Semaglutide 6-months therapy of type 2 diabetes mellitus restores adipose progenitors potential to develop metabolically active adipocytes.

BACKGROUND: Nowadays type 2 diabetes mellitus (T2DM) leads to population mortality growth. Today glucagon-like peptide type 1 receptor agonists (GLP-1 RA) are one of the most promising glucose-lowered drugs with anorexigenic and cardioprotective effects. The present study aims to determine the effects of GLP-1 RA semaglutide 6-month therapy on T2DM patient metabolic parameters and adipose progenitor cell health. METHODS: T2DM patients (N = 8) underwent clinical characterization and subcutaneous fat biopsy at start point and after semaglutide 6-month therapy. Adipose-derived stem cells (ADSC) were isolated by enzymatic method. Cell proliferation analysis was performed by MTT and immunocytochemistry. White and beige adipogenesis was analyzed by BODIPY493/503 staining and confocal microscopy. Adipocyte's metabolic properties were estimated by 3H- and 14C-based metabolic assays. Thermogenesis analysis was performed by ERthermAC staining and confocal microscopy. Protein markers were assessed by Western blotting. RESULTS: Semaglutide 6-month therapy demonstrated significant anorexigenic and glucose-lowering effects. However, insulin sensitivity (HOMA-IR and M-index) was unchanged after therapy. Semaglutide 6-month therapy increased ADSC proliferation and white and beige adipogenesis. Moreover, lipid droplets fragmentation was observed in beige adipocytes. Both white and beige adipocytes after semaglutide therapy demonstrated 2-3 fold growth of glucose uptake without changes in insulin sensitivity. Newly formed white adipocytes demonstrated glucose utilization for active ATP synthesis, whereas beige adipocytes for canonical thermogenesis. CONCLUSIONS: Our study has revealed that semaglutide 6-month therapy has not only systemic anorexigenic effects, but can markedly improve adipose tissue health. We have demonstrated critical restoration of ADSC renewal functions, which potentially can be involved in semaglutide based weight loss.

Dihydrolipoyl dehydrogenase promotes white adipocytes browning by activating the RAS/ERK pathway and undergoing crotonylation modification.

Acetylation modification has a wide range of functional roles in almost all physiological processes, such as transcription and energy metabolism. Crotonylation modification is mainly involved in RNA processing, nucleic acid metabolism, chromosome assembly and gene expression, and it's found that there is a competitive relationship between crotonylation modification and acetylation modification. Previous study found that dihydrolipoyl dehydrogenase (DLD) was highly expressed in brown adipose tissue (BAT) of white adipose tissue browning model mice, suggesting that DLD is closely related to white fat browning. This study was performed by quantitative real-time PCR (qPCR), Western blotting (WB), Enzyme-linked immunosorbent assay (ELISA), Immunofluorescence staining, JC-1 staining, Mito-Tracker Red CMXRos staining, Oil red O staining, Bodipy staining, HE staining, and Blood lipid quadruple test. The assay revealed that DLD promotes browning of white adipose tissue in mice. Cellularly, DLD was found to promote white adipocytes browning by activating mitochondrial function through the RAS/ERK pathway. Further studies revealed that the crotonylation modification and acetylation modification of DLD had mutual inhibitory effects. Meanwhile, DLD crotonylation promoted white adipocytes browning, while DLD acetylation did the opposite. Finally, protein interaction analysis and Co-immunoprecipitation (Co-IP) assays identified Sirtuin3 (SIRT3) as a decrotonylation and deacetylation modification enzyme of regulates DLD. In conclusion, DLD promotes browning of white adipocytes by activating mitochondrial function through crotonylation modification and the RAS/ERK pathway, providing a theoretical basis for the control and treatment of obesity, which is of great significance for the treatment of obesity and obesity-related diseases in the future.

Sex and age affect depot expression of Ca2+ channels in rat white fat adipocytes.

White adipose tissue (WAT) requires extracellular Ca2+ influx for lipolysis, differentiation, and expansion. This partly occurs via plasma membrane Ca2+ voltage-dependent channels (CaVs). However, WFA exists in different depots whose function varies with age, sex, and location. To explore whether their CaV expression profiles also differ we used RNAseq and qPCR on gonadal, mesenteric, retroperitoneal, and inguinal subcutaneous fat depots from rats of different ages and sex. CaV expression was found dependent on age, sex, and WFA location. In the gonadal depots of both sexes a significantly lower expression of CaV1.2 and CaV1.3 was seen for adults compared to pre-pubescent juveniles. A lower level of expression was also seen for CaV3.1 in adult male but not female gonadal WFA, the latter of whose expression remained unchanged with age. Relatively little expression of CaV3.2 and 3.2 was observed. In post-pubescent inguinal subcutaneous fat, where the third and fourth mammary glands are located, CaV3.1 was decreased in males but increased in females - thus suggesting that this channel is associated with mammogenesis; however, no difference in intracellular Ca2+ levels or adipocyte size were noted. For all adult depots, CaV3.1 expression was larger in females than males - a difference not seen in pre-pubescent rats. These observations are consistent with the changes of CaV3.1 expression seen in 3T3-L1 cell differentiation and the ability of selective CaV3.1 antagonists to inhibit adipogensis. Our results show that changes in CaV expression patterns occur in fat depots related to sexual dimorphism: reproductive tracts and mammogenesis.

IL-4 activates the futile triacylglyceride cycle for glucose utilization in white adipocytes.

The development of cardiometabolic complications during obesity is strongly associated with chronic latent inflammation in hypertrophied adipose tissue (AT). IL-4 is an anti-inflammatory cytokine, playing a protective role against insulin resistance, glucose intolerance and weight gain. The positive effects of IL-4 are associated not only with the activation of anti-inflammatory immune cells in AT, but also with the modulation of adipocyte metabolism. IL-4 is known to activate lipolysis and glucose uptake in adipocytes, but the precise regulatory mechanisms and physiological significance of these processes remain unclear. In this study, we detail IL-4 effects on glucose and triacylglycerides (TAGs) metabolism and propose mechanisms of IL-4 metabolic action in adipocytes. We have shown that IL-4 activates glucose oxidation, lipid droplet (LD) fragmentation, lipolysis and thermogenesis in mature 3T3-L1 adipocytes. We found that lipolysis was not accompanied by fatty acids (FAs) release from adipocytes, suggesting FA re-esterification. Moreover, glucose oxidation and thermogenesis stimulation depended on adipocyte triglyceride lipase (ATGL) activity, but not the uncoupling protein (UCP1) expression. Based on these data, IL-4 may activate the futile TAG-FA cycle in adipocytes, which enhances the oxidative activity of cells and heat production. Thus, the positive effect of IL-4 on systemic metabolism can be the result of the activation of non-canonical thermogenic mechanism in AT, increasing TAG turnover and utilization of excessive glucose.

Role of hydrogen sulfide in the regulation of lipid metabolism: Implications on cardiovascular health.

The World Health Organization (WHO) defines obesity as an urgency for health and a social emergency. Today around 39 % of people is overweight, of these over 13 % is obese. It is well-consolidated that the adipose cells are deputy to lipid storage under caloric excess; however, despite the classical idea that adipose tissue has exclusively a passive function, now it is known to be deeply involved in the regulation of systemic metabolism in physiological as well as under obesogenic conditions, with consequences on cardiovascular health. Beside two traditional types of adipose cells (white and brown), recently the beige one has been highlighted as the consequence of the healthy remodeling of white adipocytes, confirming their metabolic adaptability. In this direction, pharmacological, nutraceutical and nutrient-based approaches are addressed to positively influence inflammation and metabolism, thus contributing to reduce the obese-associated cardiovascular risk. In this scenario, hydrogen sulfide emerges as a new mediator that may regulate crucial targets involved in the regulation of metabolism. The current evidence demonstrates that hydrogen sulfide may induce peroxisome proliferator activated receptor γ (PPARγ), a crucial mediator of adipogenesis, inhibit the phosphorylation of perlipin-1 (plin-1), a protein implicated in the lipolysis, and finally promote browning process, through the release of irisin from skeletal muscle. The results summarized in this review suggest an important role of hydrogen sulfide in the regulation of metabolism and in the prevention/treatment of obese-associated cardiovascular diseases and propose new insight on the putative mechanisms underlying the release of hydrogen sulfide or its biosynthesis, delineating a further exciting field of application.

Obesity causes mitochondrial fragmentation and dysfunction in white adipocytes due to RalA activation.

Mitochondrial dysfunction is a characteristic trait of human and rodent obesity, insulin resistance and fatty liver disease. Here we show that high-fat diet (HFD) feeding causes mitochondrial fragmentation in inguinal white adipocytes from male mice, leading to reduced oxidative capacity by a process dependent on the small GTPase RalA. RalA expression and activity are increased in white adipocytes after HFD. Targeted deletion of RalA in white adipocytes prevents fragmentation of mitochondria and diminishes HFD-induced weight gain by increasing fatty acid oxidation. Mechanistically, RalA increases fission in adipocytes by reversing the inhibitory Ser637 phosphorylation of the fission protein Drp1, leading to more mitochondrial fragmentation. Adipose tissue expression of the human homolog of Drp1, DNM1L, is positively correlated with obesity and insulin resistance. Thus, chronic activation of RalA plays a key role in repressing energy expenditure in obese adipose tissue by shifting the balance of mitochondrial dynamics toward excessive fission, contributing to weight gain and metabolic dysfunction.

NG ,NG -Dimethylarginine Dimethylaminohydrolase 1 Expression Is Dispensable for Cold- or Diet-Induced Thermogenesis.

The strategy to activate thermogenic adipocytes has therapeutic potential to overcome obesity as they dissipate surplus energy as heat through various mechanisms. NG,NG-dimethylarginine dimethylaminohydrolases (DDAHs) are enzymes involved in the nitric oxide-protein kinase G signaling axis which increases thermogenic gene expression. However, the role of DDAHs in thermogenic adipocytes has not been elucidated. The adipocyte-specific Ddah1 knockout mice are generated by crossing Ddah1fl/fl mice with adiponectin Cre recombinase mice. Adipocyte-specific DDAH1 overexpressing mice are generated using adeno-associated virus-double-floxed inverse open reading frame (AAV-DIO) system. These mice are analyzed under basal, cold exposure, or high-fat diet (HFD) conditions. Primary inguinal white adipose tissue cells from adipocyte-specific Ddah1 knockout mice expressed comparable amounts of Ucp1 mRNA. Adipocyte-specific DDAH1 overexpressing mice do not exhibit enhanced activation of thermogenic adipocytes. In addition, when these mice are exposed to cold environment or fed an HFD, their body temperature/weight and thermogenesis-related gene and protein expressions are unchanged. These findings indicate that DDAH1 does not play a role in either cold- or diet-induced thermogenesis. Therefore, adipocyte targeting DDAH1 gene therapy for the treatment of obesity is unlikely to be effective.

Direct effects of adipocyte lipolysis on AMPK through intracellular long-chain acyl-CoA signaling.

Long-chain acyl-CoAs (LC-acyl-CoAs) are important intermediary metabolites and are also thought to function as intracellular signaling molecules; however, the direct effects of LC-acyl-CoAs have been difficult to determine in real-time and dissociate from Protein Kinase A (PKA) signaling. Here, we examined the direct role of lipolysis in generating intracellular LC-acyl-CoAs and activating AMPK in white adipocytes by pharmacological activation of ABHD5 (also known as CGI-58), a lipase co-activator. Activation of lipolysis in 3T3-L1 adipocytes independent of PKA with synthetic ABHD5 ligands, resulted in greater activation of AMPK compared to receptor-mediated activation with isoproterenol, a ß-adrenergic receptor agonist. Importantly, the effect of pharmacological activation of ABHD5 on AMPK activation was blocked by inhibiting ATGL, the rate-limiting enzyme for triacylglycerol hydrolysis. Utilizing a novel FRET sensor to detect intracellular LC-acyl-CoAs, we demonstrate that stimulation of lipolysis in 3T3-L1 adipocytes increased the production of LC-acyl-CoAs, an effect which was blocked by inhibition of ATGL. Moreover, ATGL inhibition blocked AMPKß1 S108 phosphorylation, a site required for allosteric regulation. Increasing intracellular LC-acyl-CoAs by removal of BSA in the media and pharmacological inhibition of DGAT1 and 2 resulted in greater activation of AMPK. Finally, inhibiting LC-acyl-CoA generation reduced activation of AMPK; however, did not lower energy charge. Overall, results demonstrate that lipolysis in white adipocytes directly results in allosteric activation of AMPK through the generation of LC-acyl-CoAs.

Adiponectin stimulates Sca1+CD34--adipocyte precursor cells associated with hyperplastic expansion and beiging of brown and white adipose tissue.

BACKGROUND: The adipocyte hormone adiponectin improves insulin sensitivity and there is an inverse correlation between adiponectin levels and type-2 diabetes risk. Previous research shows that adiponectin remodels the adipose tissue into a more efficient metabolic sink. For instance, mice that overexpress adiponectin show increased capacity for hyperplastic adipose tissue expansion as evident from smaller and metabolically more active white adipocytes. In contrast, the brown adipose tissue (BAT) of these mice looks "whiter" possibly indicating reduced metabolic activity. Here, we aimed to further establish the effect of adiponectin on adipose tissue expansion and adipocyte mitochondrial function as well as to unravel mechanistic aspects in this area. METHODS: Brown and white adipose tissues from adiponectin overexpressing (APN tg) mice and littermate wildtype controls, housed at room and cold temperature, were studied by histological, gene/protein expression and flow cytometry analyses. Metabolic and mitochondrial functions were studied by radiotracers and Seahorse-based technology. In addition, mitochondrial function was assessed in cultured adiponectin deficient adipocytes from APN knockout and heterozygote mice. RESULTS: APN tg BAT displayed increased proliferation prenatally leading to enlarged BAT. Postnatally, APN tg BAT turned whiter than control BAT, confirming previous reports. Furthermore, elevated adiponectin augmented the sympathetic innervation/activation within adipose tissue. APN tg BAT displayed reduced metabolic activity and reduced mitochondrial oxygen consumption rate (OCR). In contrast, APN tg inguinal white adipose tissue (IWAT) displayed enhanced metabolic activity. These metabolic differences between genotypes were apparent also in cultured adipocytes differentiated from BAT and IWAT stroma vascular fraction, and the OCR was reduced in both brown and white APN heterozygote adipocytes. In both APN tg BAT and IWAT, the mesenchymal stem cell-related genes were upregulated along with an increased abundance of Lineage-Sca1+CD34- "beige-like" adipocyte precursor cells. In vitro, the adiponectin receptor agonist Adiporon increased the expression of the proliferation marker Pcna and decreased the expression of Cd34 in Sca1+ mesenchymal stem cells. CONCLUSIONS: We propose that the seemingly opposite effect of adiponectin on BAT and IWAT is mediated by a common mechanism; while reduced adiponectin levels are linked to lower adipocyte OCR, elevated adiponectin levels stimulate expansion of adipocyte precursor cells that produce adipocytes with intrinsically higher metabolic rate than classical white but lower metabolic rate than classical brown adipocytes. Moreover, adiponectin can modify the adipocytes' metabolic activity directly and by enhancing the sympathetic innervation within a fat depot.

Modulation of angiogenic switch in reprogramming browning and lipid metabolism in white adipocytes.

Thermogenic activation via trans-and de novo browning of white adipocytes is a promising strategy to accelerate lipid metabolism for regulating obesity-related disorders. In this study, we investigated the intricate interplay between angiogenic regulation and browning in white adipocytes using the bioactive compound, resveratrol (Rsv). Rsv has previously been documented for its regulatory influence on the trans and de novo browning of white adipocytes. Our findings revealed that concurrent activation of angiogenesis is prerequisite for inducing browning within the microenvironment of white adipocytes when exposed to browning activators. Additionally, we observed a significant browning effect on white adipocytes when the local adipose tissue environment was prompted to undergo angiogenesis, notably facilitated by a proangiogenic molecule known as Vascular endothelial growth factor (VEGF). Intriguingly, this effect was reversed when angiogenesis was inhibited by treatment with the antiangiogenic agent thalidomide. Furthermore, the study revealed the role of VEGF in paracrine activation of white adipocytes resulting in the induction of browning in both 3T3-L1 cell lines and primary mouse white adipocytes. The cross-talk between angiogenesis and browning was found to be initiated via the transcriptional activation of Estrogen receptor α (ERα) triggering the VEGF/VEGFR2 signaling pathway leading to browning and a reconfiguration of lipid metabolism within adipocytes. In conclusion, this study sheds light on the intricate cross-talk between angiogenesis and browning of white adipocytes. Notably, the findings underscore the reciprocal relationship between these processes, wherein inhibition of one process exerts discernible effects on the other.

miR-889-3p Facilitates the Browning Process of White Adipocyte Precursors by Targeting the SON Gene.

It is well-established that beige/brown adipose tissue can dissipate stored energy through thermogenesis; hence, the browning of white adipocytes (WAT) has garnered significant interest in contemporary research. Our preceding investigations have identified a marked downregulation of miR-889-3p concurrent with the natural maturation of brown adipose tissue. However, the specific role and underlying molecular mechanisms of miR-889-3p in the browning process of white adipose tissue warrant further elucidation. In this research, we initially delved into the potential role of miR-889-3p in preadipocyte growth via flow cytometry and CCK-8 assay, revealing that miR-889-3p can stimulate preadipocyte growth. To validate the potential contribution of miR-889-3p in the browning process of white adipose tissue, we established an in vitro rabbit white adipocyte browning induction, which exhibited a significant upregulation of miR-889-3p during the browning process. RT-qPCR and Western blot analysis indicated that miR-889-3p overexpression significantly amplified the mRNA levels of UCP1, PRDM16, and CIDEA, as well as UCP1 protein levels. Furthermore, miR-889-3p overexpression fostered intracellular triglyceride accumulation. Conversely, the downregulation of miR-889-3p hindered the browning of rabbit preadipocytes. Subsequently, based on target gene prediction and luciferase reporter gene determination, we demonstrated that miR-889-3p directly targets the 3'-UTR region of SON. Lastly, we observed that inhibiting SON could facilitate the browning of rabbit preadipocytes. In conclusion, our findings suggest that miR-889-3p facilitates the browning process of white adipocyte precursors by specifically targeting the SON gene.

NRF2 is essential for adaptative browning of white adipocytes.

White adipose tissue browning, defined by accelerated mitochondrial metabolism and biogenesis, is considered a promising mean to treat or prevent obesity-associated metabolic disturbances. We hypothesize that redox stress acutely leads to increased production of reactive oxygen species (ROS), which activate electrophile sensor nuclear factor erythroid 2-Related Factor 2 (NRF2) that over time results in an adaptive adipose tissue browning process. To test this, we have exploited adipocyte-specific NRF2 knockout mice and cultured adipocytes and analyzed time- and dose-dependent effect of NAC and lactate treatment on antioxidant expression and browning-like processes. We found that short-term antioxidant treatment with N-acetylcysteine (NAC) induced reductive stress as evident from increased intracellular NADH levels, increased ROS-production, reduced oxygen consumption rate (OCR), and increased NRF2 levels in white adipocytes. In contrast, and in line with our hypothesis, longer-term NAC treatment led to a NRF2-dependent browning response. Lactate treatment elicited similar effects as NAC, and mechanistically, these NRF2-dependent adipocyte browning responses in vitro were mediated by increased heme oxygenase-1 (HMOX1) activity. Moreover, this NRF2-HMOX1 axis was also important for ß3-adrenergic receptor activation-induced adipose tissue browning in vivo. In conclusion, our findings show that administration of exogenous antioxidants can affect biological function not solely through ROS neutralization, but also through reductive stress. We also demonstrate that NRF2 is essential for white adipose tissue browning processes.

IL-27 increases energy storage in white adipocytes by enhancing glucose uptake and fatty acid esterification.

The cytokine interleukin (IL)-27 has been reported to induce thermogenesis in white adipocytes. However, it remains unknown whether IL-27-mediated adipocyte energy dissipation is paralleled by an elevated energy supply from lipids and/or carbohydrates. We hypothesized that IL-27 increases lipolysis and glucose uptake in white adipocytes, thereby providing substrates for thermogenesis. Unexpectedly, we found that treatment of 3T3-L1 adipocytes with IL-27 reduced intra- and extracellular free fatty acid (FFA) concentrations and that phosphorylation of hormone-sensitive lipase (HSL) was not affected by IL-27. These results were confirmed in subcutaneous white adipocytes. Further, application of IL-27 to 3T3-L1 adipocytes increased intracellular triglyceride (TG) content but not mitochondrial ATP production nor expression of enzymes involved in beta-oxidation indicating that elevated esterification rather than oxidation causes FFA disappearance. In addition, IL-27 significantly increased GLUT1 protein levels, basal glucose uptake as well as glycolytic ATP production, suggesting that increased glycolytic flux due to IL-27 provides the glycerol backbone for TG synthesis. In conclusion, our findings suggest IL-27 increases glucose uptake and TG deposition in white adipocytes.

Intermuscular adipose tissue in obesity and related disorders: cellular origins, biological characteristics and regulatory mechanisms.

Intermuscular adipose tissue (IMAT) is a unique adipose depot interspersed between muscle fibers (myofibers) or muscle groups. Numerous studies have shown that IMAT is strongly associated with insulin resistance and muscular dysfunction in people with metabolic disease, such as obesity and type 2 diabetes. Moreover, IMAT aggravates obesity-related muscle metabolism disorders via secretory factors. Interestingly, researchers have discovered that intermuscular brown adipocytes in rodent models provide new hope for obesity treatment by acting on energy dissipation, which inspired researchers to explore the underlying regulation of IMAT formation. However, the molecular and cellular properties and regulatory processes of IMAT remain debated. Previous studies have suggested that muscle-derived stem/progenitor cells and other adipose tissue progenitors contribute to the development of IMAT. Adipocytes within IMAT exhibit features that are similar to either white adipocytes or uncoupling protein 1 (UCP1)-positive brown adipocytes. Additionally, given the heterogeneity of skeletal muscle, which comprises myofibers, satellite cells, and resident mesenchymal progenitors, it is plausible that interplay between these cellular components actively participate in the regulation of intermuscular adipogenesis. In this context, we review recent studies associated with IMAT to offer insights into the cellular origins, biological properties, and regulatory mechanisms of IMAT. Our aim is to provide novel ideas for the therapeutic strategy of IMAT and the development of new drugs targeting IMAT-related metabolic diseases.

Adipocyte Subpopulations Mediate Growth Hormone-induced Lipolysis and Glucose Tolerance in Male Mice.

In adipose tissue, growth hormone (GH) stimulates lipolysis, leading to an increase in plasma free fatty acid levels and a reduction in insulin sensitivity. In our previous studies, we have found that GH increases lipolysis by reducing peroxisome proliferator-activated receptor γ (PPARγ) transcription activity, leading to a reduction of tat-specific protein 27 (FSP27, also known as CIDEC) expression. In previous studies, our laboratory uncovered 3 developmentally distinct subpopulations of white adipocytes. In this manuscript, we show that one of the subpopulations, termed type 2 adipocytes, has increased GH-induced signaling and lipolysis compared to other adipocyte subtypes. To assess the physiological role of GH-mediated lipolysis mediated by this adipocyte subpopulation, we specifically expressed human FSP27 (hFSP27) transgene in type 2 adipocytes (type2Ad-hFSP27tg mice). Systemically, male type2Ad-hFSP27tg mice displayed reduced serum glycerol release and nonesterified fatty acids levels after acute GH treatment, and improvement in acute, but not chronic, GH-induced glucose intolerance. Furthermore, we demonstrate that type2Ad-hFSP27tg mice displayed improved hepatic insulin signaling. Taken together, these results indicate that this adipocyte subpopulation is a critical regulator of the GH-mediated lipolytic and metabolic response. Thus, further investigation of adipocyte subpopulations may provide novel treatment strategies to regulate GH-induced glucose intolerance in patients with growth and metabolic disorders.

Transcriptomics profiling reveal the heterogeneity of white and brown adipocyte.

The marker genes associated with white adipocytes and brown adipocytes have been previously identified; however, these markers have not been updated in several years, and the differentiation process of preadipocytes remains relatively fixed. Consequently, there has been a lack of exploration into alternative differentiation schemes. In this particular study, we present a transcriptional signature specific to brown adipocytes and white adipocytes. Notably, our findings reveal that ZNF497, ZIC1, ZFY, UTY, USP9Y, TXLNGY, TTTY14, TNNT3, TNNT2, TNNT1, TNNI1, TNNC1, TDRD15, SOX11, SLN, SFRP2, PRKY, PAX3KLHL40, PAX3, INKA2-AS1, SOX11, and TDRD15 exhibit high expression levels in brown adipocytes. XIST, HOXA10, PCAT19, HOXA7, PLSCR3, and AVPR1A exhibited high expression levels in white adipocytes, suggesting their potential as novel marker genes for the transition from white to brown adipocytes. Furthermore, our analysis revealed the coordinated activation of several pathways, including the PPAR signaling pathway, focal adhesion, retrograde endocannabinoid signaling, oxidative phosphorylation, PI3K-Akt signaling pathway, and thermogenesis pathways, in brown adipocytes. Moreover, in contrast to prevailing culture techniques, we conducted a comparative analysis of the differentiation protocols for white preadipocytes and brown preadipocytes, revealing that the differentiation outcome remained unaffected by the diverse culture schemes employed. However, the expression levels of certain marker genes in both adipocyte types were found to be altered. This investigation not only identified potential novel marker genes for adipocytes but also examined the impact of different differentiation methods on preadipocyte maturation. Consequently, these findings offer significant insights for further research on the differentiation processes of diverse adipocyte subtypes.

The Emerging Importance of Mitochondria in White Adipocytes: Neither Last nor Least.

The growing recognition of mitochondria's crucial role in the regulation of white adipose tissue remodeling and energy balance underscores its significance. The marked metabolic diversity of mitochondria provides the molecular and cellular foundation for enabling adipose tissue plasticity in response to various metabolic cues. Effective control of mitochondrial function at the cellular level, not only in thermogenic brown and beige adipocytes but also in energy-storing white adipocytes, exerts a profound influence on adipose homeostasis. Furthermore, mitochondria play a pivotal role in intercellular communication within adipose tissue via production of metabolites with signaling properties. A more comprehensive understanding of mitochondrial regulation within white adipocytes will empower the development of targeted and efficacious strategies to enhance adipose function, leading to advancements in overall metabolic health.

Acacetin alleviates energy metabolism disorder through promoting white fat browning mediated by AC-cAMP pathway

Acacetin (ACA), a flavone isolated from Chinese traditional medical herbs, has numerous pharmacological activities. However, little is known about the roles in white fat browning and energy metabolism. In the present study, we investigated whether and how ACA would improve energy metabolism in vivo and in vitro. ACA (20 mg/kg) was intraperitoneally injected to the mice with obesity induced by HFD for 14 consecutive days (in vivo); differentiated 3T3-L1 adipocytes were treated with ACA (20 µmol/L and 40 µmol/L) for 24 h (in vitro). The metabolic profile, lipid accumulation, fat-browning and mitochondrial contents, and so on were respectively detected. The results in vivo showed that ACA significantly reduced the body weight and visceral adipose tissue weight, alleviated the energy metabolism disorder, and enhanced the browning-related protein expressions in adipose tissue of rats. Besides, the data in vitro revealed that ACA significantly reduced the lipid accumulation, induced the expressions of the browning-related proteins and cAMP-dependent protein kinase A (PKA), and increased the mitochondrium contents, especially enhanced the energy metabolism of adipocytes; however, treatment with beta-adrenergic receptor blocker (propranolol, Pro) or adenyl cyclase (AC) inhibitor (SQ22536, SQ) abrogated the ACA-mediated effects. The data demonstrate that ACA alleviates the energy metabolism disorder through the pro-browning effects mediated by the AC-cAMP pathway. The findings would provide the experimental foundation for ACA to prevent and treat obesity and related metabolism disorders. (AU)

Inhibition of AXL receptor tyrosine kinase enhances brown adipose tissue functionality in mice.

The current obesity epidemic and high prevalence of metabolic diseases necessitate efficacious and safe treatments. Brown adipose tissue in this context is a promising target with the potential to increase energy expenditure, however no pharmacological treatments activating brown adipose tissue are currently available. Here, we identify AXL receptor tyrosine kinase as a regulator of adipose function. Pharmacological and genetic inhibition of AXL enhance thermogenic capacity of brown and white adipocytes, in vitro and in vivo. Mechanistically, these effects are mediated through inhibition of PI3K/AKT/PDE signaling pathway, resulting in induction of nuclear FOXO1 localization and increased intracellular cAMP levels via PDE3/4 inhibition and subsequent stimulation of the PKA-ATF2 pathway. In line with this, both constitutive Axl deletion as well as inducible adipocyte-specific Axl deletion protect animals from diet-induced obesity concomitant with increases in energy expenditure. Based on these data, we propose AXL receptor as a target for the treatment of obesity.

Acteoside improves adipocyte browning by CDK6-mediated mTORC1-TFEB pathway.

Adipocyte browning increases energy expenditure by thermogenesis, which has been considered a potential strategy against obesity and its related metabolic diseases. Phytochemicals derived from natural products with the ability to improve adipocyte thermogenesis have aroused extensive attention. Acteoside (Act), a phenylethanoid glycoside, exists in various medicinal or edible plants and has been shown to regulate metabolic disorders. Here, the browning effect of Act was evaluated by stimulating beige cell differentiation from the stromal vascular fraction (SVF) in the inguinal white adipose tissue (iWAT) and 3T3-L1 preadipocytes, and by converting the iWAT-SVF derived mature white adipocytes. Act improves adipocyte browning by differentiation of the stem/progenitors into beige cells and by direct conversion of mature white adipocytes into beige cells. Mechanistically, Act inhibited CDK6 and mTOR, and consequently relieved phosphorylation of the transcription factor EB (TFEB) and increased its nuclear retention, leading to induction of PGC-1α, a driver of mitochondrial biogenesis, and UCP1-dependent browning. These data thus unveil a CDK6-mTORC1-TFEB pathway that regulates Act-induced adipocyte browning.