Adipocyte Differentiation Markers Research Articles

Pleomorphic liposarcoma of bone: a rare primary malignant bone tumour

BackgroundLiposarcoma is an extremely rare primary bone sarcoma.Case presentationWe report a case of primary pleomorphic liposarcoma that arose in an 18 year old male in the metaphysis of the left tibia. Plain radiographs showed a partly sclerotic lesion and MR imaging a heterogeneous tumour predominantly isointense on T1- and high-signal on T2-weighted sequences with focal areas of increased T1 signal that suppressed with fat saturation. PET/CT showed marked FDG uptake (SUV = 17.1) in the primary tumour as well as a metastasis in the right distal femur and multiple small pulmonary metastases. Histologically, the tumour was a pleomorphic liposarcoma containing large tumour cells with vacuolated cytoplasm and hyperchromatic pleomorphic nuclei as well as numerous lipoblasts and scattered brown fat-like cells. Tumour cells strongly expressed FABP4/aP2, a marker of adipocyte differentiation, and UCP1, a marker of brown fat, but not S100. The case was treated with neoadjuvant MAP chemotherapy, resulting in extensive (> 95%) necrosis in the primary tumour and almost complete resolution of the femoral and pulmonary metastases.ConclusionsPleomorphic liposarcoma can present as a sclerotic primary malignant bone tumour; markers of adipose differentiation are useful in histological diagnosis and neoadjuvant MAP chemotherapy results in significant tumor necrosis.

Effect of prehypertensive losartan therapy on AT1R and ATRAP methylation of adipose tissue in the later life of high‑fat‑fed spontaneously hypertensive rats.

Hypertension is frequently associated with metabolic disorders. The present study was designed to investigate the long-term effect of prehypertensive losartan therapy on metabolic disorders in high-fat-fed spontaneously hypertensive rats (SHRs), and to examine the role of epigenetic regulation of angiotensin II type 1 receptor (AT1R) and AT1 receptor-associated protein (ATRAP) expression in adipose tissue. A total of 32 4-week-old male SHRs were divided into four groups (n=8 rats/group): Standard chow; standard chow + losartan; high-fat diet; and high-fat diet + losartan. At 10 weeks of age, treatment with losartan was discontinued. Rats were followed up until 26 weeks of age. Obesity, dyslipidemia, hyperglycemia, abnormal adipokine secretion, larger adipocytes and decreased expression of markers of adipocyte differentiation were present in high-fat-fed SHRs, and were attenuated in losartan-treated rats. The increased expression and promoter hypomethylation of AT1R subtype a (AT1aR) in the adipose tissue of high-fat-fed SHRs were reversed by treatment with losartan. No difference was observed in the expression and promoter methylation of AT1R subtype b (AT1bR) among the four groups. Decreased expression and promoter hypermethylation of ATRAP were demonstrated in the adipose tissue of high-fat-fed SHRs. However, losartan made no difference to the expression and promoter methylation of ATRAP. Prehypertensive losartan therapy may relieve metabolic disorders in the later life of high-fat-fed SHRs. Differential epigenetic regulation of AT1aR and ATRAP expression through DNA methylation in adipose tissue may be involved in the long-term beneficial effect.

Serotonin prevents differentiation into brown adipocytes and induces transdifferentiation into white adipocytes.

Serotonin is synthesized by many cells in the periphery to affect vasoconstriction, intestinal motility, and glucose and lipid metabolism. It has recently been shown that serotonin leads to fat accumulation in white adipose tissue (WAT). However, the direct effect of serotonin on brown adipose tissue differentiation and metabolism is limited. Therefore, our aim was to investigate the effect of serotonin on brown adipocyte metabolism and differentiation. Non-differentiated HIB1B cells and differentiated HIB1B brown adipocytes were treated with serotonin and their metabolism and differentiation examined. Differentiated HIB1B brown adipocytes treated with serotonin had reduced levels of the thermogenic markers uncoupling protein 1 (UCP1) and fibroblast growth factor 21 (FGF21) and increased levels of UCP2. In parallel, serotonin led to 3-6-fold reduction in the gene expression of brown adipocyte differentiation markers, that is, Prdm16 (positive regulatory domain 16), Bmp7 (bone morphogenic protein 7) and Pparγ (peroxisome-proliferator-activated receptor γ). Serotonin treatment reduced catabolism and mitochondrial activity shifting metabolism towards fatty acid synthesis rather than oxidation. Strikingly, non-differentiated HIB1B preadipocytes incubated with serotonin failed to differentiate into brown adipocytes. Moreover, although BMP6-treated myoblasts can readily differentiate into brown adipocytes, serotonin interfered with this process. Serotonin leads to whitening of brown adipocytes, shifting their metabolism to fat accumulation rather than oxidation. In addition, serotonin interferes with the differentiation process into brown adipocytes.

Brown adipocytes postnatally arise through both differentiation from progenitors and conversion from white adipocytes in Syrian hamster.

To investigate the postnatal development of brown adipose tissue (BAT) in Syrian hamsters, we histologically examined interscapular fat tissue from 5-16-day-old pups, focusing on how brown adipocytes arise. Interscapular fat of 5-day-old hamsters mainly consisted of white adipocytes containing large unilocular lipid droplets, as observed in typical white adipose tissue (WAT). On day 7, clusters of small, proliferative nonadipocytes with a strong immunoreactivity for Ki67 appeared near the edge of the interscapular fat tissue. The area of the Ki67-positive regions expanded to ~50% of the total tissue area by day 10. The interscapular fat showed the typical BAT feature by day 16. A brown adipocyte-specific marker, uncoupling protein-1, was clearly detected on day 10 and thereafter, while not detected on day 7. During conversion of interscapular fat from WAT to BAT, unilocular adipocytes completely and rapidly disappeared without obvious apoptosis. Dual immunofluorescence staining for Ki67 and monocarboxylate transporter 1 (MCT1), another selective marker for brown adipocytes, revealed that most of the proliferating cells were of the brown adipocyte lineage. Electron microscopic examination showed that some of the white adipocytes contained small lipid droplets in addition to the large droplet and expressed MCT1 as do progenitor and mature brown adipocytes, implying a direct conversion from white to brown adipocytes. These results suggest that BAT of Syrian hamsters develops postnatally through two different pathways: the proliferation and differentiation of brown adipocyte progenitors and the conversion of unilocular adipocytes to multilocular brown adipocytes. NEW & NOTEWORTHY Brown and white adipose tissues (BAT and WAT, respectively) are quite different in morphological features and function; however, the boundary between these tissues is obscure. In this study, we histologically evaluated the process of BAT development in Syrian hamsters, which shows postnatal conversion of WAT to BAT. Our results suggest that brown adipocytes arise through two different pathways: the proliferation and differentiation of brown adipocyte progenitors and the conversion from white adipocytes.

Characterization of the visfatin gene and its expression pattern and effect on 3T3-L1 adipocyte differentiation in chickens

Visfatin is a newly identified adipocytokine that plays an important role in the determination of fat traits. In this study, we investigated the characterization of visfatin and the relationship between gene expression and chicken development to provide a theoretical basis for studying visfatin functions. The main results are summarized as follows: The 1482-bp full coding sequence of the visfatin gene of silky fowl was obtained and found to encode 493 amino acids. This gene contains 26 phosphorylation sites and a conserved domain of the NAPRTase family but no signal peptide sequence. It exhibits six functional motifs, including an amidation site. In chickens, visfatin is a highly conserved protein. The highest expression of visfatin was found in breast muscle and the lowest in bone marrow. There was no difference in expression between visceral fat and subcutaneous fat. However, the expression of visfatin in the bone marrow, liver, kidneys, and subcutaneous and visceral fat of broiler chickens was significantly higher than that in silky fowl (P<0.05). Visfatin mRNA levels in the bone marrow decreased with development (P<0.05) but increased in the liver and leg muscle. Visfatin gene expression in the liver, heart and bone marrow did not differ in silky fowl according to sex. A visfatin fusion protein caused a significant increase in the expression of adipocyte differentiation markers (PPARγ, aP2, C/EBPα, and FAS) compared with the control group and a decrease compared with the insulin group. Taken together, the results of the present study contribute to a better understanding of the expression and role of the visfatin gene in chickens.

Optimized &lt;i&gt;Cirsium setidens&lt;/i&gt; Nakai Fermented by &lt;i&gt;Lentinula edodes&lt;/i&gt; Attenuates Lipid Accumulation by Regulating Fatty Acid Oxidation-mediated Lipolysis in 3T3-L1 Cells and High Calorie Diet-induced Obese Zebrafish

Cirsium setidens Nakai is an edible herb. Previously we found that fermented Cirsium setidens Nakai (FCSN) has a large amount of major bioactive compound compared to Cirsium setidens Nakai. In this study, we aimed to examine the anti-obesity effect of FCSN using 3T3-L1 cells in vitro and high calorie diet-induced obese (HDIO) zebrafish model in vivo. Our results demonstrated that FCSN significantly inhibited intracellular lipid accumulation in 3T3-L1 cells. FCSN was shown to reduce the expressions of crucial adipocyte differentiation markers, including PPARγ and aP2. FCSN also decreased the production of ROS due to the up-regulated expressions of SOD1, SOD2, GPx, and catalase. Furthermore, we observed that FCSN also altered the levels of energy metabolism and β-oxidation-associated genes such as AMPK, ACC, and CPT-1. In addition, ATGL, a key lipolysis enzyme, was stimulated while the differentiation of 3T3-L1 was suppressed by FCSN. Strikingly, we found that FCSN dramatically increased both the energy metabolism and β-oxidation associated genes and subsequently prevented the increase of body fat accumulation in high calorie diet-induced obese zebrafish. Taken together, this is the first study that demonstrates that FCSN has the beneficial activity to suppress adipogenesis in 3T3-L1 cells and ameliorate an obese-associated health condition in vivo.

Haematopoietic TLR4 deletion attenuates perivascular brown adipose tissue inflammation in atherosclerotic mice

AimsTo investigate whether haematopoietic TLR4 deletion attenuates perivascular brown adipose tissue inflammation in atherosclerotic mice. Methods and ResultsExperiments were performed using irradiated LDL receptor-deficient (LDLR−/−) mice with marrow from either TLR4-deficient (TLR4−/−) or age-matched wild-type (WT) mice. After 12 weeks of being fed a high-cholesterol diet, TLR4−/−→LDLR−/− mice developed fewer atherosclerotic lesions in the aorta compared to WT→LDLR−/− mice. This effect was associated with an increase in multilocular lipid droplets and mitochondria in perivascular adipose tissue (PVAT). Immunofluorescence analysis confirmed that there was an increase in capillary density and M2 macrophage infiltration, accompanied by a decrease in tumour necrosis factor (TNF)-α expression in the localized PVAT of TLR4−/−→LDLR−/− mice. In vitro studies indicated that bone marrow-derived macrophages (BMDMs) from WT mice demonstrated an M1-like phenotype and expression of inflammatory cytokines induced by palmitate. These effects were attenuated in BMDMs isolated from TLR4−/− mice. Furthermore, brown adipocytes incubated with conditioned medium (CM) derived from palmitate-treated BMDMs, exhibited larger and more unilocular lipid droplets, and reduced expression of brown adipocyte-specific markers and perilipin-1 compared to those observed in brown adipocytes exposed to CM from palmitate-treated BMDMs of TLR4−/− mice. This decreased potency was primarily due to TNF-α, as demonstrated by the capacity of the TNF-α neutralizing antibody to reverse these effects. ConclusionsThese results suggest that haematopoietic-specific deletion of TLR4 promotes PVAT homeostasis, which is involved in reducing macrophage-induced TNF-α secretion and increasing mitochondrial biogenesis in brown adipocytes.

Characterization Of Expression Levels Of Genes Involved In Adipogenesis And Inflammation In Congenic Mice Carrying Obesity and Hyperlipidemia QTL on Chromosome 1

Obesity is an international health crisis and is considered the leading cause of preventable death. The development of obesity is likely due to interactions of susceptibility genes with obesity‐promoting environments such as high fat diets (HFD). These interactions lead to an expansion of adipose tissue, through an increase in adipocyte size and/or number. Adipose tissue inflammation is a well‐recognized characteristic of obesity. Previously, we identified quantitative trait loci (QTL) linked to obesity and hyperlipidemia on mouse chromosome (Chr) 1 in an intercross of obese and diabetic TALLYHO/Jng (TH) mice with C57BL/6 (B6) mice. Subsequently, we established a congenic mouse strain carrying the Chr1 QTL, 128 Mb in size, derived from TH on a B6 background (B6.TH‐Chr1–128Mb). The congenic mice were found to be more susceptible to diet‐induced obesity and hyperlipidemia than their B6 counterparts. The purpose of this study was to characterize the congenic mice by examining mRNA and protein levels of adipocyte differentiation and inflammation markers in white adipose tissue and liver. B6.TH‐Chr1–128Mb congenic and B6 mice were weaned onto chow and HFD and maintained until 16 weeks of age. Total RNA was isolated from adipose tissue and reverse‐transcribed with SUPERSCRIPT RT using oligo dT as primer to synthesize first‐strand cDNA. mRNA levels of peroxisome proliferator‐activated receptor gamma (Pparg), CCAAT/enhancer binding protein beta (Cebpb), preadipocyte factor 1 (Pref‐1), and Interleukin 6 (Il‐6) genes were measured by real‐time quantitative RT‐PCR. For protein levels, adipose tissue and liver were lysed in RIPA buffer and lysates subjected to SDA‐PAGE and Western blotting. Blots were probed with specific antibodies against PPARG and IL‐6, and detection of immunoreactive bands performed using enhanced chemiluminescence. Data analysis was conducted by ANOVA with GraphPad Prism 7. Congenic mice had a 2‐fold increase in gene expression level of Pparg, a major regulator of adipogenesis and lipogenesis, in adipose tissue compared to B6 mice on chow. HFD significantly increased the gene expression level of Pparg in both B6 and congenic mice, with greater degree in congenic mice. This trend was also the case for the protein expression level of PPARG in adipose tissue, as well as in liver, but it did not reach statistical significance. There were no significant differences in gene expression levels of Pref‐1, the most upstream repressor to adipocyte differentiation, and Cebpb, a major transcription factor during an early stage of adipogenesis, in adipose tissue between congenic and B6 mice for either diet. The gene expression level of Il‐6, a proinflammatory marker, was increased by HFD in both congenic and B6 mice, without genotype difference. A similar trend was also shown in IL‐6 protein levels in adipose tissue, as well as in liver, but it did not reach statistical significance. Collectively, we demonstrated that gene expression levels of Pparg were significantly upregulated in adipose tissue of B6.TH‐Chr1–128Mb congenic mice on chow and HFD.Support or Funding InformationThis work was supported by NASA WVSGC Graduate Fellowship to JKP and in part by NIH/NIDDK Grant R01DK077202 and institutional start‐up funding from Marshall University to JHK.

A novel role for the Wnt inhibitor APCDD1 in adipocyte differentiation: Implications for diet-induced obesity

Impaired adipogenic differentiation during diet-induced obesity (DIO) promotes adipocyte hypertrophy and inflammation, thereby contributing to metabolic disease. Adenomatosis polyposis coli down-regulated 1 (APCDD1) has recently been identified as an inhibitor of Wnt signaling, a key regulator of adipogenic differentiation. Here we report a novel role for APCDD1 in adipogenic differentiation via repression of Wnt signaling and an epigenetic linkage between miR-130 and APCDD1 in DIO. APCDD1 expression was significantly up-regulated in mature adipocytes compared with undifferentiated preadipocytes in both human and mouse subcutaneous adipose tissues. siRNA-based silencing of APCDD1 in 3T3-L1 preadipocytes markedly increased the expression of Wnt signaling proteins (Wnt3a, Wnt5a, Wnt10b, LRP5, and β-catenin) and inhibited the expression of adipocyte differentiation markers (CCAAT/enhancer-binding protein α (C/EBPα) and peroxisome proliferator-activated receptor γ (PPARγ)) and lipid droplet accumulation, whereas adenovirus-mediated overexpression of APCDD1 enhanced adipogenic differentiation. Notably, DIO mice exhibited reduced APCDD1 expression and increased Wnt expression in both subcutaneous and visceral adipose tissues and impaired adipogenic differentiation in vitro Mechanistically, we found that miR-130, whose expression is up-regulated in adipose tissues of DIO mice, could directly target the 3'-untranslated region of the APCDD1 gene. Furthermore, transfection of an miR-130 inhibitor in preadipocytes enhanced, whereas an miR-130 mimic blunted, adipogenic differentiation, suggesting that miR-130 contributes to impaired adipogenic differentiation during DIO by repressing APCDD1 expression. Finally, human subcutaneous adipose tissues isolated from obese individuals exhibited reduced expression of APCDD1, C/EBPα, and PPARγ compared with those from non-obese subjects. Taken together, these novel findings suggest that APCDD1 positively regulates adipogenic differentiation and that its down-regulation by miR-130 during DIO may contribute to impaired adipogenic differentiation and obesity-related metabolic disease.

Ginsenoside F1 attenuates lipid accumulation and triglycerides content in 3T3-L1 adipocytes with the modulation of reactive oxygen species (ROS) production through PPAR-γ/JAK2 signaling responses

Obesity is the severe health concern worldwide, causing a highest risk of mortality rate every year in different countries. During adipogenesis interactions with peroxisome proliferator-activated receptor gamma are mainly responsible for the development of obesity with regulation of the various transcription factors. It causes the increase of lipid accumulation, triglyceride content as well as reactive oxygen species production in adipocyte differentiation. Current drugs used as anti-obesity associated with several side effects. The present study was therefore conducted, to evaluate the protective and inhibitory effect of ginsenoside F1 from Panax ginseng on lipid accumulation and reactive oxygen species production by an in silico and in vitro study using 3T3-L1 cells. The structures of F1 compound has been obtained from own in-house Panax ginseng saponin data base. In silico molecular docking and drug likeness properties including absorption, distribution, metabolism and excretion, and toxicity and prediction of biological activity were performed in order to assess and investigate the antiobesity activity of F1. Molecular docking study showed that F1 exhibited strong hydrogen binding affinity with Janus activated protein kinase-2 greater than −9.2 Kcal/mol compared to co-crystalized ligand 3kc and nicotinamide (control drug) −8.2 kcal/mol and −4.6 Kcal/mol respectively. In addition, our in vitro results indicated that F1 also showed significant inhibitory effect on the triglyceride content and reactive oxygen species production in adipocyte differentiation. Moreover, F1 significantly reduced the mRNA expression of adipogenesis markers such as peroxisome proliferator-activated receptor gamma, adipocyte fatty acid-binding protein, and Janus activated prot, which are responsible for lipid accumulation. Our findings demonstrated that ginsenoside F1 inhibits lipid accumulation and reactive oxygen species generation by downregulating the expression adipocyte differentiation markers in mature 3T3-L1 cells.

Thyroid hormone acting via TRβ induces expression of browning genes in mouse bone marrow adipose tissue.

Mutant hypothyroid mouse models have recently shown that thyroid hormone is critical for skeletal development during an important prepubertal growth period. Additionally, thyroid hormone negatively regulates total body fat, consistent with the well-established effects of thyroid hormone on energy and fat metabolism. Since bone marrow mesenchymal stromal cells differentiate into both adipocytes and osteoblasts and a relationship between bone marrow adipogenesis and osteogenesis has been predicted, we hypothesized thyroid hormone deficiency during the postnatal growth period increases marrow adiposity in mice. Marrow adiposity in TH-deficient (Tshr -/-) mice treated with T3/T4, TH receptor β-specific agonist GC-1, or vehicle control was evaluated via dual-energy X-ray absorptiometry and osmium micro-computed tomography. To further examine the mechanism for thyroid hormone regulation of marrow adiposity, we used real-time RT-PCR to measure the effects of thyroid hormone on adipocyte differentiation markers in primary mouse bone marrow mesenchymal stromal cells and two mouse cell lines in vitro and in Tshr -/- mice in vivo. Marrow adiposity increased >20% (P < 0.01) in Tshr -/- mice at 3 weeks of age, and treatment with T3/T4 when serum thyroid hormone normally increases (day 5-14) rescued this phenotype. Furthermore, GC-1 rescued this phenotype equally well, suggesting this thyroid hormone effect is in part mediated via TRβ signaling. Treatment of bone marrow mesenchymal stromal or ST2 cells with T3 or GC-1 significantly increased expression of several brown/beige fat markers. Moreover, injection of T3/T4 increased browning-specific markers in white fat of Tshr -/- mice. These data suggest that thyroid hormone regulation of marrow adiposity is mediated at least in part via activation of TRβ signaling.

Peptide Fraction pOh2 Exerts Antiadipogenic Activity through Inhibition of C/EBP-α and PPAR-γ Expression in 3T3-L1 Adipocytes.

Many studies have comprehensively examined the venom of Ophiophagus hannah snake. Its venom comprises different compounds exhibiting a wide range of pharmacological activities. In this investigation, four peptide fractions (PFs), ranging from 3 kDa to 10 kDa, isolated from the Vietnamese snake venom of O. hannah were separated by HPLC and investigated for their inhibitory activity on adipogenesis in 3T3-L1 adipocytes. The most effective PF was then further purified, generating two peptides, pOh1 and pOh2. Upon investigation of these two peptides on 3T3-L1 adipocytes, it was revealed that, at 10 μg/mL, pOh2 was able to inhibit the lipid accumulation in 3T3-L1 adipocytes by up to 56%, without affecting cell viability. Furthermore, the pOh2 downregulated the gene expression of important transcription factors C/EBP-α and PPAR-γ. In addition, aP2 and GPDH adipocyte-specific markers were also significantly reduced compared to untreated differentiated cells. Taken together, pOh2 inhibited the expression of key transcription factors C/EBP-α and PPAR-γ and their target genes, aP2 and GPDH, thereby blocking the adipocyte differentiation. In conclusion, this novel class of peptide might have potential for in vivo antiobesity effects.

Critical role of the peroxisomal protein PEX16 in white adipocyte development and lipid homeostasis

The importance of peroxisomes for adipocyte function is poorly understood. Herein, we provide insights into the critical role of peroxin 16 (PEX16)-mediated peroxisome biogenesis in adipocyte development and lipid metabolism. Pex16 is highly expressed in adipose tissues and upregulated during adipogenesis of murine and human cells. We demonstrate that Pex16 is a target gene of the adipogenesis “master-regulator” PPARγ. Stable silencing of Pex16 in 3T3-L1 cells strongly reduced the number of peroxisomes while mitochondrial number was unaffected. Concomitantly, peroxisomal fatty acid (FA) oxidation was reduced, thereby causing accumulation of long- and very long-chain (polyunsaturated) FAs and reduction of odd-chain FAs. Further, Pex16-silencing decreased cellular oxygen consumption and increased FA release. Additionally, silencing of Pex16 impaired adipocyte differentiation, lipogenic and adipogenic marker gene expression, and cellular triglyceride stores. Addition of PPARγ agonist rosiglitazone and peroxisome-related lipid species to Pex16-silenced 3T3-L1 cells rescued adipogenesis. These data provide evidence that PEX16 is required for peroxisome biogenesis and highlights the relevance of peroxisomes for adipogenesis and adipocyte lipid metabolism.

Combinations of bio-active dietary constituents affect human white adipocyte function in-vitro.

BackgroundSpecific bio-active dietary compounds modulate numerous metabolic processes in adipose tissue (AT), including pre-adipocyte proliferation and differentiation. AT dysfunction, rather than an increased fat mass per se, is strongly associated with the development of insulin resistance and is characterized by impaired adipogenesis, hypertrophic adipocytes, inflammation, and impairments in substrate metabolism. A better understanding of mechanisms underlying AT dysfunction may provide new strategies for the treatment of obesity-associated metabolic diseases. Here we evaluated the role of (all-E)-lycopene (Lyc), eicosapentaenoic acid (EPA) or trans-resveratrol (Res) and combinations thereof on human white adipocyte function.MethodsIn-vitro differentiating human pre-adipocytes were treated with EPA, Lyc and Res or their combinations for 14 days. The effects on intracellular lipid droplet (LD) accumulation, secreted anti- and pro-inflammatory cyto-/adipokines (e.g. adiponectin, IL-6, IL-8/CXCL-8 and MCP-1/CCL2) and on gene expression of markers of adipocyte differentiation and substrate metabolism (e.g. PPAR-gamma, C/EBP-alpha, GLUT-4, FAS, ATGL, HSL, and PLIN-1) were measured by fluorescent microscopy (Cellomics™), multi-parametric LiquiChip® technology and quantitative RT-PCR, respectively.ResultsTreatment of differentiating adipocytes for 14 days with the combination of Lyc/Res and EPA/Res resulted in significantly inhibited LD formation (~ -25 and -20%, respectively) compared to the effects of the single compounds. These morphological changes were accompanied by increased mRNA levels of the adipogenic marker PPAR-gamma and the lipase ATGL and by decreased expression levels of lipogenic markers (LPL, FAS, GLUT-4) and the LD-covering protein PLIN-1. In addition, a blunted adipocyte secretion of pro-inflammatory cytokines (IL-6 and MCP-1) and adiponectin was observed following treatment with these compounds.ConclusionThe combination of the dietary bio-actives Lyc and EPA with Res might influence adipocyte function by affecting the balance between adipogenic, lipogenic and lipolytic gene expression, resulting in a reduced LD storage and a less inflammatory secretion profile. Taken together, our results indicate that combinations of dietary compounds may be beneficial for the prevention and treatment of metabolic disorders via effects on human white adipocyte function.Electronic supplementary materialThe online version of this article (doi:10.1186/s12986-016-0143-5) contains supplementary material, which is available to authorized users.

Graphene/single-walled carbon nanotube hybrids promoting osteogenic differentiation of mesenchymal stem cells by activating p38 signaling pathway.

Carbon nanomaterials are becoming increasingly significant in biomedical fields since they exhibit exceptional physicochemical and biocompatible properties. Today, the stem cells offer potentially new therapeutic approaches in tissue engineering and regenerative medicine. However, the induction of differentiation into specific lineages remains challenging, which provoked us to explore the biomedical applications of carbon nanomaterials in stem cells. In this study, we investigated the interactions between graphene/single-walled carbon nanotube (G/SWCNT) hybrids and rat mesenchymal stem cells (rMSCs) and focused on the proliferation and differentiation of rMSCs treated with G/SWCNT hybrids. Cell viability and morphology were evaluated using cell counting kit-8 assay and immunofluorescence staining, respectively. Osteogenic differentiation evaluated by alkaline phosphatase activity of MSCs proved to be higher after treatment with G/SWCNT hybrids, and the mineralized matrix nodule formation was also enhanced. In addition, the expression levels of osteogenic-associated genes were upregulated, while the adipocyte-specific markers were downregulated. Consistent with these results, we illustrated that the effect of G/SWCNT hybrids on the process of osteogenic differentiation of rMSCs can be modulated by activating the p38 signaling pathway and inhibiting the extracellular signal-regulated kinase 1/2 pathway. Nevertheless, our study suggests that carbon nanomaterials offer a promising platform for regenerative medicine in the near future.

6-Gingerol Suppresses Adipocyte-Derived Mediators of Inflammation In Vitro and in High-Fat Diet-Induced Obese Zebra Fish.

The present study was performed to investigate the molecular mechanism of 6-gingerol on adipocyte-mediated systemic inflammation in vitro and in high-fat diet-induced obese zebra fish. 6-Gingerol decreased adipogenesis due to the suppression of adipocyte differentiation markers, including peroxisome proliferator-activated receptor gamma, CCAATT enhancer binding protein α, and adipocyte protein 2, and triglyceride synthesis enzymes, including sterol regulatory element-binding protein-1, fatty acid synthase, lysophosphatidic acid acyltransferase, and acyl-coA : diacylglycerol acyltransferase 1, in 3T3-L1. A coculture insert system using 3T3-L1 with RAW 264.7 (coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages) revealed that 6-gingerol increased anti-inflammatory cytokine interleukin-10. The expression of TNFα, monocyte chemotactic protein-1, interleukin-1β, and interleukin-6 were decreased in the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages treated with 6-gingerol. Moreover, the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages treated with 6-gingerol inhibited the protein expression of TNFα and monocyte chemotactic protein-1 in RAW 264.7. 6-Gingerol decreased c-JUN N-terminal kinase and I kappa B kinase beta and its downstream target AP-1 expression in the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages. Furthermore, 6-gingerol decreased the expression of inducible nitric oxide synthase stimulated by the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages in RAW 264.7 and attenuated nitric oxide production in diet-induced obese zebra fish. Our results suggest that 6-gingerol suppresses inflammation through the regulation of the c-JUN N-terminal kinase-I kappa B kinase beta and its downstream targets.

Genomic and lipidomic actions of nandrolone on detached rotator cuff muscle in sheep

Reversal of fatty infiltration of pennate rotator cuff muscle after tendon release is hitherto impossible. The administration of nandrolone starting at the time of tendon release prevents the increase in fat content, but does not revert established fatty infiltration.We hypothesised that tendon release and myotendinous retraction cause alterations in lipid related gene expression leading to fatty muscle infiltration, which can be suppressed by nandrolone through its genomic actions if applied immediately after tendon release.The effects of infraspinatus tendon release and subsequent tendon repair at 16 weeks were studied in six Swiss Alpine sheep. In the interventional groups, 150mg nandrolone was administered weekly after tendon release until sacrifice (N22W, n=6) or starting at the time of repair (N6W, n=6). Infraspinatus volume, composition, expressed transcripts, lipids, and selected proteins were analyzed at baseline, 16 and 22 weeks.Tendon release reduced infraspinatus volume by 22% and increased fat content from 11% to 38%. These changes were not affected by repair. Fatty infiltration was associated with up-regulation of 227 lipid species, and increased levels of the adipocyte differentiation marker PPARG2 (peroxisome proliferator-activated receptor gamma 2). Nandrolone abrogated lipid accumulation, halved the loss in fiber area percentage, and up-regulated androgen receptor levels and transcript expression in the N22W but not the N6W group.The results document that nandrolone mitigates muscle-to-fat transformation after tendon release via a general down-regulation of lipid accumulation concomitantly with up-regulated expression of its nuclear receptor and downstream transcripts in skeletal muscle. Reduced responsiveness of retracted muscle to nandrolone as observed in the N6W group is reflected by a down-regulated transcript response.

Adipogenesis, lipogenesis and lipolysis is stimulated by mild but not severe hypoxia in 3T3-L1 cells

In-vitro investigation of the effects of hypoxia is limited by physical laws of gas diffusion and cellular O2 consumption, making prolonged exposures to stable O2 concentrations impossible. Using a gas-permeable cultureware, chronic effects of mild and severe hypoxia on triglyceride accumulation, lipid droplet size distribution, spontaneous lipolysis and gene expression of adipocyte-specific markers were assessed. 3T3-L1 cells were differentiated under 20%, 4% or 1% O2 using a gas-permeable cultureware. Triglyceride accumulation, expression of genes characteristic for advanced adipocyte differentiation and involvement of key lipogenesis enzymes were assessed after exposures. Lipogenesis increased by 375% under mild hypoxia, but dropped by 43% in severe hypoxia. Mild, but not severe, hypoxia increased formation of large lipid droplets 6.4 fold and strongly induced gene expression of adipocyte-specific markers. Spontaneous lipolysis increased by 488% in mild, but only by 135% in severe hypoxia. Inhibition of ATP-dependent citrate lyase suppressed hypoxia-induced lipogenesis by 81% and 85%. Activation of HIF inhibited lipogenesis by 59%. Mild, but not severe, hypoxia stimulates lipolysis and promotes adipocyte differentiation, probably through excess of acetyl-CoA originating from tricarboxylic acid cycle independently of HIF activation.

Abstract 344: Antimitotic effect and complex of nonmitotic effect on tumor biology of eribulin mesilate in soft tissue sarcoma model

Abstract Background and objectives: Eribulin mesilate (Eribulin), a first-in-class halichondrin B-based microtubule dynamics inhibitor, has been shown to have novel mechanism of actions (reversion of epithelial-mesenchymal transition of tumor cell and promotion of vascular remodeling) beyond its established antimitotic activity in breast cancer models, leading to decrease of tumor metastatic potency. Recently, eribulin showed trends towards greater overall survival compared to Dacarbazine in subjects with leiomyosarcoma and liposarcoma patients in phase 3 trial. The objective of this study was to investigate the effect of eribulin on tumor cell proliferation of soft tissue sarcoma (STS). Furthermore, the effects of eribulin on morphology and gene expression of differentiation markers against STS and tumor blood perfusion in a leiomyosarcoma xenograft model were examined. Methods: Anti-proliferation activity of eribulin was examined in 6 human STS cell lines. Anti-tumor activity of eribulin was examined in xenografts of A-673 (Ewing's sarcoma), SK-LMS-1 (leiomyosarcoma), SW 872 (liposarcoma), and HT-1080 (fibrosarcoma) at several doses. To analyze the effect of eribulin on in vitro morphological change, 3 human STS cell lines were treated with eribulin at 1, 3, and 10 nmol/L for 7 days. Gene expression change was analyzed by microarray and differentiation marker genes in 2 cell lines were confirmed by qPCR analysis. For tumor blood perfusion analysis, eribulin was administered at doses of 0.38, 0.75, and 1.5 mg/kg one shot in SK-LMS-1 xenografts, and tumor blood perfusion was calculated as Hoechst-positive areas in tumor cryosections 5 days after the administration. Results: Eribulin showed anti-proliferation activity in vitro against all 6 cell lines in a dose dependent manner with 50% inhibitory concentration values of around 1 nmol/L. Eribulin showed significant antitumor activity against 4 xenografts in a dose dependent manner. Eribulin treatment for 7 days induced apparent morphological change in 3 cell lines. Of these, expression level of smooth muscle differentiation marker gene CNN1 was 7.1-fold upregulated in SK-UT-1 and adipocyte differentiation marker genes MYLK, C/EBPβ, and KIF23 were 1.9-, 1.4-, and 1.3-fold upregulated in SW 872, respectively, by 10 nmol/L treatment compared to vehicle control. In addition, eribulin significantly enhanced tumor blood perfusion compared to vehicle control 5 days after the administration in SK-LMS-1 xenografts at all doses.Conclusions: Eribulin showed anti-proliferation activity against all of STS cell lines we tested in vitro and in vivo. Furthermore, eribulin might cause differentiation of STS cells and remodeling of tumor vasculature to enhance tumor blood perfusion. These data supported that eribulin showed mitotic effect and non-mitotic effect, which may decrease tumor metastatic potency in STS, as well in breast cancer. Citation Format: Satoshi Kawano, Makoto Asano, Yusuke Adachi, Junji Matsui. Antimitotic effect and complex of nonmitotic effect on tumor biology of eribulin mesilate in soft tissue sarcoma model. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 344.

Circulating adipocyte-derived extracellular vesicles are novel markers of metabolic stress.

• Extensive characterization of 3T3L1 EVs identified perilipin A in their composition. • Circulating EVs are elevated in obese mice and associated with glucose intolerance. • Circulating EVs are elevated in obese human and correlated with metabolic factors. • Perilipin A and EV levels are increased in the circulation of obese mice and human. • Circulating EV and perilipin A levels decrease with low calorie intervention.