Differentiation Of Preadipocytes Research Articles

The IL-33/ST2 Axis Affects Adipogenesis Through Regulating the TRAF6/RelA Pathway

Understanding the regulatory mechanisms of adipogenesis is essential for preventing obesity. Interleukin-33 (IL-33) has recently attracted increasing attention for its role in adipogenesis. The purpose of this study was to explore the function and regulatory mechanism of IL-33 and its receptor suppression of tumorigenicity 2 (ST2) on adipogenesis. Here, Oil Red O staining was used to detect the accumulation of intracellular lipid droplets. Molecular techniques such as qRT-PCR and Western blotting were used to detect the expression of pivotal genes and adipogenic marker genes. Gains and losses of function experiments were used to explore the potential regulatory mechanism of the IL-33/ST2 axis in adipogenesis. Functionally, IL-33 is negatively associated with adipogenesis in 3T3-L1 preadipocytes, while ST2 is positively associated with it, encompassing both the trans-membrane receptor ST2 (ST2L) and the soluble ST2 (sST2). Mechanistically, the IL-33/ST2 axis affects adipogenesis by regulating the expression of the TRAF6/RelA pathway in 3T3-L1 preadipocytes. Downregulating the expression of ST2 suppressed the activation of the IL-33/ST2 axis, which subsequently inhibits the expression of TRAF6. This further attenuates the expression of RelA, ultimately resulting in the suppression of adipogenesis in 3T3-L1 preadipocytes. This study reveals a new mechanism by which the IL-33/ST2 axis regulates the differentiation of preadipocytes and provides a new idea for improving obesity prevention.

Integrative 3D genomics with multi-omics analysis and functional validation of genetic regulatory mechanisms of abdominal fat deposition in chickens

Chickens are the most abundant agricultural animals globally, with controlling abdominal fat deposition being a key objective in poultry breeding. While GWAS can identify genetic variants associated with abdominal fat deposition, the precise roles and mechanisms of these variants remain largely unclear. Here, we use male chickens from two lines divergently selected for abdominal fat deposition as experimental models. Through the integration of genomic, epigenomic, 3D genomic, and transcriptomic data, we build a comprehensive chromatin 3D regulatory network map to identify the genetic regulatory mechanisms that influence abdominal fat deposition in chickens. Notably, we find that the rs734209466 variant functions as an allele-specific enhancer, remotely enhancing the transcription of IGFBP2 and IGFBP5 by the binding transcription factor IRF4. This interaction influences the differentiation and proliferation of preadipocytes, which ultimately affects phenotype. This work presents a detailed genetic regulatory map for chicken abdominal fat deposition, offering molecular targets for selective breeding.

LncBNIP3 Inhibits Bovine Intramuscular Preadipocyte Differentiation via the PI3K-Akt and PPAR Signaling Pathways.

Intramuscular fat (IMF) content is an economic trait in beef cattle that improves the meat quality. Studies have highlighted the correlation between long noncoding RNAs (lncRNAs) and IMF development. In this study, lncBNIP3 knockdown promoted bovine intramuscular preadipocyte differentiation. RNA-seq analysis of intramuscular preadipocytes with lncBNIP3 knockdown identified 230 differentially expressed genes. The PI3K-Akt and PPAR signaling pathways were enriched. lncBNIP3 interference promoted mRNA and protein expression of key genes in PI3K-Akt signaling pathway. LncBNIP3 interference reversed the effects of an AKT-inhibitor MK-2206 on Akt protein expression and lipid droplet accumulation, promoted mRNA and protein expression of essential genes in the PPAR signaling pathway, and ameliorated the inhibitory effects of a PPARg antagonist GW9662 on lipid accumulation. Therefore, lncBNIP3 inhibition of bovine intramuscular preadipocyte differentiation is likely mediated via the PI3K-Akt and PPAR signaling pathways. This study identified a valuable lncRNA with functional roles in IMF accumulation and revealed new strategies to improve beef quality.

Chi-circ_0009659 modulates goat intramuscular adipocyte differentiation through miR-3431-5p/STEAP4 axis.

Circular RNAs (circRNAs) are widely involved in the regulation of lipid deposition in animals, but there are few reports on key circRNAs regulating intramuscular adipocyte differentiation in goats. Therefore, this study took an abundantly expressed in goat adipocytes chi-circ_0009659 as the object. Based on the identification of back splicing site in chi-circ_0009659, its expression level during the goat intramuscular preadipocyte differentiation was detected by qPCR. The chi-circ_0009659 loss-of-function and gain-of-function cell models were obtained by adenovirus and smarter silencer, respectively. and the adipocyte differentiation were explored by Oil Red O staining, Bodipy staining and qPCR. Its major cytoplasmic localization was determined by FISH, nucleocytoplasmic separation and qPCR. The interaction between chi-circ_0009659, miR-3431-5p, and STEAP family member 4 (STEAP4) was verified by bioinformatics, RNA pull down and dual luciferase reporter assay. Silencing chi-circ_0009659 inhibited lipid droplet accumulation and the expression of differentiation-determining genes in goat intramuscular adipocytes, while overexpression of chi-circ_0009659 reversed these results. chi-circ_0009659 was predominantly localized to the cytoplasm and could regulate miR-3431 expression which in turn affects STEAP4. Consistent with expectations, miR-3431-5p acted as a negative regulator of GIMPA differentiation, while STEAP4 promoted differentiation. We demonstrated chi-circ_0009659 positively regulates goat intramuscular preadipocyte differentiation by sponging miR-3431-5p to further regulate the expression of STEAP4. This research provides a new reference for in-depth understanding of the effects of circRNA on adipocyte differentiation.

Adipogenic Effects of Cresyl Diphenyl Phosphate (Triphenyl Phosphate Alternative) through Peroxisome Proliferator-Activated Receptor Gamma Pathway: A Comprehensive Study Integrating In Vitro, In Vivo, and In Silico from Molecule to Health Risk.

Cresyl diphenyl phosphate (CDP), a novel organophosphate ester (OPE), has been detected in various environmental and human samples. However, there is very limited knowledge regarding its toxicity, mechanisms of action, and potential health risks. Using new alternative methods (NAMs), across the molecular interactions, signaling pathways, cell functions, animal effects, and population risks, we investigated the potential adipogenic effects and associated risks of CDP and legacy OPE triphenyl phosphate (TPHP) by acting on peroxisome proliferator-activated receptor gamma (PPARγ). Among the 19 screened OPEs, CDP bound to PPARγ with the highest binding potency, followed by TPHP. CDP activated PPARγ through fitting into the binding pocket with strong hydrophobicity and hydrogen bond interactions; CDP exhibited higher potency compared to TPHP. In 3T3-L1 cells, CDP enhanced the PPARγ-mediated adipogenesis activity, exhibiting greater potency than TPHP. The intracellular concentration and receptor-bound concentrations (RBC) of CDP were also higher than those of TPHP in both HEK293 cells and 3T3-L1 cells. In mice, exposure to CDP activated the PPARγ-mediated adipogenic pathway, leading to an increased white adipose tissue weight gain. Overall, CDP could bind to and activate PPARγ, thereby promoting preadipocyte differentiation and the development of white adipose tissue. Its potential obesogenic risks should be of high concern.

8700 Mono(ADP-Ribosyl)ation of Ribosomal Proteins Promotes Differentiation During Adipogenesis

Abstract Disclosure: X. Tan: None. S. Challa: None. C.V. Camacho: None. T.S. Nandu: None. M.S. Stokes: None. W.L. Kraus: Other; Self; W.L.K. is a founder and stockholder for Ribon Therapeutics, Inc; a founder, member of the SAB, member of the BOD, and a stockholder for and ARase Therapeutics, Inc. He is also coholder of U.S. Patent. ADP-ribosylation, a crucial post-translational modification, involves the covalent attachment of ADP-ribose units to amino acids on proteins through ADP-ribosyl transferase (ART) enzymes, utilizing NAD+ synthesized by nicotinamide mononucleotide adenylyl transferases (NMNATs). We previously showed that NMNAT-2, a cytoplasmic NAD+ synthase, increases during adipogenesis, leading to reduced nuclear NAD+ levels through compartmentalized NMN depletion and enhanced differentiation of preadipocytes. However, the implications of increased cytoplasmic NAD+ synthesis by NMNAT-2 in adipocytes remain unknown. Our current findings demonstrate elevated ribosome MARylation during 3T3-L1 preadipocyte differentiation, driven by NMNAT-2 induction. Inhibiting NAD+ biosynthesis by knockdown of NMNAT-2 or FK866 results in reduced ribosome MARylation, with PARP16 identified as the primary ART mediating MARylation. Knockdown of NMNAT-2 or PARP16 impaired global protein synthesis and preadipocyte differentiation. Treatment with the PARP16 inhibitor DB008 hindered ribosomal MARylation and preadipocyte differentiation. Immunofluorescence and subcellular fractionation analysis demonstrated that MARylated ribosomes are mostly associated with the endoplasmic reticulum. Using TMT mass spectrometry, we identified numerous high-confidence ADP-ribosylation sites on glutamate and aspartate residues in both undifferentiated and differentiated cells. We examined five proteins with specific sites of MARylation that are consistently more modified in the differentiated state. Among these, ADP-ribosylation of RPSA significantly promote preadipocyte differentiation; mutation of the site inhibits this effect. Ongoing investigations involve studying the impact of Parp16 knockout on high fat diet-induced obesity in mice to further determine the biological role of ribosome protein MARylation during adipogenesis. This work is supported by a grant from the NIH/NIDDK (R01 DK069710) and the DOD OCRP (OC200311). Presentation: 6/1/2024

Chemical Investigation and Regulation of Adipogenic Differentiation of Cultivated Moringa oleifera.

Background/Objectives: Moringa oleifera is a matrix plant with the high potential to cure several diseases with its medicinal and ethnopharmacological value and nutraceutical properties. In this study, we investigated the chemical and biological properties of this plant cultivated in our local region. Methods: Leaves, roots, seeds, stem bark, and twigs of oleifera were extracted and evaluated bioactivities targeting intracellular lipid accumulation and adipocyte differentiation in 3T3-L1 preadipocytes, and UHPLC-ESI-Orbitrap-MS/MS-Based molecular networking guided isolation and dereplication of metabolites from these extracts. Results: Five extracts of different organs of M. oleifera significantly stimulated intracellular lipid accumulation and adipocyte differentiation in 3T3-L1 preadipocytes in a concentration-dependent manner. These extracts markedly increased the expression of genes related to adipogenesis and lipogenesis. Notably, these extracts promoted peroxisome proliferator-activated receptor γ (PPARγ) activity and the expression of its target genes, including phosphoenolpyruvate carboxykinase, fatty acid-binding protein 4, and perilipin-2. These adipogenic and lipogenic effects of Moringa extracts through the regulation of PPARγ activity suggests their potential efficacy in preventing or treating type 2 diabetes. Furthermore, chemical investigation revealed high contents of phytonutrients as rich sources of secondary metabolites including glycosides, flavones, fatty acids, phenolics, and other compounds. In addition, in silico studies on major components of these extracts revealed the bioavailability of major components through their binding affinity to respective proteins targeting adipocyte differentiation.

Whole transcriptome sequencing analysis reveals the effect of circZFYVE9/miR-378a-3p/IMMT axis on mitochondrial function in adipocytes

Recent research highlights the complex regulation of lipid accumulation and mitochondrial function in adipocytes via non-coding RNAs like microRNAs and circular non-coding RNAs. Circular non-coding RNAs act as endogenous regulators, impacting lipid metabolism and mitochondrial function by interacting with miRNAs. Sequencing white and brown adipose tissues in mice revealed significant variations in 1936 mRNAs, 127 miRNAs, and 171 circRNAs. Analyses showed these RNAs' involvement in vital processes like mitochondrial biogenesis, oxidative phosphorylation, and the citric acid cycle, crucial for lipid metabolism. Focus on top differentially regulated miRNAs led to the construction of a regulatory network involving circRNAs, miRNAs, and mRNAs, illuminating the role of endogenous RNAs in lipid metabolism and mitochondrial function. The circZFYVE9/miR-378a-3p/IMMT axis was identified as influential in adipogenic differentiation of 3T3-L1 preadipocytes by regulating mitochondrial function. This study expands the understanding of non-coding RNAs in adipose tissue, particularly their connection to mitochondrial function and metabolism.

TRPV4 modulation affects mitochondrial parameters in adipocytes and its inhibition upregulates lipid accumulation

Enhanced lipid-droplet formation by adipocytes is a complex process and relevant for obesity. Using knock-out animals, involvement of TRPV4, a thermosensitive ion channel in the obesity has been proposed. However, exact role/s of TRPV4 in adipogenesis and obesity remain unclear and contradictory. Here we used in vitro culture of 3T3L-1 preadipocytes and primary murine-mesenchymal stem cells as model systems, and a series of live-cell-imaging to analyse the direct involvement of TRPV4 exclusively at the adipocytes that are free from other complex signalling as expected in in-vivo condition. Functional TRPV4 is endogenously expressed in pre- and in mature-adipocytes. Pharmacological inhibition of TRPV4 enhances differentiation of preadipocytes to mature adipocytes, increases expression of adipogenic and lipogenic genes, enhances cholesterol, promotes bigger lipid-droplet formation and reduces the lipid droplet temperature. On the other hand, TRPV4 activation enhanced the browning of adipocytes with increased UCP-1 levels. TRPV4 regulates mitochondrial-temperature, Ca2+-load, ATP, superoxides, cardiolipin, membrane potential (ΔΨm), and lipid-mitochondrial contact sites. TRPV4 also regulates the extent of actin fibres, affecting the cells mechanosensing ability. These findings link TRPV4-mediated mitochondrial changes in the context of lipid-droplet formation involved in adipogenesis and confirm the direct involvement of TRPV4 in adipogenesis. These findings may have broad implication in treating adipogenesis and obesity in future.

Ascochlorin Attenuates the Early Stage of Adipogenesis via the Wnt/β-Catenin Pathway and Inhibits High-Fat-Diet-Induced Obesity in Mice.

This study investigated the effects of ascochlorin (ASC), a natural compound derived from the fungus Ascochyta viciae, on adipogenesis and obesity. We determined the effects of ASC on 3T3-L1 preadipocytes and whether it ameliorated to mitigate high-fat diet (HFD)-induced obesity in C57BL/6J mice. We found that ASC significantly inhibited the differentiation of preadipocytes by modulating the Wnt/β-catenin signaling pathway, a key regulator of adipogenic processes. Treatment with ASC not only reduced the mRNA and protein expression of key adipogenic transcription factors such as C/EBPα and PPARγ, but also reduced lipid accumulation both in vitro and in vivo. In addition, treatment HFD-fed mice with ASC significantly reduced their weight gain and adiposity vs. control mice. These results suggest that ASC has considerable potential as a therapeutic agent for obesity, owing to its dual action of inhibiting adipocyte differentiation and reducing lipid accumulation. Thus, ASC represents a promising candidate as a natural anti-obesity agent.

Identification and functional analysis of a new cold induced LncRNA44154

With the development of high-throughput sequencing technology, numerous unknown long noncoding RNAs (lncRNAs) have been discovered. Preliminary research found that after being subjected to low-temperature enviroment, lncRNA44154 expression in the subcutaneous fat of pigs significantly increased. In order to analyze its biological function and regulatory mechanism, qRT-PCR was used to detect its spatiotemporal expression pattern. The nucleoplasmic isolation and FISH were used to locate it subcellular. Through overexpression or interference assays discussed the effect of lncRNA44154 on adipocyte proliferation and differentiation. RNA-seq was used to screen the biological pathways that lncRNA44154 may be involved in. RNA-pull down was used to find the interaction protein and double luciferase reporter gene detection experiment was used to validate the target gene. It was found that the expression of lncRNA44154 had tissue specificity and played a role in adipocyte proliferation. It was expressed in both cytoplasm and nucleus. After overexpression of lncRNA44154 in adipocytes, the expression level of Cyclins B, D, and E were all up-regulated and the cell proliferation ability was significantly enhanced. The number of positive cells increased significantly and the DNA replication activity was significantly enhanced. LncRNA44154 can promote the proliferation of porcine preadipocytes by encouraging cells to enter the S phase of the cell cycle. The result of interfering with lncRNA44154 is the opposite. Further analysis revealed that lncRNA44154 regulates intracellular lipid metabolism, inhibits the production of intracellular lipid droplets, and inhibits preadipocyte differentiation. RNA-seq results indicate that lncRNA44154 may be involved in biological pathways such as AMPK, PPAR and cAMP signaling pathway. It may work by regulating the RPS4X gene. This research may provide a new perspective for investigating the regulation of adipose metabolism by lncRNAs.

138 Regulation of post protein translational modification in adipogenesis and fat accumulation in pigs

Abstract In this study, we explored the role of protein transcriptional modification (PTM) in the adipogenesis and fat accumulation of intramuscular adipocytes. Intramuscular fat (IMF) is mainly stored in adipocytes interspersed in the perimysial space or within fascicles, and its content is a key factor attributing to meat quality in farm animals, especially in terms of tenderness, flavor and juiciness. The early stage from the fetus to the neonate is crucial for skeletal muscle development involving myogenesis, adipogenesis, and fibrogenesis, and is susceptible to environmental factors. Previous studies have been subjected to explore the molecular mechanism involved in IMF development. However, it was obscured by muscular background of IMF adipocytes. Therefore, we investigated PTM dynamics from three aspects: 1) first analyzed PTM levels of lysine acetylation (Kac), 2-hydroxyisobutyrylation (Khib), crotonylation (Kcr) and succinylation (Ksu) in three distinct differentiation potenctial primary stem cells, including myogenic precursor cells (MPCs), fibro-adipogenic progenitors (FAPs) and MSCs isolated from neonatal longissimus dorsi muscle of pigs within 3-d of age; 2) Comparing the Khib levels of proliferative and differentiated FAPs; and 3) Exploring the role of Khib modification in the adipose tissue of obese- and lean-types of pigs. We found that only Khib level in FAPs was significantly greater than MSCs. During adipogenic differentiation of FAPs, the modification level of Khib was significantly increased quadratically in parallel with protein levels of acylation regulatory enzymes KAT5 (writer) and HDAC2 (Eraser). In comparison of Khib levels in FAPs in proliferation and in differentiation and Khib levels in backfat tissues from obese pigs (Laiwu pigs) and lean pigs (Duroc pigs), we also demonstrated that Khib modification level was positive correlated with adipogenic differentiation and fat accumulation. Accordingly, the expression patterns of KAT5 and HDAC2 were parallel to the degree of lipid accumulation in the backfat adipose tissue of finishing pigs. Furthermore, we showed that KAT5 knockdown in FAPs inhibited adipogenic differentiation, while HDAC2 knockdown enhanced adipogenic differentiation of FAPs. By employing C3H10T1/2 cells, we identified EON1 involved in Khib modification in regulation of adipo gesis and fat accumulation with analysis of Liquid chromatography-tandem mass spectrometry (LC-MS/MS) using the Tandem Mass Tag (TMT Through conditional loss- and gain-of-function mutations, we further demonstrated that khib modification of ENO1 is necessary for preadipocyte differentiation and subsequent fat accumulation by mediating cellular glycolysis. In conclusion, we are the first to reveal the role of Khib modification in adipogenesis and fat deposition in pigs, and provided new clues for the improvement of fat accumulation and distribution via genetic selection and nutritional strategy in pigs.

Genome-Wide Analysis Reveals Copy Number Variant Gene TGFBR3 Regulates Pig Back Fat Deposition.

BFT is closely related to meat quality and lean meat percentage in pigs. The BFT traits of European LW pigs significantly differ from those of Chinese indigenous fatty MZ pigs. CNV is a prevalent genetic variation that plays an important role in economically important traits in pigs. However, the potential contribution of CNV to BFT in LW and MZ pigs remains unclear. In this study, whole-genome CNV detection was performed using next-generation sequencing data from LW and MZ pigs, and transcriptome data from back fat tissue of 180-day-old LW and MZ pigs were integrated for expression quantitative trait loci (eQTL) analysis. We identified a copy number variation in the TGFBR3 gene associated with BFT, showing a dose effect between the genome and transcriptome levels of the TGFBR3 gene. In porcine preadipocytes, TGFBR3 expression continuously increased during differentiation. Knockdown of TGFBR3 using specific siRNA inhibited preadipocyte differentiation and proliferation. Our study provides insights into the genetic regulation of pork quality and offers a theoretical basis for improving carcass quality by modulating BFT in pigs.

Edible Vitalmelon Fruit Extract Promotes Lipolysis of MDI-stimulated 3T3-L1 Adipocytes Through the Stimulation of Glycerol Release and β-oxidation

Background and Purpose: The anti-obesity effects of vitalmelon extracts have been well verified in only adipogenesis and lipogenesis of adipocytes and high-fat diet-induced obesity mice except lipolysis. Therefore, we were investigated the novel function of edible vitalmelon fruit water extract (VW) on lipolysis. Methods: The changes in the free glycerol release, cyclic adenosine monophosphate (cAMP)-dependent signaling pathway, and expression of β-oxidation-related regulators were analyzed in 3‐isobutyl‐1‐methylxanthine, dexamethasone, and insulin (MDI)-stimulated 3T3‐L1 adipocytes (MiSt-3T3 adipocytes) after a VW treatment. Results: The successful differentiation of pre-adipocytes was confirmed with increased oil-red O (ORO)-stained lipid accumulation and PPARγ gene transcription. After maturation of adipocytes, the levels of free glycerol, intracellular cAMP, and hormone-sensitive lipase phosphorylation increased in a dose-dependent manner in the MDI+VW-treated group. In addition, the protein and mRNA levels of three β-oxidation-related genes (ACADs, ACO1, and CTP) were remarkably increased in the MiSt-3T3 adipocytes after VW treatment, while ATPCL phosphorylation was decreased. The changes in the expression levels of these proteins were accompanied by alteration on the expression levels of the PPARα transcription factors. The levels of PPARα protein phosphorylation and mRNA transcription in MiSt-3T3 adipocytes increased dose-dependently after the VW treatment. Conclusion: These results show that VW can promote lipolysis by controlling the cAMP-mediated glycerol release pathway and β-oxidation in MiSt-3T3 adipocytes.

CTGF Inhibits the Differentiation of Chicken Preadipocytes via the TGFβ/Smad3 Signaling Pathway or by Inducing the Expression of ACTG2.

Chicken is the main source of protein for humans in most parts of the world. However, excessive fat deposition in chickens has become a serious problem. This adversely affects the growth of chickens and causes economic losses. Fat formation mainly occurs through preadipocyte differentiation, and excessive fat deposition results from the accumulation of preadipocytes after differentiation. Our previous studies have found that the connective tissue growth factor (CTGF) may be an important candidate gene for fat deposition. However, its function and mechanism in preadipocyte differentiation are still unclear. In this study, the RT-qPCR and Western blot results showed that the expression of CTGF mRNA and protein in the abdominal adipose of lean chickens was significantly higher than that of fat chickens. Therefore, we studied the function and mechanism of the CTGF in the differentiation of chicken preadipocytes. Functionally, the CTGF inhibited the differentiation of chicken preadipocytes. Mechanistically, the CTGF mediated the TGFβ1/Smad3 signaling pathway, thereby inhibiting the differentiation of chicken preadipocytes. In addition, we used the unique molecular identifier (UMI) RNA-Seq technology to detect genes that can be regulated by the CTGF in the whole genome. Through transcriptome data analysis, we selected actin gamma 2 (ACTG2) as a candidate gene. Regarding the function of the ACTG2 gene, we found that it inhibited the differentiation of chicken preadipocytes. Furthermore, we found that the CTGF can inhibit the differentiation of preadipocytes through the ACTG2 gene. In summary, this study found the CTGF as a new negative regulator of chicken preadipocyte differentiation. The results of this study help improve the understanding of the molecular genetic mechanism of chicken adipose tissue growth and development and also have reference significance for the study of human obesity.

IGFBP7 promotes the proliferation and differentiation of primary myoblasts and intramuscular preadipocytes in chicken

Though it is well known that insulin-like growth factor (IGF) binding protein 7 (IGFBP7) plays an important role in myogenesis and adipogenesis in mammals, its impact on the proliferation, differentiation, and lipid deposition in chicken primary myoblasts (CPM) and intramuscular preadipocytes remains unexplored. In the present study, we firstly examined the correlation between SNPs within the genomic sequence of the IGFBP7 gene and carcass and blood chemical traits in a F2 resource population by genetic association analysis, and found that a significant correlation between the SNP (4_49499525) located in the intron region of IGFBP7 and serum high-density lipoproteins (HDL). We then examined the expression patterns of IGFBP7 across different stages of proliferation and differentiation in CPMs and intramuscular preadipocytes via qPCR, and explored the biological functions of IGFBP7 through gain- and loss-of-function experiments and a range of techniques including qPCR, CCK-8, EdU, flow cytometry, Western blot, immunofluorescence, and Oil Red O staining to detect the proliferation, differentiation, and lipid deposition in CPMs and intramuscular preadipocytes. We ascertained that the expression levels of the IGFBP7 gene increased as cell differentiation progresses in CPMs and intramuscular preadipocytes, and that IGFBP7 promotes the proliferation and differentiation of these cells, as well as facilitates intracellular lipid deposition. Furthermore, we investigated the regulatory mechanism of IGFBP7 expression by using co-transfection strategy and dual-luciferase reporter assay, and discovered that the myogenic transcription factors (MRF), myoblast determination factor (MyoD) and myogenin (MyoG), along with the adipocyte-specific transcription factor (TF) CCAAT/enhancer-binding protein α (C/EBPα), can bind to the core transcription activation region of the IGFBP7 promoter located 500 bp upstream from the transcription start site, thereby promoting IGFBP7 transcription and expression. Taken together, our study underscores the role of IGFBP7 as a positive regulator for myogenesis and adipogenesis, while also elucidating the functional and transcriptional regulatory mechanisms of IGFBP7 in chicken skeletal muscle development and intramuscular adipogenesis.

A natural small molecule pinocembrin resists high-fat diet-induced obesity through GPR120-ERK1/2 pathway

Obesity is a widely concerned health problem. Mobilizing white adipose tissue and reducing fat synthesis are considered as effective strategies in the treatment of obesity. Here, using Connectivity Map (CMap) approach, we identified the pinocembrin (PB), a natural flavonoid primarily found in propolis, as a potential anti-obesity drug. Therefore, high-fat-diet (HFD) mice were randomly divided into two groups and fed a HFD or HFD with PB in this study. In vivo experiments showed that supplementation of PB reduced the body weight gain and ameliorated insulin resistance in HFD-induced mice. More importantly, PB did not cause side effect through detecting the levels of alanine transaminase (ALT), aspartate aminotransferase (AST), creatinine (CRE) and blood urea nitrogen (BUN) in serum of mice. Additionally, PB reduced expansion of white adipose tissue with upregulation of genes related lipolysis and downregulation of genes related lipogenesis. Furthermore, in vitro experiments revealed that PB treatment dose-dependently inhibited lipid droplet formation with upregulation of genes related lipolysis and downregulation of genes related lipogenesis. Molecular docking analysis combined with cellular thermal shift assay (CETSA) suggested that PB has a high affinity to the G protein-coupled receptor 120 (GPR120). Meanwhile, we confirmed that PB efficiently inhibited adipogenic differentiation of preadipocytes by directly binding to GPR120 and subsequently activating the downstream phosphorylation extracellular regulated kinase 1/2 (ERK1/2). Collectively, PB exerted anti-obesity effect through GPR120-ERK1/2 signaling pathway, providing a novel and promising natural drug for the treatment of obesity.

MiR-196a Promotes Lipid Deposition in Goat Intramuscular Preadipocytes by Targeting MAP3K1 and Activating PI3K-Akt Pathway.

Meat quality in goats is partly determined by the intramuscular fat (IMF) content, which is associated with the proliferation and differentiation of intramuscular preadipocytes. Emerging studies have suggested that miRNA plays a crucial role in adipocyte proliferation and differentiation. In our recent study, we observed the expression variations in miR-196a in the longissimus dorsi muscle of Jianzhou goats at different ages. However, the specific function and underlying mechanism of miR-196a in IMF deposition are still unclear. This study demonstrated that miR-196a significantly enhanced adipogenesis and apoptosis and reduced the proliferation of preadipocytes. Subsequently, RNA-seq was employed to determine genes regulated by miR-196a, and 677 differentially expressed genes were detected after miR-196a overexpression. The PI3K-Akt pathway was identified as activated in miR-196a regulating intramuscular adipogenesis via Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and further verified via Western blot and rescue assays. Lastly, using RT-qPCR, Western blot, dual-luciferase, and rescue assays, we found that miR-196a promoted adipogenesis and suppressed the proliferation of intramuscular preadipocytes by the downregulation of MAP3K1. In summary, these results suggest that miR-196a regulates IMF deposition by targeting MAP3K1 and activating the PI3K-Akt pathway and provide a theoretical foundation for improving goat meat quality through molecular breeding.

Inhibitory Effect and Mechanism of Epigallocatechin Gallate on the Differentiation of 3T3-L1 Preadipocytes.

Green tea possesses a range of beneficial effects, including anti-obesity, antioxidant, and anti-inflammatory properties, owing to its biologically active components, primarily catechins such as epicatechin (EC), epicatechin gallate (ECG), epigallocatechin (EGC), and epigallocatechin gallate (EGCG). However, few studies have investigated the four catechin monomers simultaneously, and the molecular mechanisms of their anti-obesity effects have not been fully elucidated. In this study, we investigated the effects of four catechin monomers on the differentiation of 3T3-L1 preadipocytes of mice. Our findings demonstrated that four catechin monomers EC/ECG/EGC/EGCG (12, 25, 50 µM) dose-dependently inhibited the differentiation of 3T3-L1 preadipocytes and reduced triglyceride content. EGCG exhibited the most potent inhibitory effect with an optimal concentration of 50 µM. In addition, transcriptome sequencing and lipidomic analysis of EGCG-treated 3T3-L1 preadipocytes revealed that Ptgs2 and Pim1 were the most differentially expressed genes involved in regulating adipocyte differentiation. The results suggested that EGCG up-regulated the expression of the Pla2g2e gene and down-regulated the expression of the Pla2g4a and Pla2g2a genes via the glycerophospholipid metabolic pathway, which subsequently elevated lysophosphatidylcholine (LPC) levels, influencing the differentiation process of 3T3-L1 preadipocytes.

Endothelin 3/EDNRB signaling induces thermogenic differentiation of white adipose tissue

Thermogenic adipose tissue, consisting of brown and beige fat, regulates nutrient utilization and energy metabolism. Human brown fat is relatively scarce and decreases with obesity and aging. Hence, inducing thermogenic differentiation of white fat offers an attractive way to enhance whole-body metabolic capacity. Here, we show the role of endothelin 3 (EDN3) and endothelin receptor type B (EDNRB) in promoting the browning of white adipose tissue (WAT). EDNRB overexpression stimulates thermogenic differentiation of human white preadipocytes through cAMP-EPAC1-ERK activation. In mice, cold induces the expression of EDN3 and EDNRB in WAT. Deletion of EDNRB in adipose progenitor cells impairs cold-induced beige adipocyte formation in WAT, leading to excessive weight gain, glucose intolerance, and insulin resistance upon high-fat feeding. Injection of EDN3 into WAT promotes browning and improved whole-body glucose metabolism. The findings shed light on the mechanism of WAT browning and offer potential therapeutics for obesity and metabolic disorders.