Lipid Droplet Formation In Adipocytes Research Articles

Perilipin 1-3 in grass carp Ctenopharyngodon idella: molecular characterization, gene structure, tissue distribution, and mRNA expression in DHA-induced lipid droplet formation in adipocytes.

Perilipin family is the main structural proteins of lipid droplet (LD) that is intracellular neutral lipid store ponds, and regulates LD assembly and formation, and is crucial for lipid metabolism. Here three paralogs of perilipin family were characterized from grass carp and their complete coding sequences (CDS) were obtained, including perilipin1, perilipin2, and perilipin3, coding peptides of 492, 454, and 419 amino acids, respectively. The alignment of the homology of grass carp perilipin deduced amino acid sequences with other teleost species showed that the homology with mammalian was less than 55%. PAT (perilipin) domain in mammalian was also predicted in grass carp perilipin 1-3 proteins. Genomic organization analysis revealed that grass carp perilipin1 contained 6 coding exons, while both perilipin2 and perilipin3 consisted of 7 coding exons. The mRNA encoding three paralogs were expressed in a wide range of tissues; perilipin1-3 were primarily expressed in adipose tissue and liver; besides, perilipin3 was also highly expressed in the heart. In vitro, 200μM DHA increased the proportion of smaller lipid droplets effectively in fully differentiated adipocytes of grass carp. The mRNA expression of perilipin1, perilipin2, and perilipin3 was significantly increased in the adipocytes treated with DHA (P < 0.05, P < 0.01). The same responses of different paralogs in the adipocytes during DHA treatment suggest that they might play synergistic roles in the formation of LDs.

Role of LXR alpha in regulating expression of glucose transporter 4 in adipocytes — Investigation on improvement of health of diabetic patients

BackgroundThe insulin-responsive glucose transporter 4 (GLUT4) plays prominent role in insulin-mediated facilitated glucose uptake into most of the cell types, majorly muscle, liver and adipose tissue. Impaired expression of GLUT4 has been linked to insulin resistance and diabetes. In adipocytes, excess lipids that are generated from liver and by de novo lipogenesis utilizing blood sugar, are stored. There are various nuclear factors, co-factors and signaling mechanisms that directly and indirectly regulate glucose transporter activity in adipocytes. Molecular mechanism behind the regulation of GLUT4 in adipocytes has not elucidated well. ObjectiveTherefore, the present study focuses to explore the role of Liver X receptor- alpha (LXRα) on GLUT4 expression and the possible co-factors involved during in vitro adipogenesis and is assessed by modulating the activity of LXRα with specific agonist and antagonist ligands in 3T3L1 differentiated cells. ResultsThe results demonstrate that SR 9238 (300nM), a strong inhibitor of LXRα, decreased the rate of adipogenesis through reduced lipid droplet formation in adipocytes without affecting the cell morphology. The FMOC-l-Leucine (FLL), a known partial ligand of PPARγ, enhanced the expression of LXRα. Thus, it can be concluded that LXRα has a significant role in adipocyte differentiation and FLL interaction promoted the transcription of LXRα, thereby promotes GLUT4 expression. The results in this study identified the role of LXRα in regulating the expression of GLUT4 through SRC1. ConclusionThe study is of much relevance in treatment of diabetes and also opens the possibility of identifying new drug molecules that target LXRα.

ANGPTL8 regulates adipocytes differentiation and adipogenesis in bovine

The formation of bovine adipose tissue involves complex developmental and physiological processes that play a vital role in determining the quality of beef; however, the regulatory mechanisms are largely unknown. Angiopoietin-like protein 8 (ANGPTL8) has been reported to be involved in the development of adipose tissue; however, the mechanism of adipogenesis which is regulated by ANGPTL8 has not been revealed in cattle. In this study, RT-qPCR and Oil Red O staining were performed to detect the expression of ANGPTL8 and adipocyte differentiation in bovine. We found that ANGPTL8 had a high expression level in adipose tissue and that the expression pattern was consistent with those of PPARγ, C/EBPα and LPL which are key regulators and transcription factors involved in preadipocyte differentiation and adipogenesis. The overexpression of ANGPTL8 by the adenovirus vector promoted lipid droplet formation in adipocytes. Thus, we speculated that ANGPTL8 could significantly enhance lipid deposition. Moreover, the expression of LPL and SREBP1, key genes inhibiting adipogenesis, was significantly decreased by ANGPTL8 overexpression. These results suggested that ANGPTL8 promotes adipocyte differentiation. In conclusion, we consider that ANGPTL8 regulates adipocyte differentiation and adipogenesis in bovine.

Inhibition of lipid droplet formation by Ser/Thr protein phosphatase PPM1D inhibitor, SL-176.

Obesity is a worldwide public health problem, which is associated with various severe diseases including diabetes, hypertension, atherosclerosis, and cancer. Recent studies have revealed that combination treatment of several different compounds using low doses is effective to reduce side effects. Thus, there is a need to develop an efficient inhibitor for reducing lipid droplets with a divergent target/pathway. Ser/Thr protein phosphatase PPM1D is involved in cellular metabolic processes and is a promising target for anti-obesity treatment. We have previously developed a potent and specific PPM1D inhibitor, SL-176. In this study, we demonstrated that significant reduction of lipid droplet formation in adipocytes by the PPM1D specific inhibitor, SL-176. Using Oil-red O staining and fluorescent imaging of lipid droplet, we found that treatment of SL-176 significantly suppressed lipid droplet formation of 3T3-L1 cells both in amount and in size. SL-176 also repressed mRNA and protein expression of PPARγ and C/EBPα, adipogenic markers, at nontoxic conditions. Thus, SL-176 is a unique and potent inhibitor of lipid droplet formation that acts via PPM1D, a novel target toward inhibiting adipocyte differentiation.

An emerging interplay between altered human lipid metabolism, lipodystrophy and aging

LMNA -Lipodystrophies are a group of heterogeneous syndromes, with either genetic or acquired origin, characterized by the accumulation of prelamin A, an inmature form of the protein lamin A, one of the major components of the nuclear lamina. Several molecular studies suggest lamin A is involved in adipocyte development, the disruption of which leads to compromised regulation of adipogenesis, adipocyte lipid droplet formation and maintenance, and subsequent secondary dysfunctions in fat metabolism. Moreover, these diseases clinically present with generalized or partial fat atrophy connected with metabolic complications, such as insulin-resistant diabetes and dyslipidemia, in addition to age associated manifestations. There is a real need to increase our understanding regarding these syndromes because of their import in human health and the lack of knowledge of their etiopathology. To gain deeper insights into these metabolic diseases, we have taken advantage of a previously generated “disease in a dish” model of human LMNA -lipodystrophy based on the pathological accumulation of the precursor prelamin A in stem cell derived adipocytes. This experimental model recapitulates phenotypes observed in lipodystrophic patient’s samples and animal models, and it has been critical in elucidating new insights into the molecular mechanisms governing this set of disorders. Recently, we have identified alterations in fundamental processes of lipid homeostasis such as lipolysis, as well as mitochondrial and endoplasmic reticulum functions, similar to what can be observed in some metabolic and aging phenotypes. Additionally, the lipidomic profile of this lipodystrophic experimental model displayed a lipid metabolic signature similar to aging systems, providing new information concerning metabolic pathways affected during the aging process. By clarifying the fundamental mechanisms governing these aging associated diseases, future novel interventions could be developed that will at least delay the appearance of aging phenotypes and thereby increase the healthspan or disease-free time of an individual.

Repressive effects of oat extracts on intracellular lipid-droplet formation in adipocytes and a three-dimensional subcutaneous adipose tissue model.

We assessed the repression of lipid-droplet formation in mouse mesenchymal stromal preadipocytes OP9 by specified oat extracts (Hatomugi, Coix lacryma-jobi var. ma-yuen) named "SPH" which were proteolytically and glucosyl-transferredly prepared from finely-milled oat whole-grain. Stimulation of OP9 preadipocytes with insulin-containing serum-replacement promoted differentiation to adipocytes, concurrently with an increase in the intracellular lipid droplets by 51.5%, which were repressed by SPH-bulk or SPH-water-extract at 840ppm, to 33.5% or 46.9%, respectively, but not by SPH-ethanol-extract at the same dose, showing the hydrophilic property of the anti-adipogenetic ingredients. The intracellular lipid droplets were scanty for intact preadipocytes, small-sized but abundant for the SPH-unadministered adipocytes, and large-sized but few for SPH-bulk-administered adipocytes being coexistent with many lipid-droplet-lacking viable cells, suggesting "the all-or-none rule" for lipid-droplet generation in cell-to-cell. Hydrogen-peroxide-induced cell death in human epidermal keratinocytes HaCaT was prevented by SPH-bulk at 100 or 150ppm by 5.6-8.1%, being consistent with higher viabilities of SPH-bulk-administered OP9 cells, together with repressions of both cell shrinkage and cell detachment from the culture substratum. In three-dimensional subcutaneous adipose tissue models reconstructed with HaCaT-keratinocytes and OP9-preadipocytes, lipid droplets were accumulated in dermal OP9-cell-parts, and repressed to 43.5% by SPH-bulk at 840ppm concurrently with marked diminishment of huge aggregates of lipid droplets. Thus SPH-bulk suppresses adipogenesis-associated lipid-droplet accumulation during differentiation of OP9 preadipocytes together with lowered cytotoxicity to either HaCaT keratinocytes or the preadipocytes.

Perilipin-mediated lipid droplet formation in adipocytes promotes sterol regulatory element-binding protein-1 processing and triacylglyceride accumulation.

Sterol regulatory element-binding protein-1 (SREBP-1) has been thought to be a critical factor that assists adipogenesis. During adipogenesis SREBP-1 stimulates lipogenic gene expression, and peroxisome proliferator-activated receptor γ (PPARγ) enhances perilipin (plin) gene expression, resulting in generating lipid droplets (LDs) to store triacylglycerol (TAG) in adipocytes. Plin coats adipocyte LDs and protects them from lipolysis. Here we show in white adipose tissue (WAT) of plin−/− mice that nuclear active SREBP-1 and its target gene expression, but not nuclear SREBP-2, significantly decreased on attenuated LD formation. When plin−/− mouse embryonic fibroblasts (MEFs) differentiated into adipocytes, attenuated LDs were formed and nuclear SREBP-1 decreased, but enforced plin expression restored them to their original state. Since LDs are largely derived from the endoplasmic reticulum (ER), alterations in the ER cholesterol content were investigated during adipogenesis of 3T3-L1 cells. The ER cholesterol greatly reduced in differentiated adipocytes. The ER cholesterol level in plin−/− WAT was significantly higher than that of wild-type mice, suggesting that increased LD formation caused a change in ER environment along with a decrease in cholesterol. When GFP-SREBP-1 fusion proteins were exogenously expressed in 3T3-L1 cells, a mutant protein lacking the S1P cleavage site was poorly processed during adipogenesis, providing evidence of the increased canonical pathway for SREBP processing in which SREBP-1 is activated by two cleavage enzymes in the Golgi. Therefore, LD biogenesis may create the ER microenvironment favorable for SREBP-1 activation. We describe the novel interplay between LD formation and SREBP-1 activation through a positive feedback loop.

Transcriptional activation of Cidec by PPARγ2 in adipocyte

Cidec is a lipid droplet-associated protein, which inhibits lipolysis, leading to the accumulation of triglycerides in adipocytes. However, the transcriptional regulation of Cidec in adipocyte remains unknown. In the present study we investigated that the mouse Cidec transcript is regulated by PPARγ2. After the differentiation of adipocyte, the expression pattern of Cidec was similar to that of PPARγ2. In the presence of a PPARγ agonist, the level of Cidec mRNA was highly increased. In addition, putative PPRE sites were identified in the Cidec promoter. By chromatin immunoprecipitation assay and reporter assay, we observed the binding of PPARγ2 to the promoter of Cidec. Gel shift assay and the mutagenesis study were showed that the −219/−207 region of the Cidec promoter could function as a PPRE of the Cidec promoter. These results suggest that PPARγ2 is required for the transcriptional activity of Cidec during adipogenesis, which could be contributed to understand the molecular mechanism of lipid droplet formation in adipocytes.