Lipid Droplet Levels Research Articles

Global impacts of peroxisome and pexophagy dysfunction revealed through multi-omics analyses of lon2 and atg2 mutants.

Peroxisomes house diverse metabolic pathways that are essential for plant and animal survival, including enzymes that produce or inactivate toxic byproducts. Despite the importance of peroxisomes and their collaborations with other organelles, the mechanisms that trigger or prevent peroxisome turnover and the cellular impacts of impaired peroxisomes are incompletely understood. When Arabidopsis thaliana LON2, a peroxisomal protein with chaperone and protease capacity, is disrupted, metabolic dysfunction and protein instability in peroxisomes ensue. Paradoxically, preventing autophagy in lon2 mutants appears to normalize peroxisomal metabolism and stabilize peroxisomal proteins-hinting at a role for autophagy in causing the peroxisomal defects observed in lon2 seedlings. Using a combination of transcriptomics, proteomics, and in silico investigations, we compared wild type to lon2 and autophagy null mutants and double mutants. Through this analysis, we found that impeding autophagy via an atg2 null mutation alleviated several of the global defects observed when LON2 is absent. Moreover, we revealed processes influenced by LON2 that are independent of autophagy, including impacts on lipid droplet and chloroplast protein levels. Finally, we identified and classified potential LON2 substrates, which include proteins that might provide signal(s) for pexophagy.

A novel fluorescent probe for viscosity and polarity detection in real tobacco root cells and biological imaging.

The disruption of lipid droplet function is associated with the pathogenesis of various diseases. Clarifying the response behavior of lipid droplets to the microenvironment at the cellular level is of great significance. Plant lipids not only exist in phospholipids in cell membranes, but also in aromatic essential oils. Monitoring the level of lipid droplets in plant cells using fluorescent probes provides a simple method for screening lipid-rich varieties. We synthesized a polarity-viscosity responsive coumarin fluorescent probe, Cou-CN, which achieved sensitive detection of polarity and viscosity in dilute solution environments by constructing this simple probe with ICT and TICT properties and verifying it using Gaussian computational simulation. Cou-CN exhibited good lipid droplet illumination effects in HepG2 cells with a correlation coefficient of 0.92 compared to the commercial lipid droplet dye BODIPY. Additionally, co-staining the probe with the lipophilic commercial dye Nile Red in tobacco root stem seedling cells resulted in a high correlation coefficient of 0.9.

Hepatic and metabolic outcomes induced by sub-chronic exposure to polystyrene microplastics in mice.

Microplastics (MPs) have attracted significant attention due to their global distribution in living environments. Although some studies have reported MP-induced hepatotoxicity in mouse models, a systematic approach to MP-mediated liver toxicity was still lacking. Therefore, we used a mouse model to study the sub-chronic effects of MP exposure on the liver. Female C57BL/6 mice, aged 6weeks, received an oral administration of 0.3mg of Nile Red-labeled polystyrene (PS) microplastics, with particle sizes of 0.5µm (submicron) and 5µm (micron), via gavage, while control mice received vehicle only. Each mouse was exposed to MPs twice a week for 12weeks. After sacrifice, the levels of MP accumulation, oxidative stress, inflammation, and pathological changes were measured in the mouse liver, and blood samples were collected for serum biochemistry analysis. Our results demonstrated that 0.5µm PS-MPs were accumulated in mouse livers post-MP exposure, but not in the 5µm MP exposure group. Simultaneously, increased levels of glucose, triglyceride, alanine transaminase (ALT), aspartate transaminase (AST), superoxide dismutase, 4-hydroxy-2-nonenal-mercapturic acid (HNE-MA), interleukin-6, and lipid droplets were found in the 0.5µm MP exposure group, while the fewer responses, including elevated liver weight index, glucose, high-density lipoprotein, AST, and decreased HNE-MA were observed in 5µm MP exposure group. These results indicate that sub-chronic exposure to submicron MPs causes MP deposition in mouse livers, which further induces oxidative stress, increases inflammatory cytokines and perturbs glucose and lipid homeostasis, which might trigger more severe metabolic dysfunction or non-alcoholic steatohepatitis-like hepatotoxicity.

Tanshinone IIA promotes osteogenic differentiation potential and suppresses adipogenic differentiation potential of bone marrow mesenchymal stem cells.

Tanshinone IIA (Tan IIA) may have therapeutic effects on avascular necrosis of the femoral head (ANFH) by targeting bone marrow mesenchymal stem cells (BMSCs). The effect and underlying mechanism of Tan IIA on adipogenesis and osteogenesis ability of BMSCs remain to be elucidated. In the present study BMSCs were treated with osteogenic or adipogenic differentiation medium with or without Tan IIA under hypoxic environment. Osteogenic differentiation potential was evaluated by alkaline phosphatase (ALP) measurement, alizarin red staining and reverse transcription‑quantitative (RT‑q) PCR of osteogenic marker genes. Adipogenic differentiation potential was evaluated with oil red staining and RT‑qPCR of adipogenic marker genes. Detailed mechanism was explored by RNA‑seq and small molecular treatment during osteogenesis and adipogenesis of BMSCs. ALP level, mineralized nodules and expression level of osteogenic marker genes significantly increased following Tan IIA treatment during osteogenic differentiation of BMSCs. Lipid droplet and expression levels of adipogenic marker genes significantly decreased following Tan IIA treatment during adipogenic differentiation of BMSCs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses of RNA‑seq data indicated increased Akt and TGFβ signaling following Tan IIA treatment. Further western blot assay confirmed that Tan IIA significantly activated Akt/cAMP response element‑binding protein signaling and TGFβ/Smad3 signaling. Application of Akti1/2 (an Akt inhibitor) significantly decreased the promotion effect of osteogenesis induced by Tan IIA, while the addition of SB431542 significantly reduced inhibition effect of adipogenesis caused by Tan IIA. Tan IIA could promote osteogenic differentiation potential of BMSCs by activating AKT signaling and suppress adipogenic differentiation potential of BMSCs by activating TGFβ signaling.

Docosahexaenoic acid supplementation during porcine oocyte in vitro maturation improves oocyte quality and embryonic development by enhancing the homeostasis of energy metabolism

Although supplementation with docosahexaenoic acid (DHA) during porcine oocyte IVM is well-established, the available data are limited due to the lack of consistency. Moreover, to our knowledge, the anti-oxidant effects of DHA on porcine oocytes have not been reported. Hence, this study aimed to examine the effects of DHA supplementation on the regulation of energy metabolism during porcine oocyte maturation to improve oocyte maturation and embryonic development. By supplementing the IVM medium with various DHA concentrations, 25 μM DHA was identified as the optimal concentration which improved intraoocyte glutathione content and enhanced embryonic development after parthenogenesis. Compared to embryos derived from the control group, those derived from SCNT or IVF showed significantly improved blastocyst formation upon DHA supplementation during IVM. In addition, various transcription factors associated with oocyte development and apoptosis in mature oocytes were beneficially regulated in the DHA-treated oocytes. Moreover, DHA improved the AMP-activated protein kinase (AMPK)-regulatory ability of porcine oocytes and ameliorated nuclear maturation and embryonic development, which were decreased by artificially downregulating AMPK. To our knowledge, this is the first study to examine the effects of DHA as an AMPK regulator on oocyte maturation and embryo development in pigs. Furthermore, DHA addition to the IVM medium upregulated the relative expression of genes associated with mitochondrial potential and lipid metabolism. Therefore, the membrane potential of mitochondria (evaluated based on the JC-1 aggregate/JC-1 monomer ratio) and the levels of fatty acids and lipid droplets in matured oocytes increased, resulting in increased ATP synthesis. In conclusion, the DHA treatment of porcine oocytes with 25 μM DHA during IVM enhances the homeostasis of energy metabolism by improving mitochondrial function and lipid metabolism, leading to improved quality of matured oocytes and enhanced embryonic developmental potential of in vitro produced (IVP) embryos. Thus, 25 μM DHA supplementation could serve as a tool for improving the quality of IVP embryos. The study findings provide a basis for further research on improving the production efficiency of cloned animals by securing high-quality matured oocytes and enhancing energy metabolism in mammalian oocytes, including those of pigs.

Benzoate glycosides from Gentiana scabra Bge. and their lipid-lowering activity

Seven undescribed benzoate glycosides (1−7) and five known ones (8−12) were isolated from the rhizomes of Gentiana scabra Bge. Their structures were characterized by comprehensive NMR and MS spectroscopic data analysis. The lipid-lowering effects of these compounds were evaluated by measuring the triglyceride (TG) contents and intracellular lipid droplets (LDs) in oleic acid (OA)-treated HepG2 cells. The results showed that compounds 1, 5, 7, and 11 significantly reduced the TG content at 20 μM, and the Bodipy staining displayed that OA enhanced the levels of LDs in the cell, while these compounds reversed the lipid accumulation caused by OA. These findings provide a basis for further development and utilization of G. scabra as a natural source of potential lipid-lowering agents.

Ultrastructural changes in the adrenal gland in terms of age and gender-related

Objectives: The adrenal glands are endocrine organs that synthesise hormones with crucial functions in the body. This irreplaceable structure performs multiple functions, from metabolism of carbohydrates, fats and proteins to sex development. Any disorder concerning the adrenal glands can give rise to life-threatening conditions. The aging process inevitably affects the adrenal glands. The effects of ageing are marked by a rise in cortisol secretion in the zona fasciculata and a decrease in androgen secretion in the zona reticularis. Elevated cortisol levels in the blood disturb most bodily systems in favour of catabolism, amplifying the cellular decline and destruction that accompanies ageing. The deterioration of the adrenal glands relates not just to spongiocytes and chromaffin cells, but also to the endothelium, which enables circulation. It is clear that a decline in vascular function will have a negative impact on endocrine activities. Our study aims to explore the influence of ageing on the adrenal glands in rats, analysing sex-specific ultrastructural scales. Materials and Methods: A total of 28 Sprague-Dawley rats, 14 males and 14 females, were planned to be used in the study. Of these 28 rats, 4 females and 4 males will constitute the control group. The rats in the control group will be approximately 10 weeks old, and the rats in the experimental group, which will represent the aged group, will be 19 weeks old. All animals in the study will be anaesthetised and then sacrificed by removal of the heart. The right and left common carotid arteries were removed from the sacrificed animals. Collected vessels prepared for TEM examination. For each animal, at least four TEM images were taken from four different sections from the same block. Results: Our findings demonstrate that the ageing process not only affects spongiocytes within the adrenal gland, but also contributes to the deterioration of endothelial cells. As anticipated, our results indicate the presence of senescence and apoptosis in endothelial cells. The observed vascular separations and ruptures are due to the endothelial deterioration. Spongiocytes experienced hypertrophy to compensate for the functional deficiencies following a decrease in their number due to ageing. Elevated levels of lipofuscin, lipid droplets, and lysosomes were discovered in spongiocytes. Impaired endothelium potentially contributes to certain changes in spongiocytes. Conclusions: While ageing-related changes appear similar in both genders, males tend to be more impacted.

Berberine attenuates obesity-induced insulin resistance by inhibiting miR-27a secretion.

Berberine (BBR) is an alkaloid found in plants. It has neuroprotective, anti-inflammatory and lipid-lowering activity. However, the efficacy of treatment with BBR and the mechanisms through which it acts need further study. This study investigated the therapeutic effects and the mechanism of action of BBR on obesity-induced insulin resistance in peripheral tissues. High-fat-fed C57BL/6J mice and low-fat-fed C57BL/6J mice with miR-27a overexpression were given BBR intervention (100 mg/kg, po), and the oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) were performed. Palmitic acid-stimulated hypertrophic adipocyte models were treated with BBR (10 μM). Related indicators and protein expression levels were examined. The AUCs of the OGTT and the ITT in the BBR intervention group were reduced significantly (p < 0.01) (p < 0.05), and the serum biochemical parameters, including FBG, TC, TG and LDL-C were significantly reduced after BBR intervention. In the invitro experiments, the triglyceride level and volume of lipid droplets decreased significantly after BBR intervention (p < 0.01) (p < 0.05). Likewise, BBR ameliorates skeletal muscle and pancreas insulin signalling pathways invivo and invitro. The results showed that BBR significantly ameliorated insulin resistance, reduced body weight and percent body fat and improved serum biochemical parameters in mice. Likewise, BBR reduced triglyceride level and lipid droplet volume in hypertrophic adipocytes, BBR improved obesity effectively. Meanwhile, BBR ameliorated the histomorphology of the pancreas, and skeletal muscle and pancreas insulin related signalling pathways of islets in invitro and invivo experiments. The results further demonstrated that BBR inhibited miR-27a levels in serum from obese mice and supernatant of hypertrophic adipocytes. miR-27a overexpression in low-fat fed mice indicated that miR-27a caused insulin resistance, and BBR intervention significantly improved the miR-27a induced insulin resistance status. This study demonstrates the important role of BBR in obesity-induced peripheral insulin resistance and suggest that the mechanism of its effect may be inhibition of miR-27a secretion.

BCAA metabolism in pancreatic cancer affects lipid balance by regulating fatty acid import into mitochondria

BackgroundPancreatic ductal adenocarcinoma (PDAC) has been associated with the host dysmetabolism of branched-chain amino acids (BCAAs), however, the implications for the role of BCAA metabolism in PDAC development or progression are not clear. The mitochondrial catabolism of valine, leucine, and isoleucine is a multistep process leading to the production of short-chain R-CoA species. They can be subsequently exported from mitochondria as short-chain carnitines (SC-CARs), utilized in anabolic pathways, or released from the cells.MethodsWe examined the specificities of BCAA catabolism and cellular adaptation strategies to BCAA starvation in PDAC cells in vitro. We used metabolomics and lipidomics to quantify major metabolic changes in response to BCAA withdrawal. Using confocal microscopy and flow cytometry we quantified the fluorescence of BODIPY probe and the level of lipid droplets (LDs). We used BODIPY-conjugated palmitate to evaluate transport of fatty acids (FAs) into mitochondria. Also, we have developed a protocol for quantification of SC-CARs, BCAA-derived metabolites.ResultsUsing metabolic profiling, we found that BCAA starvation leads to massive triglyceride (TG) synthesis and LD accumulation. This was associated with the suppression of activated FA transport into the mitochondrial matrix. The suppression of FA import into mitochondria was rescued with the inhibitor of the acetyl-CoA carboxylase (ACC) and the activator of AMP kinase (AMPK), which both regulate carnitine palmitoyltransferase 1A (CPT1) activation status.ConclusionsOur data suggest that BCAA catabolism is required for the import of long chain carnitines (LC-CARs) into mitochondria, whereas the disruption of this link results in the redirection of activated FAs into TG synthesis and its deposition into LDs. We propose that this mechanism protects cells against mitochondrial overload with LC-CARs and it might be part of the universal reaction to amino acid perturbations during cancer growth, regulating FA handling and storage.

Abstract 1796: Role of FASN in pancreatic neuroendocrine cancer cells survival

Abstract Introduction. Pancreatic neuroendocrine tumors (pNETs) are among the most frequently occurring neuroendocrine neoplasms. Up-regulation of fatty acid synthase (FASN) has been reported in many cancers, but the role of lipid metabolism in pNETs remains unclear. The purpose of this study was to determine whether FASN inhibition can be used as a target for pNET treatment. Methods. Immunohistochemistry and western blot analysis were used to determine protein levels of FASN in pNET tissues and cell lines. Expression of FASN was evaluated in pancreatic neuroendocrine primary and metastatic tumors and scored by a pathologist. Neutral lipid droplet levels in pNET cell lines were determined with BODIPY 493/503 staining. Clonogenic and SRB assays were used to evaluate cytotoxic effects of FASN inhibition by TVB-3664 in QGP-1 and BON pancreatic NET cell lines. Western blot analysis was used to determine levels of cleaved PARP and cyclin D1 after FASN inhibition. FASN knockdown was used to determine the effects of FASN downregulation on proliferation and lipid synthesis. Results. FASN expression was determined at the maximum scores of 6 and 5 in 98% of patient samples. BON, QGP-1 and NT-3 cell lines were stained with BODIPY 493/503; the highest levels of neutral lipid droplets were observed in the BON cell line. BON and QGP-1 cells were treated with TVB-3664 in either complete media with fatty acids (FA+) or complete media prepared with fatty acid depleted FBS (FA-). Treatment with TVB-3664 resulted in significant inhibition of BON and QGP-1 cell proliferation and cyclin D1 expression in both FA+ and FA- conditions. Depletion of extracellular FA enhanced the effect of TVB-3664 therapy on NET cells, especially in the BON cell line. FASN inhibition with TVB-3634 suppressed pNET cell proliferation and colony formation. Conclusions. Our study identified high protein levels of FASN in pNET patient samples and cell lines. Moreover, we demonstrated variability in the response of pNET cells to FASN inhibition. QGP cell viability was significantly decreased with FASN inhibition even in the presence of extracellular lipids. FASN inhibition in BON cells had only a minimal effect on cell viability and required depletion of extracellular lipids to achieve a decrease in cell viability and cyclin D1 expression. These data demonstrate that de novo lipid synthesis is involved in the pathogenesis and progression of pNETs. Importantly, our findings show that some pNETs are highly sensitive to FASN inhibition, suggesting that FASN inhibition may be considered as a treatment option in selected pNETs. Citation Format: Benjamin M. Evers, Zeta Chow, Yekaterina Zaytseva, Eun Y. Lee, Courtney M. Townsend, Tianyan Gao, B. Mark Evers, Piotr G. Rychahou. Role of FASN in pancreatic neuroendocrine cancer cells survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1796.

Enhanced lipid biosynthesis in oral squamous cell carcinoma cancer-associated fibroblasts contributes to tumor progression: Role of IL8/AKT/p-ACLY axis.

Lipid metabolic reprogramming of tumor cells has been proven to play a critical role in tumor initiation and development. However, lipid metabolism in cancer-associated fibroblasts (CAFs) has rarely been studied, particularly in CAFs of oral squamous cell carcinoma (OSCC). Additionally, the molecular mechanism by which tumor cells regulate lipid metabolism in fibroblasts is unclear. In this study, we found that phosphorylated ATP citrate lyase (p-ACLY), a key lipid metabolic enzyme, was upregulated in OSCC CAFs. Compared to paracancerous normal fibroblasts, CAFs showed enhanced lipid synthesis, such as elevated cytosolic acetyl-CoA level and accumulation of lipid droplets. Conversely, reduction of p-ACLY level blocked this biological process. In addition, blocking lipid synthesis in CAFs or inhibiting fatty acid uptake by OSCC cells reduced the promotive effects of CAFs on OSCC cell proliferation, invasion, and migration. These findings suggested that CAFs are one of lipid sources required for OSCC progression. Mechanistically, AKT signaling activation was involved in the upregulation of p-ACLY level and lipid synthesis in CAFs. Interleukin-8 (IL8), an exocrine cytokine of OSCC cells, could activate AKT and then phosphorylate ACLY in fibroblasts. This study suggested that the IL8/AKT/p-ACLY axis could be considered as a potential target for OSCC treatment.

The Effect of Thymoquinone on the Protein Levels of PLA2G7, UCP2, and NEDD4L Genes Associated with Lipid Droplets Formation in Prostate Cancer

Objective: Prostate cancer (PCa) patients suffer severe side effects of standard treatment beside the resistance to castration. PCa cells shows increased lipogenesis. Thymoquinone (TQ) inhibits cell proliferation, metastasis, and invasion. However, there was no study on the effect of TQ on the levels of NEDDL4, PLA2G7, and UCP2 lipid droplets (LD) related proteins. Hence, the study aims to investigate the impact of TQ on PLA2G7, UCP2, and NEDD4L proteins on DU145 and PC3 cell lines.&#x0D; Materials and Methods: Cells were cultured and treated with TQ with a IC50 of 60 µM and 80 µM for DU145 and PC3, respectively. PLA2G7, UCP2, and NEDD4L levels were measured using the ELISA.&#x0D; Results: TQ has significantly increased the level of NEDD4L (p

Protein-encapsulated long-wavelength fluorescent probe hybrid for imaging lipid droplets in living cells and mice with non-alcoholic fatty liver

Non-alcoholic fatty liver disease (NAFLD) is prevalent worldwide as a chronic liver disease that not only gives rise to hepatic complications, but leads to other chronic diseases such as type 2 diabetes and atherosclerosis. The aberrant accumulation of lipid droplets (LDs) in hepatocytes is a prominent signature of NAFLD. However, conventional techniques lack the capability to effectively monitor the dynamic changes in LD levels during NAFLD with living organisms. Hence, it is imperative to develop LD-specific long-wavelength fluorescent probes with high imaging contrast for the in-situ diagnosis of NAFLD. In this study, we synthesized a new LD-selective long-wavelength fluorescent probe, denoted as LD-1, based on the twisted intramolecular charge transfer (TICT) mechanism. The probe exhibits a large Stokes shift and intensive fluorescence emission in nonpolar and viscous solutions. By self-assembling LD-1 with bovine serum albumin (BSA), a biocompatible, long-wavelength fluorescent probe hybrid, LD-1@BSA, was formed, allowing for LDs to be selectively imaged in hepatocytes. Moreover, LD-1@BSA successfully discriminates NAFLD cells before and after drug treatment, and achieves non-invasive and real-time monitoring of LD accumulation in a mouse model of NAFLD.

Smart and low-cost fluorometer for identifying breast cancer malignancy based on lipid droplets accumulation.

The most common cause of breast cancer-related death is tumor recurrence. To develop more effective treatments, the identification of cancer cell specific malignancy indicators is therefore critical. Lipid droplets are known as an emerging hallmark in aggressive breast tumors. A common technique that can be used for observing molecules in cancer microenvironment is fluorescence microscopy. We describe the design, development and applicability of a smart fluorometer to detect lipid droplet accumulation based on the emitted fluorescence signals from highly malignant (MDA-MB-231) and mildly malignant (MCF7) breast cancer cell lines, that are stained with BODIPY dye. This device uses a visible-range light source as an excitation source and a spectral sensor as the detector. A commercial imaging system was used to examine the fluorescent cancer cell lines before being validated in a preclinical setting with the developed prototype. The outcomes indicate that this low-cost fluorometer can effectively detect the alterations levels of lipid droplets and hence distinguish between "moderately malignant" and "highly malignant" cancer cells. In comparison to prior research that used fluorescence spectroscopy techniques to detect cancer biomarkers, this study revealed enhanced capability in classifying mildly and highly malignant cancer cell lines.

TGF-β1 Decreases Microglia-Mediated Neuroinflammation and Lipid Droplet Accumulation in an In Vitro Stroke Model.

Hypoxia triggers reactive microglial inflammation and lipid droplet (LD) accumulation under stroke conditions, although the mutual interactions between these two processes are insufficiently understood. Hence, the involvement of transforming growth factor (TGF)-β1 in inflammation and LD accumulation in cultured microglia exposed to hypoxia were analyzed herein. Primary microglia were exposed to oxygen-glucose deprivation (OGD) injury and lipopolysaccharide (LPS) stimulation. For analyzing the role of TGF-β1 patterns under such conditions, a TGF-β1 siRNA and an exogenous recombinant TGF-β1 protein were employed. Further studies applied Triacsin C, an inhibitor of LD formation, in order to directly assess the impact of LD formation on the modulation of inflammation. To assess mutual microglia-to-neuron interactions, a co-culture model of these cells was established. Upon OGD exposure, microglial TGF-β1 levels were significantly increased, whereas LPS stimulation yielded decreased levels. Elevating TGF-β1 expression proved highly effective in suppressing inflammation and reducing LD accumulation in microglia exposed to LPS. Conversely, inhibition of TGF-β1 led to the promotion of microglial cell inflammation and an increase in LD accumulation in microglia exposed to OGD. Employing the LD formation inhibitor Triacsin C, in turn, polarized microglia towards an anti-inflammatory phenotype. Such modulation of both microglial TGF-β1 and LD levels significantly affected the resistance of co-cultured neurons. This study provides novel insights by demonstrating that TGF-β1 plays a protective role against microglia-mediated neuroinflammation through the suppression of LD accumulation. These findings offer a fresh perspective on stroke treatment, suggesting the potential of targeting this pathway for therapeutic interventions.

Glyceryl triacetate promotes blood–brain barrier recovery after ischemic stroke through lipogenesis-mediated IL-33 in mice

BackgroundLipid metabolism has a crucial role in neural repair in neurodegenerative diseases. We recently revealed that lipogenesis-mediated interleukin-33 (IL-33) upregulation lead to blood–brain barrier (BBB) repair after ischemic stroke. However, manipulating the key enzyme fatty acid synthase (FASN) to enhance lipogenesis was very challenging. Glyceryl triacetate (GTA) was used as a donor of acetate and precursor of acetyl coenzyme A, the key substrate for de novo lipogenesis catalyzed by FASN. Therefore, we hypothesized that GTA would promote lipogenesis the peri-infarct after ischemic stroke and contribute to the BBB repair through IL-33.MethodsMiddle cerebral artery occlusion (MCAO) was performed on C57BL mice and GTA was gavage administrated (4 g/kg) on day 2 and 4 after MCAO. Lipogenesis was evaluated by assessment of the protein level of FASN, lipid droplets, and fatty acid products through liquid chromatography-mass spectrometry in the peri-infarct area on day 3 after MCAO, respectively. BBB permeability was determined by extravasation of Evans blue, IgG and dextran, and levels of tight junction proteins in the peri-infarct area on day 7 after MCAO, respectively. Infarct size and neurological defects were assessed on day 7 after MCAO. Brain atrophy on day 30 and long-term sensorimotor abilities after MCAO were analyzed as well. The inhibitor of FASN, C75 and the virus-delivered FASN shRNA were used to evaluate the role of FASN-driven lipogenesis in GTA-improved BBB repair. Finally, the therapeutic potential of recombinant IL-33 on BBB repair and neurological recovery was evaluated.ResultsWe found that treatment with GTA increased the lipogenesis as evidenced by lipid droplets level and lauric acid content, but not the FASN protein level. Treatment with GTA increased the IL-33 level in the peri-infarct area and decreased the BBB permeability after MCAO. However, infarct size and neurological defect score were unchanged on day 7 after MCAO, while the long-term recovery of sensorimotor function and brain atrophy were improved by GTA. Inhibition of lipogenesis using C75 or FASN shRNA reversed the beneficial effect of GTA. Finally, exogenous IL-33 improved BBB repair and long-term functional recovery after stroke.ConclusionCollectively, we concluded that treatment with GTA improved the BBB repair and functional recovery after ischemic stroke, probably by the enhancement of lipogenesis and IL-33 expression.Graphical

Preventing Adipogenesis and Preserving Mitochondria and GLUT-4 Functions by Extracts and Isolated Compounds of Australian Acacia saligna.

Acacia saligna's secondary metabolites show promise in treating type 2 diabetes mellitus and its related conditions. We previously discovered that methanolic extracts, isolated flavonoids, and cyclitols effectively preserve mitochondria in 3T3-L1 adipocytes. In this current work, quantification of lipid droplet levels with Oil Red O assay showed a noticeable decrease in lipogenesis in 3T3-L1 cells. Methanolic leaf and bark extracts and isolated compounds, (-)-epicatechin 6 and myricitrin 8, reduced cellular lipid levels by 21.15% to 25.28%, respectively. mRNA levels of key regulators of mitochondrial biogenesis, such as adiponectin, PGC-1α, and mtTFA, were increased. Methanolic flower extract (FL-MeOH) and its chemical components, naringenin 1 and D-(+)-pinitol 5a, increased these gene levels from 10% to 29% at the higher dose. Our study found that FL-MeOH slightly reduced pro-inflammatory cytokines TNF-α and IL-6, attributed to two phytochemicals, naringenin-7-O-α-L-arabinofuranoside 2 and D-(+)-pinitol 5a. Western blot analysis also showed that adipocytes treated with MeOH extracts had higher GLUT-4 expression levels than untreated adipocytes. Overall, A. saligna extracts and their isolated compounds demonstrated anti-lipogenesis activity during 3T3-L1 cell differentiation, modulation of transcriptional levels of adiponectin, PGC-1α, and mtTFA, reducing TNF-α and IL-6 mRNA levels, promoting mitochondrial biogenesis, and enhancing GLUT-4 expression.

Lipid droplets modulate proteostasis, SQST-1/SQSTM1 dynamics, and lifespan in C. elegans

In several long-lived Caenorhabditis elegans strains, such as insulin/IGF-1 receptor daf-2 mutants, enhanced proteostatic mechanisms are accompanied by elevated intestinal lipid stores, but their role in longevity is unclear. Here, while determining the regulatory network of the selective autophagy receptorSQST-1/SQSTM1, we uncovered an important role for lipid droplets in proteostasis and longevity. Using genome-wide RNAi screening, we identified several SQST-1 modulators, including lipid droplets-associated and aggregation-prone proteins. Expansion of intestinal lipid droplets by silencing the conserved cytosolic triacylglycerol lipase gene atgl-1/ATGL enhanced autophagy, and extended lifespan. Notably, a substantial amount of ubiquitinated proteins were found on lipid droplets. Reducing lipiddroplet levels exacerbated the proteostatic collapse when autophagy or proteasome function wascompromised, and significantly reduced the lifespan of long-lived daf-2 animals. Altogether, our study uncovered a key role for lipid droplets in C.elegans as a proteostatic mediator that modulates ubiquitinated protein accumulation, facilitates autophagy, and promotes longevity.

Investigations of Lipid Droplets Role in Attenuating Chemotherapeutic Responses to 5-FU in Cancer Cells in vitro

Cancer has been considered as a main cause of death worldwide. Despite the effectiveness of traditional anticancer therapies such as 5-Flurouracil (5-FU), poor therapeutic outcomes are reported in many cases, due to tumor recurrence and chemoresistance. Intracellular lipid accumulation as lipid droplets (LDs) is now a well-recognized hallmark of cancer. However, the influence of LDs accumulation in cancer progression and treatment remains to be elucidated. Adjuvant use of non-chemotherapeutic drugs (e.g. NSAIDs and corticosteroids) with anticancer drugs to manage cancer related symptoms, may though serve as factors modulating the therapeutic response to anticancer agents via LDs related mechanisms. The aim of this study is to evaluate possible strategies influencing chemoresistance by attenuating LDs biogenesis and function. LDs levels in eight human cancer cell lines were measured. The existence of correlation between the cellular levels of LDs and cytotoxicity of ten chemotherapeutic agents was evaluated. A moderate correlation between basal LDs levels and the half inhibitory concentration (IC50) values of 5-FU on selected cell lines was established (r2= 0.5235). Nevertheless, LDs levels were significantly elevated following exposure to 5-FU. A549 human lung cancer cells showed the highest increase in LD accumulation (***P &gt;0.001) compared to their basal levels of LDs. LDs levels were also assessed following exposure to 5-FU in the presence of sub-lethal doses of celecoxib (CXB), dexamethasone (DEX), and simvastatin (SMV). Interestingly, CXB and DEX exposure to 5-FUtreated cells resulted in an alleviation in the antiproliferative activities of 5-FU in MDA-MB-468 and HCT116 but not in A549 cells. While, SMV exposure to 5-FU-treated cells resulted in reduced antiproliferative activities of 5-FU in MDA-MB-468 only. These results strongly suggest that increased LDs levels caused by CXB, DEX, and SMV may contribute in the development of a resistance mechanism exist only in some cancer types, which therefore, attenuates responses to 5- FU. The inhibition of phospholipid metabolism by DEX, as well as the inhibition of 3-hydroxy-3- methylglutaryl-CoA reductase (HMGCR) by SMV were expected to result in the up regulation of triacylglycerol (TAG), leading to LDs accumulation. This study highlights the importance of assessing drug-drug interaction before designing integrated therapy regimens for cancer patients receiving 5-FU treatment. Further studies are needed to discover the role of TAG inhibition on the sensitivity of cancer cells against 5-FU.

Multifaceted Nature of Lipid Droplets in Viral Interactions and Pathogenesis.

Once regarded as inert organelles with limited and ill-defined roles, lipid droplets (LDs) have emerged as dynamic entities with multifaceted functions within the cell. Recent research has illuminated their pivotal role as primary energy reservoirs in the form of lipids, capable of being metabolized to meet cellular energy demands. Their high dynamism is underscored by their ability to interact with numerous cellular organelles, notably the endoplasmic reticulum (the site of LD genesis) and mitochondria, which utilize small LDs for energy production. Beyond their contribution to cellular bioenergetics, LDs have been associated with viral infections. Evidence suggests that viruses can co-opt LDs to facilitate their infection cycle. Furthermore, recent discoveries highlight the role of LDs in modulating the host's immune response. Observations of altered LD levels during viral infections suggest their involvement in disease pathophysiology, potentially through production of proinflammatory mediators using LD lipids as precursors. This review explores these intriguing aspects of LDs, shedding light on their multifaceted nature and implications in viral interactions and disease development.