such as peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), PR domain containing 16 (PRDM16), and uncoupling protein 1 (UCP1) and beige-fat specific genes were significantly increased in Cyp2f2-deficient 3T3-L1 adipocytes. Moreover, Cyp2f2 silencing led to reduced adipogenesis and lipogenesis, and enhanced lipid catabolism through the increased expression of lipolytic and fatty acid oxidative enzymes. A mechanistic study to identify molecular signals for CYP2F2-mediated negative regulation in the browning of white adipocytes revealed that CYP2F2 impairs the beta-3 adrenergic receptor (ß3-AR) activation as well as its downstream regulators including protein kinase A (PKA), p38 mitogen-activated protein kinase (p38 MAPK), and activating transcription factor 2 (ATF2). This data provides evidence that CYP2F2 is a negative regulator of lipid catabolism and browning in white adipocytes, suggesting that inhibitors of CYP2F2 could be potential drugs for the treatment of obesity with a focus on enhancing energy expenditure.

Cytochrome P450 2E1 (CYP2E1) positively regulates lipid catabolism and induces browning in 3T3-L1 white adipocytes.

AIMS: Browning induction (beiging) of white adipocytes is an emerging prospective strategy to defeat obesity and its related metabolic disorders. Cytochrome P450 2E1 (CYP2E1), a membrane protein which belongs to the cytochrome P450 superfamily, reportedly functions in the xenobiotic metabolism in the body, especially ethanol metabolism. Although previous studies have reported the effect of CYP2E1 on obesity in animal models, the data remains controversial. In the current study, we investigate for the first time, the role of CYP2E1 in lipid metabolism in 3T3-L1 white adipocytes, with a focus on fat browning. METHODS: 3T3-L1 white adipocytes and Cyp2e1 siRNA were applied to investigate the role of CYP2E1 in white adipocytes. After that, cells were seperately exposed to ß3-AR agonist, ß3-AR antagonist and p38 inhibitor to identify the pathway which CYP2E1 was involved in to regulate browning event in white adipocytes. KEY FINDINGS: We found that CYP2E1 deficiency results in reduced adipogenesis and lipogenesis as well as brown adipocyte-like phenotype induction. A mechanistic study to identify the molecular signals for CYP2E1 regulation in the browning of white adipocytes revealed that CYP2E1 inhibition deters the ß3-adrenergic receptor activation and its downstream targets. SIGNIFICANCE: Our data unveilved a previously unknown mechanism in the regulation of browning by CYP2E1 in 3T3-L1 white adipocytes, suggesting that CYP2E1 is a promising molecular target for the treatment of obesity and its related diseases.

Loss of family with sequence similarity 107, member A (FAM107A) induces browning in 3T3-L1 adipocytes.

Induction of white fat browning (beiging) and activation of brown fat has been considered a promising strategy to treat obesity and associated metabolic complications. However, the molecular mechanisms regulating brown and beige fat-mediated thermogenesis remains unclear. Our study aimed to identify genes with a hitherto unknown mechanism in the metabolic functions of adipocytes and identified family with sequence similarity 107, member A (FAM107A) as a factor that interferes with fat browning in white adipocytes. We explored physiological roles of FAM107A in cultured 3T3-L1 white adipocytes and HIB1B brown adipocytes by using FAM107A-deficient adipocytes. Significant loss in FAM107A gene functionality induced fat browning was evidenced by evaluating the gene and protein expression level of brown fat-associated markers through real-time qRT-PCR and immunoblot analysis, respectively. Deficiency of FAM107A promoted mitochondrial biogenesis and significantly upregulated core fat-browning marker proteins (PGC-1α, PRDM16, and UCP1) and beige-specific genes (Cd137, Cited1, Tbx1, and Tmem26). Furthermore, FAM107A increased adipogenesis and negatively regulated lipid metabolism in 3T3-L1 adipocytes. In addition, in-silico analysis revealed a strong interaction between FAM107A and ß3-AR based on their energy binding score. Next, mechanistic study revealed that specific knockdown of FAM107A induces browning in white adipocytes via activation of ß3-AR, AMPK and p38 MAPK-dependent signaling pathways. Our data unveiled a previously unknown mechanism of FAM107A in the regulation of lipid metabolism and identified its significant role in metabolic homeostasis. This highlighted the potential of FAM107A as a pharmacotherapeutic target in treating obesity and related metabolic disorders.

Novel regulatory roles of UCP1 in osteoblasts.

AIMS: The bone-adipose axis requires complex homeostasis in energy and global metabolism. The bioenergetics of bone establishes the necessary energy balance to coordinate endocrine functions that are affected by various factors and is not limited to matrix proteins only. UCP1 is an uncoupling protein of adipocytes, commonly known for its unique feature of promoting thermogenesis, mainly in brown fat; however, the effects of UCP1 in other cell types remain unreported. MAIN METHODS: In the current study, we determined the roles of UCP1 in osteoblasts by silencing the Ucp1 gene in MC-3T3-E1 cells, as well as C3H10T1/2 mesenchymal stem cells, and explored its functional activities. KEY FINDINGS: Our results demonstrate for the first time the presence of UCP1 in osteoblast cells. We identified that UCP1 regulates ATP and oxidative phosphorylation in MC-3T3-E1 cells. In addition, our data reveal that the lack of Ucp1 results in reduced expressions of regulatory proteins involved in scavenging of ROS by enhancing an autophagic event to balance osteogenic differentiation. SIGNIFICANCE: In conclusion, this study highlights a novel perspective on the importance of UCP1 in bone cells.

BMP11 regulates thermogenesis in white and brown adipocytes.

Bone morphogenetic protein-11 (BMP11), also known as growth differentiation factor-11 (GDF11), is implicated in skeletal development and joint morphogenesis in mammals. However, its functions in adipogenesis and energy homeostasis are mostly unknown. The present study investigates crucial roles of BMP11 in cultured 3T3-L1 white and HIB1B brown adipocytes, using Bmp11 gene depletion and pharmacological inhibition of BMP11. The silencing of Bmp11 markedly decreases the expression levels of brown-fat signature proteins and beige-specific genes in white adipocytes and significantly down-regulates the expression levels of brown fat-specific genes in brown adipocytes. The deficiency of Bmp11 reduces the expressions of lipolytic protein markers in white and brown adipocytes. Moreover, BMP11 induces browning of 3T3-L1 adipocytes via coordination of multiple signalling pathways, including mTORC1-COX2 and p38MAPK-PGC-1α as non-canonical pathways, as well as Smad1/5/8 as a canonical pathway. We believe this study is the first to provide evidence of the potential roles of BMP11 for improvement of lipid catabolism in both cultured white and brown adipocytes, as well as the effect on browning of white adipocytes. Taken together, these results demonstrate the therapeutic potential for the treatment of obesity.

Re-engineering of peptides with high binding affinity to develop an advanced electrochemical sensor for colon cancer diagnosis.

Colorectal cancer (CRC) develops from polyps in the inner large intestine or rectum and an increasing incidence and high mortality rate has been observed in humans. Currently, colonoscopy is the preferred modality for early CRC diagnosis. However, this technique has several limitations, such as high medical costs and intricate procedures, leading to increasing demands for the development of a new, simple, and affordable diagnostic method. In this study, an advanced electrochemical biosensor based on rationally designed affinity peptides was developed for discriminating adenoma to carcinoma progression. Amino acid-substituted and rationally designed synthetic peptides (BP3-1 to BP3-8) based on in silico modeling studies were chemically synthesized, and covalently immobilized onto a gold electrode using aromatic ring compounds through surface chemistry techniques. The binding performance of the developed sensor system was observed using square wave voltammetry (SWV). The peptide BP3-2 was selected depending on its relative binding affinity; SWV indicated the limit of detection of BP3-2 for LRG1 to be 0.025 µg/mL. This sensor could distinguish the adenoma-carcinoma transition with improved binding abilities (specificity and selectivity), and stability in plasma samples spiked with LRG1 and real samples from patients with CRC. These results indicate that this electrochemical sensor system can be used for early monitoring of the colorectal adenoma to carcinoma progression.