Lipid Droplet Formation Research Articles

Seipin governs phosphatidic acid homeostasis at the inner nuclear membrane

The nuclear envelope is a specialized subdomain of the endoplasmic reticulum and comprises the inner and outer nuclear membranes. Despite the crucial role of the inner nuclear membrane in genome regulation, its lipid metabolism remains poorly understood. Phosphatidic acid (PA) is essential for membrane growth as well as lipid storage. Using a genome-wide lipid biosensor screen in S. cerevisiae, we identify regulators of inner nuclear membrane PA homeostasis, including yeast Seipin, a known mediator of nuclear lipid droplet biogenesis. Here, we show that Seipin preserves nuclear envelope integrity by preventing its deformation and ectopic membrane formation. Mutations of specific regions of Seipin, some linked to human lipodystrophy, disrupt PA distribution at the inner nuclear membrane and nuclear lipid droplet formation. Investigating the Seipin co-factor Ldb16 reveals that a triacylglycerol binding site is crucial for lipid droplet formation, whereas PA regulation can be functionally separated. Our study highlights the potential of lipid biosensor screens for examining inner nuclear membrane lipid metabolism.

A Single-Cell Atlas of Porcine Skeletal Muscle Reveals Mechanisms That Regulate Intramuscular Adipogenesis

Porcine skeletal muscle development is closely linked to meat production efficiency and quality. The accumulation of porcine intramuscular fat is influenced by the hyperplasia and hypertrophy of adipocytes within the muscle. However, the cellular profiles corresponding to the two stages of muscle development remain undetermined. Single-nucleus RNA sequencing (snRNA-seq) can elucidate cell subsets in tissues, capture gene expression at the individual cell level, and provide innovative perspectives for studying muscle and intramuscular fat formation. In this study, a total of 78,302 nuclei and 9 clusters of cells, which included fibro/adipogenic progenitor (FAP), myonuclei, adipocytes, and other cell types, of Xidu black pigs, were identified on Day 1 and Day 180. The pattern of cell clustering varied between the two developmental stages. Notably, the percentage of adipocytes in the Day 180 group was higher than in the Day 1 group (0.51% vs. 0.15%). Pseudo-time sequence analysis indicated that FAPs could differentiate into adipocytes and myonuclei cells, respectively. The THRSP gene was identified as a biomarker for swine intramuscular fat cells, and its down-regulation resulted in significant reduction in lipid droplet formation in porcine preadipocytes. Our research provides new insights into the cellular characteristics of intramuscular fat formation, which may facilitate the development of novel strategies to enhance intramuscular fat deposition and improve pork quality.

A PLIN1 polymorphism is associated with fat production in male emus

The emu (Dromaius novaehollandiae) is a novel poultry species that produces meat, eggs, and fat. Although emus have recently been domesticated, genetic improvements to establish strains have scarcely progressed. In this study, we investigated the relationship between production traits and perilipin 1-encoding gene (PLIN1) polymorphisms in the emus. We determined the partial complementary DNA (cDNA) sequence of the PLIN1, which is involved in lipid droplet formation. We identified four nucleotide substitution sites (c.270C>T, c.321T>C, c.587A>T, and c.639C>T) in the PLIN1 gene of emus. Of these, c.587A>T is a non-synonymous substitution that converts lysine to methionine at the 196th codon (p.K196M). Although p.K196M was predicted to affect the production traits of emus, a large deflection in genotype frequency was observed in this study; thus, we could not investigate the relationship between genotypes and phenotypes. In males, the fat yields of the CC, CT, and TT genotypes in c.270C>T were 0.25 ± 0.06, 0.22 ± 0.06, and 0.21 ± 0.07 kg, respectively, while the meat yields of the CC, CT, and TT genotypes in c.270C>T were 0.15 ± 0.01, 0.16 ± 0.02, and 0.16 ± 0.03 kg, respectively. These results indicate that male emus with the CC genotype had a significantly higher fat content and lower meat productivity than male emus with the other genotypes (P < 0.05). Therefore, c.270C>T in PLIN1 affects fat and meat production in males. Our findings may contribute to the effective genetic improvement of the emus.

Gold Kiwi-Derived Nanovesicles Mitigate Ultraviolet-Induced Photoaging and Enhance Osteogenic Differentiation in Bone Marrow Mesenchymal Stem Cells

Bone marrow mesenchymal stem cells (BM-MSCs) play a crucial role in bone formation through their ability to differentiate into osteoblasts. Aging, however, detrimentally affects the differentiation and proliferation capacities of BM-MSCs, consequently impairing bone regeneration. Thus, mitigating the aging effects on BM-MSCs is vital for addressing bone-related pathologies. In this study, we demonstrate that extracellular nanovesicles isolated from gold kiwi (GK-NVs) protect human BM-MSCs from ultraviolet (UV)-induced photoaging, thereby alleviating aging-related impairments in cellular functions that are crucial for bone homeostasis. Notably, GK-NVs were efficiently taken up by BM-MSCs without causing cytotoxicity. GK-NVs reduced intracellular reactive oxygen species (ROS) levels upon UV irradiation, restoring impaired proliferation and migration capabilities. Furthermore, GK-NVs corrected the skewed differentiation capacities of UV-irradiated BM-MSCs by enhancing osteoblast differentiation, as evidenced by the increased expression in osteoblast-specific genes and the calcium deposition, and by reducing adipocyte differentiation, as indicated by the decreased lipid droplet formation. These findings position GK-NVs as a promising biomaterial for the treatment of bone-related diseases such as osteoporosis.

Diosgenin improves lipid metabolism in diabetic nephropathy via regulation of miR-148b-3p/DNMT1/FOXO1 axis.

The progression of diabetic nephropathy (DN) is closely associated with lipid accumulation. Diosgenin (Dio) plays a beneficial role in the lipid metabolism associated with multiple diseases. Thus, the mechanism underlying Dio's function in DN associated with aberrant lipid accumulation warrants further investigation. To model DN in vitro, HK-2 cells were treated with high glucose (HG) and palmitic acid. Cell viability was evaluated using MTT assay. The triglyceride (TG) content in HK-2 cells was measured using a commercial assay kit. The formation of lipid droplets in HK-2 cells was observed using Oil Red O staining. The expression levels of mRNA and protein were detected using RT-qPCR and western blot, respectively. The DNA methylation of FOXO1 was assessed using MSP. The interaction between DNMT1 and the FOXO1 promoter was confirmed by ChIP assay. Dio treatment reduced TG levels and lipid droplet formation in HK-2 cells co-treated with HG and palmitic acid. Simultaneously, the levels of miR-148b-3p and FOXO1 were increased by Dio, while Dio decreased the expression levels of DNMT1 and SREBP-2. Meanwhile, miR-148b-3p can bind to DNMT1, which in turn inhibits the expression of FOXO1 by mediating the DNA methylation of FOXO1. In addition, FOXO1 negatively regulates the expression of SREBP-2 by interacting with the SREBP-2 promoter. MiR-148b-3p inhibition or silencing of FOXO1 abolished the inhibitory effect of Dio on TG production and lipid droplet formation. This effect was further exacerbated by the downregulation of DNMT1. FOXO1 overexpression may counteract the promotive effects of miR-148b-3p inhibitor on lipid accumulation. Dio treatment reduced TG production and lipid droplet formation in HK-2 cells during the progression of DN by modulating the miR-148b-3p/DNMT1/FOXO1/SREBP-2 axis. This finding provides new evidence supporting the therapeutic potential of Dio for DN.

Loss of peroxiredoxin 6 (PRDX6) alters lipid composition and distribution resulting in increased sensitivity to ferroptosis.

Peroxiredoxin 6 (PRDX6) is a multifunctional enzyme involved in phospholipid peroxide repair and metabolism. In this study we investigated the global lipid composition of a human hepatocarcinoma cell line SNU475 lacking PRDX6 and lipid related cellular processes. There was a general decrease in multiple lipids species upon loss of PRDX6, in particular sphingomyelins and acylcarnitines, consistent with previously observed alterations in cell signaling pathways and mitochondrial dysfunction. Deprivation of docosahexaenoic acid and related species was also evident. However, a few striking exceptions are worth highlighting: 1) Three specific arachidonic acid (AA) containing phophatidylcholines (PC) increased significantly. The increase of sn1-stearic/sn2-PUFA containing PC and sn2-AA containing plasmenyls are indicative of a preference of PRDX6 iPLA2 activity for these AA storage glycerophospholipids. 2) Several polyunsaturated fatty acids (PUFA) and PUFA containing triacylglycerols accumulated together with increased formation of lipid droplets, an indication of altered FA flux and PUFA sequestration in PRDX6 knockout cells. Loss of PRDX6 resulted in increased sensitivity to erastin-induced ferroptosis, independent of selenium and GPX4, as a consequence of increased levels of lipid hydroperoxides, that reverted to normal levels upon rescue with PRDX6. <SPAN style="font-weight: 400;">The results presented demonstrate that all three enzymatic activities of PRDX6 contribute to the role of this multifunctional enzyme in diverse cellular processes, including membrane phospholipid remodeling and glycerophospholipid functional diversity, resulting in altered lipid peroxides and modulation of AA disposition and traffic. These contributions highlight the complexity of the changes that loss of PRDX6 exerts on cell functionality.</SPAN>.

Uncovering the protective role of lipid droplet accumulation against acid-induced oxidative stress and cell death in osteosarcoma

Extracellular acidosis stemming from altered tumor metabolism promotes cancer progression by enabling tumor cell adaptation to the hostile microenvironment. In osteosarcoma, we have previously shown that acidosis increases tumor cell survival alongside substantial lipid droplet accumulation. In this study, we explored the role of lipid droplet formation in mitigating cellular stress induced by extracellular acidosis in osteosarcoma cells, thereby enhancing tumor survival during progression. Specifically, we examined how lipid droplets shield against reactive oxygen species induced by extracellular acidosis. We demonstrated that lipid droplet biogenesis is critical for acid-exposed tumor cell survival, as it starts shortly after acid exposure (24 h) and inversely correlates with ROS levels (DCFH-DA assay), lipid peroxidation (Bodipy assay), and the antioxidant response, as also revealed by NRF2 transcript. Additionally, extracellular metabolites, such as lactate, and interaction with mesenchymal stromal cells within the tumor microenvironment intensify lipid droplet build-up in osteosarcoma cells. Critically, upon targeting two key proteins implicated in LD formation - PLIN2 and DGAT1 - cell viability significantly declined while ROS production escalated. In summary, our findings underscore the vital reliance of acid-exposed tumor cells on lipid droplet formation to scavenge oxidative stress. We conclude that the rewiring of lipid metabolism driven by microenvironmental cues is of paramount importance for the survival of metabolically altered osteosarcoma cells in acidic condition. Overall, we suggest that targeting key members of lipid droplet biogenesis may eradicate more aggressive and resistant tumor cells, uncovering potential new treatment strategies for osteosarcoma.

Inhibiting lipid droplet biogenesis enhances host protection against hypervirulent Klebsiella pneumoniae infections

Hypervirulent Klebsiella pneumoniae (hvKp), an emerging Kp subtype, has become a serious global pathogen. However, the information regarding host interactions and innate immune responses during hvKp infection is limited. Here, we found that hvKp clinical strains increased triacylglycerol synthesis, resulting in lipid droplets (LDs) formation via the mammalian target of rapamycin signaling pathway in RAW264.7 cells. Treatment with rapamycin, an inhibitor of this pathway, affected LDs formation and antimicrobial responses against clinical hvKp infections. In accordance with the role of LDs in modulating inflammation, the pharmacological inhibition of lipogenesis reduced proinflammatory cytokine expression during hvKp infections. In addition, inhibition of LDs formation using pharmacological inhibitors and knockdown of lipogenesis regulators decreased the intracellular survival of hvKp in macrophages. Moreover, inhibiting LDs biogenesis reduced mortality, weight loss, and bacterial loads in hvKp-infected mice. Collectively, these data suggest that LDs biogenesis is crucial in linking host immune responses to clinical hvKp infections.

TMET-07. RADIORESISTANT GLIOBLASTOMA EVADES MITOCHONDRIAL LIPOTOXICITY THROUGH ALTERED DGKB AND DGAT1

Abstract Glioblastoma (GBM) is the most common and aggressive primary brain tumor, with a median survival of only 15 months despite multimodal therapy. Radiotherapy is a conventional therapy of GBM treatment, but the development of radioresistance often leads to tumor recurrence and treatment failure. To elucidate the molecular mechanisms underlying radioresistance in GBM, we established radioresistant GBM cell lines through repeated exposure to ionizing radiation. Transcriptomic analysis revealed that diacylglycerol kinase beta (DGKB), an enzyme that converts diacylglycerol (DAG) to phosphatidic acid, was significantly downregulated in radioresistant GBM cells. Functional studies demonstrated that knockdown of DGKB or expression of a kinase-dead DGKB mutant resulted in DAG accumulation and reduced fatty acid oxidation. Conversely, overexpression of DGKB activated fatty acid oxidation, increased reactive oxygen species (ROS) production, and sensitized GBM cells to radiation. Interestingly, we found that radiation upregulated the expression of diacylglycerol acyltransferase 1 (DGAT1), which catalyzes the conversion of DAG to triglycerides. Knockdown of DGAT1 using shRNA or miR-3918 mimic reduced lipid droplet formation and enhanced radiosensitivity both in vitro and in vivo. Furthermore, using Connectivity Map, we identified cladribine as a potential radiosensitizing agent that increases DGKB and decreases DGAT1. Cladribine effectively sensitized GBM cells to radiation and significantly prolonged the survival of mice bearing GBM xenografts. In conclusion, our study reveals a novel mechanism by which radioresistant GBM cells maintain lipid homeostasis through downregulation of DGKB and upregulation of DGAT1 to evade lethal mitochondrial lipotoxicity. Targeting the DGKB-DGAT1 axis represents a promising therapeutic strategy to overcome radioresistance and improve the outcomes of GBM patients.

Lipid Droplet Biology in Obesity: Mechanisms of Lipid Storage and Mobilization in Metabolic Disease

Lipid droplets (LDs) are dynamic organelles that play a central role in lipid storage and mobilization, particularly in the context of metabolic diseases like obesity. Obesity is characterized by excessive accumulation of triglycerides within adipocytes, leading to metabolic dysregulation and associated comorbidities such as insulin resistance, type 2 diabetes, and cardiovascular disease. This review explores the biology of lipid droplets in the setting of obesity, focusing on the mechanisms of lipid storage and mobilization. We discuss the key regulatory proteins involved in LD formation and degradation, including perilipins, adipose triglyceride lipase (ATGL), and hormone-sensitive lipase (HSL). Additionally, the roles of lipophagy, autophagy, and lipolysis in lipid mobilization are explored. We also highlight how dysregulation of lipid droplet biology contributes to obesity-related metabolic disorders, with emphasis on the impact of lipid droplet dysfunction on cellular signaling, lipid metabolism, and inflammation. Finally, we discuss the potential of targeting LDs as a therapeutic strategy for treating obesity and its metabolic complications. Keywords: Lipid droplets, Obesity, Lipid storage, Lipid mobilization, Metabolic disease, Lipolysis, Lipophagy, Triglycerides

FABP4-mediated lipid accumulation and lipolysis in tumor-associated macrophages promote breast cancer metastasis.

A high density of tumor-associated macrophages (TAMs) is associated with poorer prognosis and survival in breast cancer patients. Recent studies have shown that lipid accumulation in TAMs can promote tumor growth and metastasis in various models. However, the specific molecular mechanisms that drive lipid accumulation and tumor progression in TAMs remain largely unknown. Herein, we demonstrated that unsaturated fatty acids (FAs), unlike saturated ones, are more likely to form lipid droplets in murine macrophages. Specifically, unsaturated FAs, including linoleic acids (LA), activate the FABP4/CEBPα pathway, leading to triglyceride synthesis and lipid droplet formation. Furthermore, FABP4 enhances lipolysis and FA utilization by breast cancer cell lines, which promotes cancer cell migration in vitro and metastasis in vivo. Notably, a deficiency of FABP4 in murine macrophages significantly reduces LA-induced lipid metabolism. Therefore, our findings suggest FABP4 as a crucial lipid messenger that facilitates unsaturated FA-mediated lipid accumulation and lipolysis in TAMs, thus contributing to the metastasis of breast cancer.

FABP4-mediated lipid accumulation and lipolysis in tumor-associated macrophages promote breast cancer metastasis

A high density of tumor-associated macrophages (TAMs) is associated with poorer prognosis and survival in breast cancer patients. Recent studies have shown that lipid accumulation in TAMs can promote tumor growth and metastasis in various models. However, the specific molecular mechanisms that drive lipid accumulation and tumor progression in TAMs remain largely unknown. Herein, we demonstrated that unsaturated fatty acids (FAs), unlike saturated ones, are more likely to form lipid droplets in murine macrophages. Specifically, unsaturated FAs, including linoleic acids (LA), activate the FABP4/CEBPα pathway, leading to triglyceride synthesis and lipid droplet formation. Furthermore, FABP4 enhances lipolysis and FA utilization by breast cancer cell lines, which promotes cancer cell migration in vitro and metastasis in vivo. Notably, a deficiency of FABP4 in murine macrophages significantly reduces LA-induced lipid metabolism. Therefore, our findings suggest FABP4 as a crucial lipid messenger that facilitates unsaturated FA-mediated lipid accumulation and lipolysis in TAMs, thus contributing to the metastasis of breast cancer.

The α/β hydrolase domain-containing protein 1 (ABHD1) acts as a lysolipid lipase and is involved in lipid droplet formation

Abstract Lipid droplets (LDs) are the major sites of lipid and energy homeostasis. However, few LD biogenesis proteins have been identified. Using model microalga Chlamydomonas, we show that ABHD1, an α/β-hydrolase domain-containing protein, is localized to the LD surface and stimulates LD formation through two actions: one enzymatic and one structural. The knockout mutants contained similar amounts of triacylglycerols (TAG) but their LDs showed a higher content of lyso-derivatives of betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS). Over-expression of ABHD1 increased LD abundance and boosted TAG content. Purified recombinant ABHD1 hydrolyzed lyso-DGTS, producing a free fatty acid and a glyceryltrimethylhomoserine. In vitro droplet-embedded vesicles showed ABHD1 promoted LD emergence. Taken together, these results identify ABHD1 as a new player in LD formation by its lipase activity on lyso-DGTS and by its distinct biophysical property. This study further suggests that lipases targeted to LDs and able to act on their polar lipid coat may be interesting tools to promote LD assembly in eukaryotic cells.

Bta-miR-224 regulates milk fat metabolism by targeting FABP4 in bovine mammary epithelial cells

Milk fat is produced and secreted by the mammary gland, which is mainly regulated by diet and gene-molecule network. Therefore, understanding the molecular mechanism of milk fat synthesis is of practical significance for improving milk quality. Fatty acid-binding protein 4 (FABP4) is a candidate messenger RNA (mRNA) closely linked to milk fat metabolism obtained from transcriptomic analysis of mammary epithelial cells of cows in the pre-existing high- and low-milk-fat groups, and its expression pattern and function are still unclear. The qRT-PCR results depicted that FABP4 was highly expressed in bovine mammary epithelial cells (BMECs) in breast tissues and the high milk fat group. Subsequently, the regulatory effects of FABP4 on BMECs were analyzed by CCK8, EdU, and flow cytometry, and the results demonstrated that FABP4 inhibited the proliferation and viability of BMECs and promoted their apoptosis. Next, the effect of FABP4 on milk lipid metabolism was explored. pEGFP-N1-FABP4 was transfected into BMECs, and FABP4 upregulated the expression levels of the milk lipid marker genes XDH, PPARG, and ACSS2, and promoted the formation of triglycerides (TGs), cholesterol, lipid droplets, and β-casein. Strong interactions between FABP4 and PPARG were identified using STRING prediction. Western blotting revealed that FABP4 interacted with PPARG to promote PPARG expression, while the opposite result was observed after interfering with FABP4. The gene regulation of microRNA (miRNA) is essential for fatty acid metabolism and synthesis. Predicted by website and combined with pre-miRNA transcriptome sequencing results, we hypothesized that FABP4 might be the target gene of bta-miR-224. The results of the dual-luciferase reporter gene and qRT-PCR revealed that bta-miR-224 negatively regulated FABP4 expression by targeting the 3′-UTR of FABP4. By exploring the function of bta-miR-224, we observed that bta-miR-224 mimics downregulated the expression of the milk fat marker genes AGPAT6, ACSS2, and XDH and inhibited TG synthesis and lipid droplet secretion. However, the bta-miR-224 inhibitor depicted the opposite results. In conclusion, FABP4 plays a crucial role in regulating BMEC proliferation and differentiation. Bta-miR-224 targeting FABP4 may promote biological processes such as TG synthesis and lipid droplet formation through PPARG, which lays a solid foundation for further analysis of the functional mechanism of milk lipid metabolism in dairy cows from a miRNA-mRNA perspective.

Metallothionein 1B attenuates inflammation and hepatic steatosis in MASH by inhibiting the AKT/PI3K pathway

Metabolic dysfunction-associated steatohepatitis (MASH) is a severe form of metabolic dysfunction-associated fatty liver disease (MAFLD), characterized by hepatic steatosis, inflammation, and fibrosis. This study investigates the role and potential mechanisms of metallothionein 1B (MT1B) in MASH through bioinformatics analysis and experimental validation. qRT-PCR and western blot analyses confirm that MT1B expression is significantly downregulated in liver tissues of MASH patients, in high-fat diet (HFD)-induced mouse models, and in hepatocytes induced by free fatty acids (FFA). Further functional experiments show that upregulation of MT1B reduces intracellular triglycerides and total cholesterol levels, lipid droplet formation, and pro-inflammatory factors. In vivo experiments demonstrate that specific downregulation of hepatic MT1B expression via AAV8-shMT1B injection significantly increases triglyceride and total cholesterol levels, exacerbates lipid accumulation, and markedly elevates liver fibrosis and inflammatory factor expression. RNA-seq and bioinformatics analyses show that the AKT/PI3K pathway is significantly suppressed in MT1B-overexpressing cells. Further experiments indicate that AKT inhibition can reverse the lipid metabolism disorders and inflammatory responses caused by MT1B downregulation. Additionally, Zinc can promote the nuclear translocation of MTF1, leading to its binding to the MT1B promoter, thereby upregulating MT1B expression and ultimately mitigating MASH progression. These findings suggest that zinc-regulated MT1B plays a critical role in lipid metabolism and inflammatory responses by regulating the AKT/PI3K signaling pathway, influencing MASH progression.

Therapeutic role of physalin A in the pathogenesis of Graves’ orbitopathy

Background Graves’ orbitopathy (GO) is an autoimmune condition that causes serious ocular symptoms; its treatment strategies are limited. Physalin A is a phytosterol that has shown various therapeutic properties, including anti-inflammatory and anti-fibrotic effects. In this study, we investigated whether physalin A could inhibit inflammation, fibrosis, hyaluronan (hyaluronic acid) production, and adipogenesis, which are crucial to the pathogenesis of GO. Methods Orbital tissue explants were obtained from patients with GO during orbital decompression surgery and healthy controls. Orbital fibroblasts (OFs) were isolated and treated with different concentrations of physalin A. Using western blot and ELISA analyses, we determined the effects of physalin A on OFs. Results Physalin A treatment suppressed the production of interleukin (IL)-1β-induced prostaglandin E2 (PGE2) and pro-inflammatory molecules, including cyclooxygenase (COX)-2, IL-6, IL-8, and intercellular adhesion molecule (ICAM)-1. We discovered that physalin A attenuated hyaluronan production induced by IL-1β or insulin-like growth factor (IGF)-1. Moreover, physalin A reduced lipid droplet formation and production of peroxisome proliferator activator (PPAR) γ, CCAAT-enhancer-binding protein (C/EBP) α, C/EBP β, sterol regulatory element binding protein (SREBP)-1, leptin, and adiponectin proteins. Physalin A suppressed the phosphorylation of extracellular signal-related kinase (ERK), nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), and suppressor of mothers against decapentaplegic (SMAD) 2 signaling protein. Conclusions Our study suggests that the major mechanisms by which physalin A suppresses GO include reducing inflammation, fibrosis, hyaluronan production, and adipogenesis in OFs. The findings of this study provide evidence of the therapeutic effect of physalin A in GO.

Metabolic checkpoints in rheumatoid arthritis

BackgroundRheumatoid Arthritis is a systemic autoimmune disease affecting 0.5–1 % of the population. Despite a growing therapeutic armamentarium, RA remains incurable, and many patients suffer significant morbidity over time. The strongest genetic risk derives from HLA class II polymorphisms, implicating T cells as pathogenic drivers. Innate immune cells, e.g. monocytes and macrophages (Mⱷ) contribute to chronic tissue inflammation through an array of pro-inflammatory functions but also present antigen to autoreactive T cells. Differentiation, survival, and effector functions of both T cells and Mⱷ are ultimately controlled by their bioenergetic and biosynthetic programs, identifying cellular metabolism as a critical disease mechanism in RA. ObjectivesSummarize current knowledge about metabolic conditions in the RA joint and disease-relevant metabolic circuits shaping the effector repertoire of RA T cells and Mⱷ. ResultsThe rheumatoid joint is a glucose deplete tissue environment, selecting for invading immune cells that can survive on non-glucose fuel sources. Inflamed synovium instead offers the amino acid glutamine and RA CD4+T cells and RA Mⱷ rely on glutamine and glutamate to support their pathogenic functions. The metabolic hallmark of RA T cells is their low mitochondrial performance, resulting in low ATP production, low generation of reactive oxygen species (ROS) and low availability of tricarboxylic acid (TCA) cycle intermediates, all shifting RA T cells towards autoreactivity. The underlying defect stems from insufficient repair of mitochondrial DNA (mtDNA). Functional consequences include reversal of the TCA cycle, accumulation of citrate and lack of malate production. Excessive citrate promotes cytoskeletal hyperacetylation, creating hypermigratory and tissue-invasive T cells. Surplus acetyl-CoA supports lipid droplet formation and lipotoxicity. Lack of malate production disrupts the malate-aspartate shuttle, restricts recovery of cytosolic NAD and drives the endoplasmic reticulum (ER) into expansion. The bioenergetically stressed ER accumulates TNF mRNA and turns RA T cells into TNF superproducers. ATP low production renders RA T cells susceptible to cell death, depositing highly inflammatory mtDNA in the tissue. Mitochondrial deficiency leads to a slowdown in glycolysis and pyruvate processing, such that RA CD4+T cells shunt glucose towards the pentose phosphate pathway to support nucleotide synthesis and clonal proliferation. Metabolically deprived CD4+T cells partner with Mⱷ that have highly functional mitochondria. A hallmark of RA Mⱷ is the high expression of the DNA binding protein RFX5, which co-ordinates adaptations to metabolic needs with function. RFX5 upregulates HLA-DR expression and induces the glutaminolytic enzyme glutamate dehydrogenase 1 (GLUD1), providing bioenergetic resources for antigen presentation and survival in the tissue. In essence, RA CD4+T cells and Mⱷ function in a metabolically challenging environment and rewire their cellular metabolism to survive. Metabolic adaptations promote immunostimulation and tissue inflammation, triggering and sustaining rheumatoid synovitis.

The Lipid Droplet Protein DHRS3 Is a Regulator of Melanoma Cell State.

Lipid droplets are fat storage organelles composed of a protein envelope and lipid-rich core. Regulation of this protein envelope underlies differential lipid droplet formation and function. In melanoma, lipid droplet formation has been linked to tumor progression and metastasis, but it is unknown whether lipid droplet proteins play a role. To address this, we performed proteomic analysis of the lipid droplet envelope in melanoma. We found that lipid droplet proteins were differentially enriched in distinct melanoma states; from melanocytic to undifferentiated. DHRS3, which converts all-trans-retinal to all-trans-retinol, is upregulated in the MITFLO/undifferentiated/neural crest-like melanoma cell state and reduced in the MITFHI/melanocytic state. Increased DHRS3 expression is sufficient to drive MITFHI/melanocytic cells to a more undifferentiated/invasive state. These changes are due to retinoic acid-mediated regulation of melanocytic genes. Our data demonstrate that melanoma cell state can be regulated by expression of lipid droplet proteins which affect downstream retinoid signaling.

Protumoral lipid droplet-loaded macrophages are enriched in human glioblastoma and can be therapeutically targeted.

Glioblastoma presents a formidable clinical challenge because of its complex microenvironment. Here, we characterized tumor-associated foam cells (TAFs), a type of lipid droplet-loaded macrophage, in human glioblastoma. Through extensive analyses of patient tumors, together with in vitro and in vivo investigations, we found that TAFs exhibit distinct protumorigenic characteristics related to hypoxia, mesenchymal transition, angiogenesis, and impaired phagocytosis, and their presence correlates with worse outcomes for patients with glioma. We further demonstrated that TAF formation is facilitated by lipid scavenging from extracellular vesicles released by glioblastoma cells. We found that targeting key enzymes involved in lipid droplet formation, such as diacylglycerol O-acyltransferase or long-chain acyl-CoA synthetase, effectively disrupted TAF functionality. Together, these data highlight TAFs as a prominent immune cell population in glioblastoma and provide insights into their contribution to the tumor microenvironment. Disrupting lipid droplet formation to target TAFs may represent an avenue for future therapeutic development for glioblastoma.

Targeted modulation of pennycress lipid droplet proteins impacts droplet morphology and seed oil content.

Lipid droplets (LDs) are unusual organelles that have a phospholipid monolayer surface and a hydrophobic matrix. In oilseeds, this matrix is nearly always composed of triacylglycerols (TGs) for efficient storage of carbon and energy. Various proteins play a role in their assembly, stability and turnover, and even though the major structural oleosin proteins in seed LDs have been known for decades, the factors influencing LD formation and dynamics are still being uncovered mostly in the "model oilseed" Arabidopsis. Here we identified several key LD biogenesis proteins in the seeds of pennycress, a potential biofuel crop, that were correlated previously with seed oil content and characterized here for their participation in LD formation in transient expression assays and stable transgenics. One pennycress protein, the lipid droplet associated protein-interacting protein (LDIP), was able to functionally complement the Arabidopsis ldip mutant, emphasizing the close conservation of lipid storage among these two Brassicas. Moreover, loss-of-function ldip mutants in pennycress exhibited increased seed oil content without compromising plant growth, raising the possibility that LDIP or other LD biogenesis factors may be suitable targets for improving yields in oilseed crops more broadly.