Lipid-associated macrophages reshape BAT cell identity in obesity.

Obesity and type 2 diabetes cause a loss in brown adipose tissue (BAT) activity, but the molecular mechanisms that drive BAT cell remodeling remain largely unexplored. Using a multilayered approach, we comprehensively mapped a reorganization in BAT cells. We uncovered a subset of macrophages as lipid-associated macrophages (LAMs), which were massively increased in genetic and dietary model of BAT expansion. LAMs participate in this scenario by capturing extracellular vesicles carrying damaged lipids and mitochondria released from metabolically stressed brown adipocytes. CD36 scavenger receptor drove LAM phenotype, and CD36-deficient LAMs were able to increase brown fat genes in adipocytes. LAMs released transforming growth factor ß1 (TGF-ß1), which promoted the loss of brown adipocyte identity through aldehyde dehydrogenase 1 family member A1 (Aldh1a1) induction. These findings unfold cell dynamic changes in BAT during obesity and identify LAMs as key responders to tissue metabolic stress and drivers of loss of brown adipocyte identity.

Gene expression and characterization of clonally derived murine embryonic brown and brite adipocytes.

White adipocytes store energy, while brown and brite adipocytes release heat via nonshivering thermogenesis. In this study, we characterized two murine embryonic clonal preadipocyte lines, EB5 and EB7, each displaying unique gene marker expression profiles. EB5 cells differentiate into brown adipocytes, whereas EB7 cells into brite (also known as beige) adipocytes. To draw a comprehensive comparison, we contrasted the gene expression patterns, adipogenic capacity, as well as carbohydrate and lipid metabolism of these cells to that of F442A, a well-known white preadipocyte and adipocyte model. We found that commitment to differentiation in both EB5 and EB7 cells can be induced by 3-Isobutyl-1-methylxanthine/dexamethasone (Mix/Dex) and staurosporine/dexamethasone (St/Dex) treatments. Additionally, the administration of rosiglitazone significantly enhances the brown and brite adipocyte phenotypes. Our data also reveal the involvement of a series of genes in the transcriptional cascade guiding adipogenesis, pinpointing GSK3ß as a critical regulator for both EB5 and EB7 adipogenesis. In a developmental context, we observe that, akin to brown fat progenitors, brite fat progenitors make their appearance in murine development by 11-12 days of gestation or potentially earlier. This result contributes to our understanding of adipocyte lineage specification during embryonic development. In conclusion, EB5 and EB7 cell lines are valuable for research into adipocyte biology, providing insights into the differentiation and development of brown and beige adipocytes. Furthermore, they could be useful for the characterization of drugs targeting energy balance for the treatment of obesity and metabolic diseases.

MORPHOLOGICAL DIFFERENCES BETWEEN THE LESSER AND THE GREATER OMENTA IN ALBINO RATS.

BACKGROUND: Publications report that all mammals have two omenta, namely, lesser omentum and greater omentum. Basically, these organs, which share the same name except for the adjective "lesser" or "greater," should not differ from each other. However, no clear description of the structure of the lesser omentum, as well as comparative morphological analysis between the lesser and greater omenta have been found in the literature, which necessitates a thorough investigation. Therefore, the aim of our study was to analyze the morphofunctional differences between the greater and lesser omenta in albino rats. METHOD: The experiment involved 20 mature male albino rats, weighing 298,28±7,36grams. The material for our study were preparations of lesser and greater omenta, fixed in 10% of neutral buffered formalin. Paraffin sections were stained with hematoxylin-eosin and Van Gieson stain. RESULTS: The findings of the study showed that the greater omentum in albino rats, unlike other derivatives of the omentum (ligaments and mesenteries), represents a free extension (mostly from the greater curvature of the stomach), in the form of an "apron," into a specific depth of the peritoneal cavity, duplicating the serous membrane. This duplication is characterized by the composition of two structurally interdependent formations. These include vascular-fatty arcades, associated with lymphoid nodules known as milky spots, and binding serous-reticular membranes. The findings of the study of the lesser omentum have established that in all cases it is located beneath the liver and becomes visualized only after hepatolifting. It is presented in the form of two ligaments: hepatoduodenal and hepatogastric, which contain two main structured formations, which we called vascular-fatty spurs, between these spurs, serous-reticular membranes are located. CONCLUSION: despite having similar names, the lesser omentum, a derivative of the peritoneum, is fundamentally different. As it is well known, the lesser omentum is represented by ligaments that extend from the liver hilus to the lesser curvature of the stomach and the duodenum. Due to this arrangement, the lesser omentum lacks the mobile activity characteristic of the greater omentum, which plays a crucial role in rapid response to damage in the gastrointestinal tract. Despite sharing the same names, both formations differ in shape, morphological structure, development and function.

Obesity-induced chronic low-grade inflammation in adipose tissue: A pathway to Alzheimer's disease.

Alzheimer's disease (AD) is a leading cause of cognitive impairment worldwide. Overweight and obesity are strongly associated with comorbidities, such as hypertension, diabetes, and insulin resistance (IR), which contribute substantially to the development of AD and subsequent morbidity and mortality. Adipose tissue (AT) is a highly dynamic organ composed of a diverse array of cell types, which can be classified based on their anatomic localization or cellular composition. The expansion and remodeling of AT in the context of obesity involves immunometabolic and functional shifts steered by the intertwined actions of multiple immune cells and cytokine signaling within AT, which contribute to the development of metabolic disorders, IR, and systemic markers of chronic low-grade inflammation. Chronic low-grade inflammation, a prolonged, low-dose stimulation by specific immunogens that can progress from localized sites and affect multiple organs throughout the body, leads to neurodystrophy, increased apoptosis, and disruption of homeostasis, manifesting as brain atrophy and AD-related pathology. In this review, we sought to elucidate the mechanisms by which AT contributes to the onset and progression of AD in obesity through the mediation of chronic low-grade inflammation, particularly focusing on the roles of adipokines and AT-resident immune cells.

Silybin restores glucose uptake after tumour necrosis factor-alpha and lipopolysaccharide stimulation in 3T3-L1 adipocytes.

Obesity is associated with a low-grade chronic inflammatory process characterized by higher circulating TNFα levels, thus contributing to insulin resistance. This study evaluated the effect of silybin, the main bioactive component of silymarin, which has anti-inflammatory properties, on TNFα levels and its impact on glucose uptake in the adipocyte cell line 3T3-L1 challenged with two different inflammatory stimuli, TNFα or lipopolysaccharide (LPS). Silybin's pre-treatment effect was evaluated in adipocytes pre-incubated with silybin (30 or 80 µM) before challenging with the inflammatory stimuli (TNFα or LPS). For the post-treatment effect, the adipocytes were first challenged with the inflammatory stimuli and then post-treated with silybin. After treatments, TNFα production, glucose uptake, and GLUT4 protein expression were determined. Both inflammatory stimuli increased TNFα secretion, diminished GLUT4 expression, and significantly decreased glucose uptake. Silybin 30 µM only reduced TNFα secretion after the LPS challenge. Silybin 80 µM as post-treatment or pre-treatment decreased TNFα levels, improving glucose uptake. However, glucose uptake enhancement induced by silybin did not depend on GLUT4 protein expression. These results show that silybin importantly reduced TNFα levels and upregulates glucose uptake, independently of GLUT4 protein expression.

Therapeutic potential of dedifferentiated fat cells in a rat model of osteoarthritis of the knee.

Introduction: Mature adipocyte-derived dedifferentiated fat cells (DFATs) represent a subtype of multipotent cells that exhibit comparable phenotypic and functional characteristics to adipose-derived stem cells (ASCs). In this study, we assessed the chondroprotective properties of intra-articularly administrated DFATs in a rat model of osteoarthritis (OA). We also investigated in vitro the expression of anti-inflammatory and chondroprotective genes in DFATs prepared from the infrapatellar fat pad (IFP) and subcutaneous adipose-tissue (SC) of human origin. Methods: In the cell transplantation experiment, rats were assigned to the DFAT and Control group (n = 10 in each group) and underwent anterior cruciate ligament transection (ACLT) accompanied by medial meniscus resection (MMx) to induce OA. One week later, they received intra-articular injections of 1 × 106 DFATs (DFAT group) or PBS (control group) four times, with a weekly administration frequency. Macroscopic and microscopic evaluations were conducted five weeks post-surgery. In the in vitro experiments. DFATs derived from the IFP (IFP-DFATs) and SC (SC-DFATs) were prepared from donor-matched tissue samples (n = 3). The gene expression of PTGS2, TNFAIP6, PRG4, BMP2, and BMP6 under TNF-α or IFN-γ stimulation in these cells was evaluated using RT-PCR. Furthermore, the effect of co-culturing synovial fibroblasts with DFATs on the gene expression of ADAMTS4 and IL-6 were evaluated. Results: Intra-articular injections of DFATs significantly inhibited cartilage degeneration in the rat OA model induced by ACLT and MMx. RT-PCR analysis revealed that both IFP-DFATs and SC-DFATs upregulated the expression of genes involved in immune regulation, anti-inflammation, and cartilage protection such as PTGS2, TNFAIP6, and BMP2, under stimulation by inflammatory cytokines. Co-culture with DFATs suppressed the expression of ADAMTS4 and IL6 in synovial fibroblasts. Conclusions: The intra-articular injection of DFATs resulted in chondroprotective effects in the rat OA model. Both SC-DFATs and IFP-DFATs induced the expression of anti-inflammatory and chondroprotective genes in vitro. These results indicate that DFATs appear to possess therapeutic potential in inhibiting cartilage degradation and could serve as a promising cellular resource for OA treatment.

Alternative isoform expression of key thermogenic genes in human beige adipocytes.

Background: The beneficial effect of thermogenic adipocytes in maintaining body weight and protecting against metabolic disorders has raised interest in understanding the regulatory mechanisms defining white and beige adipocyte identity. Although alternative splicing has been shown to propagate adipose browning signals in mice, this has yet to be thoroughly investigated in human adipocytes. Methods: We performed parallel white and beige adipogenic differentiation using primary adipose stem cells from 6 unrelated healthy subjects and assessed differential gene and isoform expression in mature adipocytes by RNA sequencing. Results: We find 777 exon junctions with robust differential usage between white and beige adipocytes in all 6 subjects, mapping to 562 genes. Importantly, only 10% of these differentially spliced genes are also differentially expressed, indicating that alternative splicing constitutes an additional layer of gene expression regulation during beige adipocyte differentiation. Functional classification of alternative isoforms points to a gain of function for key thermogenic transcription factors such as PPARG and CITED1, and enzymes such as PEMT, or LPIN1. We find that a large majority of the splice variants arise from differential TSS usage, with beige-specific TSSs being enriched for PPARγ and MED1 binding compared to white-specific TSSs. Finally, we validate beige specific isoform expression at the protein level for two thermogenic regulators, PPARγ and PEMT. Discussion: These results suggest that differential isoform expression through alternative TSS usage is an important regulatory mechanism for human adipocyte thermogenic specification.

Increased obesogenic action of palmitic acid during early stage of adipogenesis.

The functional differences between preadipocytes and fully differentiated mature adipocytes derived from stromal vascular fraction stem cells, as well as primary adipocytes have been analysed by evaluating their response to the obesogenic factor (a saturated fatty acid) and TNF-triggered inflammation. The analysis of single adipocytes shows that the saturated fatty acid (palmitic acid) accumulation is accompanied by inflammation and considerably dependent on the stage of the adipogenesis. In particular, preadipocytes show the exceptional potential for palmitic acid uptake resulting in their hypertrophy and the elevated cellular expression of the inflammation marker (ICAM-1). Our research provides new information on the impact of obesogenic factors on preadipocytes that is important in the light of childhood obesity prevention.

The role of Sirtuin 1 in regulation of fibrotic genes expression in pre-adipocytes.

Purpose: Considering inhibition of pre-adipocyte cells differentiation in adipose tissue fibrosis, we aimed to explore whether Sirt1 and Hif-1α in pre-adipocytes have a significant effect on fibrotic gene expression. Methods: 3T3-L1 pre-adipocytes were transfected with SIRT1-specific siRNA, confirmed by real-time polymerase chain reaction (RT-PCR) and western blotting. Additionally, cells were treated with varying concentrations of resveratrol and sirtinol as the activator and inhibitor of Sirt1, respectively. Involvement of Hif-1α was evaluated by treatment with echinomycin. Subsequently, we assessed the gene and protein expressions related to fibrosis in the extracellular matrix of adipose tissue, including collagen VI (Col VI), lysyl oxidase (Lox), matrix metalloproteinase-2 (Mmp-2), Mmp-9, and osteopontin (Opn) in pre-adipocytes through RT-PCR and western blot. Results: The current study demonstrated that Sirt1 knockdown and reduced enzyme activity significantly increased the expression of Col VI, Lox, Mmp-2, Mmp-9, and Opn genes in the treated 3T3-L1 cells compared to the control group. Interestingly, resveratrol significantly decreased the gene expression related to the fibrosis pathway. Inhibition of Hif-1α by echinomycin led to a significant reduction in Col VI, Mmp-2, and Mmp-9 gene expression in the treated group compared to the control. Conclusion: This study highlights that down-regulation of Sirt1 might be a predisposing factor in the emergence of adipose tissue fibrosis by enhancing the expression of extracellular matrix (ECM) components. Activation of Sirt1, similar to suppressing of Hif-1α in pre-adipocytes may be a beneficial approach for attenuating fibrotic gene expression. Supplementary Information: The online version contains supplementary material available at 10.1007/s40200-024-01389-4.

CTRP9 Promotes Brown Adipose Tissue Lipolysis in Mice Fed a High-Fat Diet.

BACKGROUND: This study aimed to elucidate the molecular mechanism through which C1q/tumor necrosis factor (TNF)-related protein 9 (CTRP9) acts in the formation and differentiation of brown adipose tissue (BAT). METHODS: Adenovirus particles encoding CTRP9 and green fluorescent protein were inoculated into the scapula of C57BL/6J mice and fed a high-fat diet for 8 weeks; the body weight, lipid droplet morphology, glucose tolerance, insulin tolerance, and protein expression levels were analyzed. In addition, CTRP9 adenovirus was transfected into brown preadipocytes, and differentiation was induced to identify the effect of CTRP9 overexpression on adipocyte differentiation. RESULTS: CTRP9 overexpression significantly increased the weight gain of mice. Additionally, the CTRP9 overexpression group exhibited significantly increased adipose tissue weight and glucose clearance rates and decreased insulin sensitivity and serum triglyceride levels compared to the control group. Furthermore, CTRP9 overexpression significantly upregulated the adipose triglyceride lipase (ATGL) and perilipin 1 protein expression levels in BAT. The cell experiment results confirmed that CTRP9 overexpression significantly inhibited the adipogenesis of brown adipocytes as evidenced by the downregulation of uncoupling protein 1, beta-3 adrenergic receptor, ATGL, and hormone-sensitive lipase mRNA levels and the significant suppression of uncoupling protein 1, ATGL, and perilipin 1 protein levels in brown adipocytes. CONCLUSIONS: The finding of this study demonstrated that CTRP9 promotes lipolysis by upregulating ATGL expression in vivo and inhibits the differentiation of brown preadipocytes in vitro.

Red Oranges and Olive Leaf Waste-Derived Bioactive Extracts Promote Adipocyte Functionality In Vitro.

Obesity is increasingly prevalent worldwide and is linked to metabolic diseases, such as insulin resistance (IR) and type 2 diabetes mellitus (T2DM), due to excessive free fatty acids (FFAs). Although lifestyle changes are effective, they often prove to be insufficient as initial treatments for obesity. Additionally, while surgical and pharmacological interventions are available, they are not entirely safe or effective. Recently, interest has grown in utilizing food waste and plant-derived phenolic compounds for their health benefits, presenting a promising avenue for managing obesity and its related disorders. Indeed, many studies have examined the potential inhibitory effects of the natural extract on adipocyte differentiation and lipid accumulation. This study focused on the evaluation of the effects of standardized extracts obtained from red oranges and olive leaf waste on 3T3-L1 murine pre-adipocyte and adipocyte functionality. Red orange extract (ROE) and olive leaf extract (OLE), alone and in combination, were tested to assess their anti-obesity and anti-inflammatory effects, as well as their potential therapeutic benefits. Three in vitro models were established to investigate the effects of the extracts on (I) adipocyte differentiation; (II) mature and hypertrophic adipocytes challenged with palmitic acid (PA) and erastin (ER), respectively; and (III) erastin-induced cytotoxicity on pre-adipocytes.

The Expression of a Subset of Aging and Antiaging Markers Following the Chondrogenic and Osteogenic Differentiation of Mesenchymal Stem Cells of Placental Origin.

Mesenchymal stem cells (MSCs) of placental origin hold great promise in tissue engineering and regenerative medicine for diseases affecting cartilage and bone. However, their utility has been limited by their tendency to undergo premature senescence and phenotypic drift into adipocytes. This study aimed to explore the potential involvement of a specific subset of aging and antiaging genes by measuring their expression prior to and following in vitro-induced differentiation of placental MSCs into chondrocytes and osteoblasts as opposed to adipocytes. The targeted genes of interest included the various LMNA/C transcript variants (lamin A, lamin C, and lamin A∆10), sirtuin 7 (SIRT7), and SM22α, along with the classic aging markers plasminogen activator inhibitor 1 (PAI-1), p53, and p16INK4a. MSCs were isolated from the decidua basalis of human term placentas, expanded, and then analyzed for phenotypic properties by flow cytometry and evaluated for colony-forming efficiency. The cells were then induced to differentiate in vitro into chondrocytes, osteocytes, and adipocytes following established protocols. The mRNA expression of the targeted genes was measured by RT-qPCR in the undifferentiated cells and those fully differentiated into the three cellular lineages. Compared to undifferentiated cells, the differentiated chondrocytes demonstrated decreased expression of SIRT7, along with decreased PAI-1, lamin A, and SM22α expression, but the expression of p16INK4a and p53 increased, suggesting their tendency to undergo premature senescence. Interestingly, the cells maintained the expression of lamin C, which indicates that it is the primary lamin variant influencing the mechanoelastic properties of the differentiated cells. Notably, the expression of all targeted genes did not differ from the undifferentiated cells following osteogenic differentiation. On the other hand, the differentiation of the cells into adipocytes was associated with decreased expression of lamin A and PAI-1. The distinct patterns of expression of aging and antiaging genes following in vitro-induced differentiation of MSCs into chondrocytes, osteocytes, and adipocytes potentially reflect specific roles for these genes during and following differentiation in the fully functional cells. Understanding these roles and the network of signaling molecules involved can open opportunities to improve the handling and utility of MSCs as cellular precursors for the treatment of cartilage and bone diseases.

Long Noncoding RNA 6302 Regulates Chicken Preadipocyte Differentiation by Targeting SLC22A16.

The excessive deposition of abdominal adipocytes in chickens is detrimental to poultry production. However, the regulatory factors that affect abdominal adipogenesis in chickens are still poorly understood. SLC22A16 is differentially expressed in abdominal preadipocytes and 10-day differentiated adipocytes in chickens, but its role in regulating chicken adipogenesis has not been reported. In this study, the function of SLC22A16 in chicken abdominal preadipocytes was investigated. SLC22A16 is significantly upregulated during abdominal adipocyte differentiation. The overexpression of SLC2A16 upregulated the expression of adipogenic marker genes and proliferation-related genes, and promoted the proliferation of adipocytes and the accumulation of triglycerides. The knockdown of SLC22A16 downregulated the expression of adipogenic marker genes and proliferation-related genes, inhibited the proliferation of adipocytes, and impaired the accumulation of triglycerides in adipocytes. In addition, LNC6302 was differentially expressed in abdominal preadipocytes and mature adipocytes, and was significantly positively correlated with the expression of SLC22A16. Interference with LNC6302 inhibits the expression of adipogenic marker genes and proliferation-related genes. The data supported the notion that LNC6302 promotes the differentiation of chicken abdominal adipocytes by cis-regulating the expression of SLC22A16. This study identified the role of SLC22A16 in the differentiation and proliferation of chicken adipocytes, providing a potential target for improving abdominal adipogenesis in chickens.

Adipo-Epithelial Transdifferentiation in In Vitro Models of the Mammary Gland.

Subcutaneous adipocytes are crucial for mammary gland epithelial development during pregnancy. Our and others' previous data have suggested that adipo-epithelial transdifferentiation could play a key role in the mammary gland alveolar development. In this study, we tested whether adipo-epithelial transdifferentiation occurs in vitro. Data show that, under appropriate co-culture conditions with mammary epithelial organoids (MEOs), mature adipocytes lose their phenotype and acquire an epithelial one. Interestingly, even in the absence of MEOs, extracellular matrix and diffusible growth factors are able to promote adipo-epithelial transdifferentiation. Gene and protein expression studies indicate that transdifferentiating adipocytes exhibit some characteristics of milk-secreting alveolar glands, including significantly higher expression of milk proteins such as whey acidic protein and ß-casein. Similar data were also obtained in cultured human multipotent adipose-derived stem cell adipocytes. A miRNA sequencing experiment on the supernatant highlighted mir200c, which has a well-established role in the mesenchymal-epithelial transition, as a potential player in this phenomenon. Collectively, our data show that adipo-epithelial transdifferentiation can be reproduced in in vitro models where this phenomenon can be investigated at the molecular level.

Calmodulin Contributes to Lipolysis and Inflammatory Responses in Clinical Ketosis Cows through the TLR4/IKK/NF-κB Pathway.

Clinical ketosis is a detrimental metabolic disease in dairy cows, often accompanied by severe lipolysis and inflammation in adipose tissue. Our previous study suggested a 2.401-fold upregulation in the calmodulin (CaM) level in the adipose tissue of cows with clinical ketosis. Therefore, we hypothesized that CaM may regulate lipolysis and inflammatory responses in cows with clinical ketosis. To verify the hypothesis, we conducted a thorough veterinary assessment of clinical symptoms and serum ß-hydroxybutyrate (BHB) concentration. Subsequently, we collected subcutaneous adipose tissue samples from six healthy and six clinically ketotic Holstein cows at 17 ± 4 days postpartum. Commercial kits were used to test the abundance of BHB, non-esterified fatty acid (NEFA), the liver function index (LFI), interleukin-6 (IL-6), IL-1ß, and tumor necrosis factor-α (TNF-α). We found that cows with clinical ketosis exhibited higher levels of BHB, NEFA, LFI, IL-6, IL-1ß, TNF-α, and lower glucose levels than healthy cows. Furthermore, the abundance of CaM, toll-like receptor 4 (TLR4), inhibitor of nuclear factor κB kinase subunit ß (IKK), phosphorylated nuclear factor κB p65/nuclear factor κB p65 (p-NF-κB p65/NF-κB p65), adipose triacylglycerol lipase (ATGL), and phosphorylated hormone-sensitive lipase/hormone-sensitive lipase (p-HSL/HSL) was increased, while that of perilipin-1 (PLIN1) was decreased in the adipose tissue of cows with clinical ketosis. To investigate the mechanism underlying the responses, we isolated the primary bovine adipocytes from the adipose tissue of healthy cows and induced the inflammatory response mediated by TLR4/IKK/NF-κB p65 with lipopolysaccharide (LPS). Additionally, we treated the primary bovine adipocytes with CaM overexpression adenovirus and CaM small interfering RNA. In vitro, LPS upregulated the abundance of TLR4, IKK, p-NF-κB p65, ATGL, p-HSL/HSL, and CaM and downregulated PLIN1. Furthermore, CaM silencing downregulated the abundance of LPS-activated p-HSL/HSL, TLR4, IKK, and p-NF-κB p65 and upregulated PLIN1 in bovine adipocytes, except for ATGL. However, CaM overexpression upregulated the abundance of LPS-activated p-HSL/HSL, TLR4, IKK, and p-NF-κB p65 and downregulated PLIN1 expression in bovine adipocytes. These data suggest that CaM promotes lipolysis in adipocytes through HSL and PINL1 while activating the TLR4/IKK/NF-κB inflammatory pathway to stimulate an inflammatory response. There is a positive feedback loop between CaM, lipolysis, and inflammation. Inhibiting CaM may act as an adaptive mechanism to alleviate metabolic dysregulation in adipose tissue, thereby relieving lipolysis and inflammatory responses.

Anti­obesity and immunostimulatory activity of Chrysosplenium flagelliferum in mouse preadipocytes 3T3­L1 cells and mouse macrophage RAW264.7 cells.

Chrysosplenium flagelliferum (CF) is known for its anti-inflammatory, antioxidant and antibacterial activities. However, there is a lack of research on its other pharmacological properties. In the present study, the bifunctional roles of CF in 3T3-L1 and RAW264.7 cells were investigated, focusing on its anti-obesity and immunostimulatory effects. In 3T3-L1 cells, CF effectively mitigated the accumulation of lipid droplets and triacylglycerol. Additionally, CF downregulated the peroxisome proliferator-activated receptor (PPAR)-γ and CCAAT/enhancer-binding protein α protein levels; however, this effect was impeded by the knockdown of ß-catenin using ß-catenin-specific small interfering RNA. Consequently, CF-mediated inhibition of lipid accumulation was also decreased. CF increased the protein levels of adipose triglyceride lipase and phosphorylated hormone-sensitive lipase, while decreasing those of perilipin-1. Moreover, CF elevated the protein levels of phosphorylated AMP-activated protein kinase and PPARγ coactivator 1-α. In RAW264.7 cells, CF enhanced the production of pro-inflammatory mediators, such as nitric oxide (NO), inducible NO synthase, interleukin (IL)-1ß, IL-6 and tumor necrosis factor-α, and increased their phagocytic capacities. Inhibition of Toll-like receptor (TLR)-4 significantly reduced the effects of CF on the production of pro-inflammatory mediators and phagocytosis, indicating its crucial role in facilitating these effects. CF-induced increase in the production of pro-inflammatory mediators was controlled by the activation of c-Jun N-terminal kinase (JNK) and nuclear factor (NF)-κB pathways, and TLR4 inhibition attenuated the phosphorylation of these kinases. The results of the pesent study suggested that CF inhibits lipid accumulation by suppressing adipogenesis and inducing lipolysis and thermogenesis in 3T3-L1 cells, while stimulating macrophage activation via the activation of JNK and NF-κB signaling pathways mediated by TLR4 in RAW264.7 cells. Therefore, CF simultaneously exerts both anti-obesity and immunostimulatory effects.

Exploring contrast-enhancing staining agents for studying adipose tissue through contrast-enhanced computed tomography.

Contrast-enhanced computed tomography offers a nondestructive approach to studying adipose tissue in 3D. Several contrast-enhancing staining agents (CESAs) have been explored, whereof osmium tetroxide (OsO4) is the most popular nowadays. However, due to the toxicity and volatility of the conventional OsO4, alternative CESAs with similar staining properties were desired. Hf-WD 1:2 POM and Hexabrix have proven effective for structural analysis of adipocytes using contrast-enhanced computed tomography but fail to provide chemical information. This study introduces isotonic Lugol's iodine (IL) as an alternative CESA for adipose tissue analysis, comparing its staining potential with Hf-WD 1:2 POM and Hexabrix in murine caudal vertebrae and bovine muscle tissue strips. Single and sequential staining protocols were compared to assess the maximization of information extraction from each sample. The study investigated interactions, distribution, and reactivity of iodine species towards biomolecules using simplified model systems and assesses the potential of the CESA to provide chemical information. (Bio)chemical analyses on whole tissues revealed that differences in adipocyte gray values post-IL staining were associated with chemical distinctions between bovine muscle tissue and murine caudal vertebrae. More specific, a difference in the degree of unsaturation of fatty acids was identified as a likely contributor, though not the sole determinant of gray value differences. This research sheds light on the potential of IL as a CESA, offering both structural and chemical insights into adipose tissue composition.

Single-Cell Landscape and a Macrophage Subset Enhancing Brown Adipocyte Function in Diabetes.

Background: Metabolic dysregulation is a hallmark of type 2 diabetes mellitus (T2DM), in which the abnormalities in brown adipose tissue (BAT) play important roles. However, the cellular composition and function of BAT as well as its pathological significance in diabetes remain incompletely understood. Our objective is to delineate the single-cell landscape of BAT-derived stromal vascular fraction (SVF) and their characteristic alterations in T2DM rats. Methods: T2DM was induced in rats by intraperitoneal injection of low-dose streptozotocin and high-fat diet feeding. Single-cell mRNA sequencing was then performed on BAT samples and compared to normal rats to characterize changes in T2DM rats. Subsequently, the importance of key cell subsets in T2DM was elucidated using various functional studies. Results: Almost all cell types in the BAT-derived SVF of T2DM rats exhibited enhanced inflammatory responses, increased angiogenesis, and disordered glucose and lipid metabolism. The multidirectional differentiation potential of adipose tissue-derived stem cells was also reduced. Moreover, macrophages played a pivotal role in intercellular crosstalk of BAT-derived SVF. A novel Rarres2+macrophage subset promoted the differentiation and metabolic function of brown adipocytes via adipose-immune crosstalk. Conclusion: BAT SVF exhibited strong heterogeneity in cellular composition and function and contributed to T2DM as a significant inflammation source, in which a novel macrophage subset was identified that can promote brown adipocyte function.

Upregulation of inflammatory genes and pathways links obesity to severe COVID-19.

Obesity is a risk factor for developing severe COVID-19. However, the mechanism underlying obesity-accelerated COVID-19 remains unclear. Here, we report results from a study in which 2-3-month-old K18-hACE2 (K18) mice were fed a western high-fat diet (WD) or normal chow (NC) over 3 months before intranasal infection with a sublethal dose of SARS-CoV2 WA1 (a strain ancestral to the Wuhan variant). After infection, the WD-fed K18 mice lost significantly more body weight and had more severe lung inflammation than normal chow (NC)-fed mice. Bulk RNA-seq analysis of lungs and adipose tissue revealed a diverse landscape of various immune cells, inflammatory markers, and pathways upregulated in the infected WD-fed K18 mice when compared with the infected NC-fed control mice. The transcript levels of IL-6, an important marker of COVID-19 disease severity, were upregulated in the lung at 6-9 days post-infection in the WD-fed mice when compared to NC-fed mice. Transcriptome analysis of the lung and adipose tissue obtained from deceased COVID-19 patients found that the obese patients had an increase in the expression of genes and the activation of pathways associated with inflammation as compared to normal-weight patients (n = 2). The K18 mouse model and human COVID-19 patient data support a link between inflammation and an obesity-accelerated COVID-19 disease phenotype. These results also indicate that obesity-accelerated severe COVID-19 caused by SARS-CoV-2 WA1 infection in the K18 mouse model would be a suitable model for dissecting the cellular and molecular mechanisms underlying pathogenesis.

Proteome Analysis Related to Unsaturated Fatty Acid Synthesis by Interfering with Bovine Adipocyte ACSL1 Gene.

Unsaturated fatty acids (UFAs) in beef play a vital role in promoting human health. Long-chain fatty acyl-CoA synthase 1 (ACSL1) is a crucial gene for UFA synthesis in bovine adipocytes. To investigate the protein expression profile during UFA synthesis, we performed a proteomic analysis of bovine adipocytes by RNA interference and non-interference with ACSL1 using label-free techniques. A total of 3558 proteins were identified in both the NC and si-treated groups, of which 1428 were differentially expressed proteins (DEPs; fold change ≥ 1.2 or ≤ 0.83 and p-value < 0.05). The enrichment analysis of the DEPs revealed signaling pathways related to UFA synthesis or metabolism, including cAMP, oxytocin, fatty acid degradation, glycerol metabolism, insulin, and the regulation of lipolysis in adipocytes (p-value < 0.05). Furthermore, based on the enrichment analysis of the DEPs, we screened 50 DEPs that potentially influence the synthesis of UFAs and constructed an interaction network. Moreover, by integrating our previously published transcriptome data, this study established a regulatory network involving differentially expressed long non-coding RNAs (DELs), highlighting 21 DEPs and 13 DELs as key genes involved in UFA synthesis. These findings present potential candidate genes for further investigation into the molecular mechanisms underlying UFA synthesis in bovines, thereby offering insights to enhance the quality of beef and contribute to consumer health in future studies.