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Per- and polyfluoroalkyl substances (PFAS) may cause various deleterious health effects. Epidemiological studies have demonstrated associations between PFAS exposure and adverse neurodevelopmental outcomes. The cytotoxicity, neurotoxicity, and mitochondrial toxicity of up to 12 PFAS including perfluoroalkyl carboxylates, perfluoroalkyl sulfonates, 6:2 fluorotelomer sulfonic acid (6:2 FTSA), and hexafluoropropylene oxide-dimer acid (HPFO-DA) were tested at concentrations typically observed in the environment (e.g., wastewater, biosolids) and in human blood using high-throughput in vitro assays. The cytotoxicity of all individual PFAS was classified as baseline toxicity, for which prediction models based on partition constants of PFAS between biomembrane lipids and water exist. No inhibition of the mitochondrial membrane potential and activation of oxidative stress response were observed below the cytotoxic concentrations of any PFAS tested. All mixture components and the designed mixtures inhibited the neurite outgrowth in differentiated neuronal cells derived from the SH-SY5Y cell line at concentrations around or below cytotoxicity. All designed mixtures acted according to concentration addition at low effect and concentration levels for cytotoxicity and neurotoxicity. The mixture effects were predictable from the experimental single compounds' concentration-response curves. These findings have important implications for the mixture risk assessment of PFAS.

A recently synthesized aminated 3,4-dioxygenated xanthone (Xantifoul2) was found to have promising antifouling (AF) effects against the settlement of the macrofouler Mytilus galloprovincialis larvae. Preliminary assessment indicated that Xantifoul2 has reduced ecotoxicological impacts: e.g., being non-toxic to the marine crustacea Artemia salina (<10 % mortality at 50 μM) and showing low bioconcentration factor in marine organisms. In order to meet the EU Biocidal Product Regulation, a preliminary hazard assessment of this new nature-inspired antifouling (NIAF) agent was conducted in this work. Xantifoul2 did not affect the swimming ability of the planktonic crustacean Daphnia magna, the growth of the diatom Phaeodactylum tricornutum, and the cellular respiration of luminescent Gram-negative bacteria Vibrio fischeri, supporting the low toxicity towards several non-target marine species. Regarding human cytotoxicity, Xantifoul2 did not affect the cell viability of retinal human cells (hTERT-RPE-1) and lipidomic studies revealed depletion of lipids involved in cell death, membrane modeling, lipid storage, and oxidative stress only at a high concentration (10 μM). Accelerated degradation studies in water were conducted under simulated sunlight to allow the understanding of putative transformation products (TPs) that could be generated in the aquatic ecosystems. Both Xantifoul2 and photolytic-treated Xantifoul2 in the aqueous matrix were therefore evaluated on several nuclear receptors (NRs). The results of this preliminary hazard assessment of Xantifoul2, combined with the high degradation rates in water, provide strong evidence of the safety of this AF agent under the evaluated conditions, and provide the support for future validation studies before this compound can be introduced in the market., Competing Interests: Declaration of competing interest Marta Correia-da-Silva has patent Xanthonic compounds and their use as antifouling agents issued to CN113226035. Emilia Sousa has patent Xanthonic compounds and their use as antifouling agents issued to CN113226035. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Metabolic adaptations in response to changes in energy supply and demand are essential for survival. The mitochondrial calcium uniporter coordinates metabolic homeostasis by regulating TCA cycle activation, mitochondrial fatty acid oxidation and cellular calcium signaling. However, a comprehensive analysis of uniporter-regulated mitochondrial metabolic pathways has remained unexplored. Here, we investigate the metabolic consequences of uniporter loss- and gain-of-function, and identify a key transcriptional regulator that mediates these effects. Using gene expression profiling and proteomic, we find that loss of uniporter function increases the expression of proteins in the branched-chain amino acid (BCAA) catabolism pathway. Activity is further augmented through phosphorylation of the enzyme that catalyzes this pathway's committed step. Conversely, in the liver cancer fibrolamellar carcinoma (FLC)-which we demonstrate to have high mitochondrial calcium levels- expression of BCAA catabolism enzymes is suppressed. We also observe uniporter-dependent suppression of the transcription factor KLF15, a master regulator of liver metabolic gene expression, including those involved in BCAA catabolism. Notably, loss of uniporter activity upregulates KLF15, along with its transcriptional target ornithine transcarbamylase (OTC), a component of the urea cycle, suggesting that uniporter hyperactivation may contribute to the hyperammonemia observed in FLC patients. Collectively, we establish that FLC has increased mitochondrial calcium levels, and identify an important role for mitochondrial calcium signaling in metabolic adaptation through the transcriptional regulation of metabolism., Competing Interests: CONFLICT OF INTEREST STATEMENT The authors have no conflicts of interest to disclose.

Per- and polyfluoroalkyl substances (PFAS) are widely used in many industrial and domestic applications, which has resulted in unintentional human exposures and bioaccumulation in blood and other organs. Perfluorooctane sulfonate (PFOS) is among the most prevalent PFAS in the environment and has been postulated to affect brain functions in exposed organisms. However, the impacts of PFOS in early neural development have not been well described. We used zebrafish larvae to assess the effects of PFOS on two fundamental complex behaviors, prey capture and learning. Zebrafish exposed to PFOS concentrations ranging from 2 to 20 µM for differing 48-h periods were viable through early larval stages. In addition, PFOS uptake was unaffected by the presence of a chorion. We employed two different experimental paradigms; first we assessed the impacts of increasing organismal PFOS bioaccumulation on prey capture and learning, and second, we probed stage-specific sensitivity to PFOS by exposing zebrafish at different developmental stages (0-2 vs. 3-5 days post fertilization). Following both assays we measured the amount of PFOS present in each larva and found that PFOS levels varied in larvae from different groups within each experimental paradigm. Significant negative correlations were observed between larval PFOS accumulation and percentage of captured prey, whereas nonsignificant negative correlations were observed between PFOS accumulation and experienced-induced prey capture learning. These findings suggest that PFOS accumulation negatively affects larval zebrafish's ability to perform complicated multisensory behaviors and highlights the potential risks of PFOS exposure to animals in the wild, with implications for human health. Environ Toxicol Chem 2024;43:847-855. © 2023 SETAC., (© 2023 SETAC.)

Mixed lineage kinase domain-like (MLKL) is a key signaling protein of necroptosis. Upon activation by phosphorylation, MLKL translocates to the plasma membrane and induces membrane permeabilization, which contributes to the necroptosis-associated inflammation. Membrane binding of MLKL is initially initiated by electrostatic interactions between the protein and membrane phospholipids. We previously showed that MLKL and its phosphorylated form (pMLKL) are S -acylated during necroptosis. Here, we characterize the acylation sites of MLKL and identify multiple cysteines that can undergo acylation with an interesting promiscuity at play. Our results show that MLKL and pMLKL undergo acylation at a single cysteine, with C184, C269, and C286 as possible acylation sites. Using all-atom molecular dynamic simulations, we identify differences that the acylation of MLKL causes at the protein and membrane levels. Through investigations of the S -palmitoyltransferases that might acylate pMLKL in necroptosis, we showed that zDHHC21 activity has the strongest effect on pMLKL acylation, inactivation of which profoundly reduced the pMLKL levels in cells and improved membrane integrity. These results suggest that blocking the acylation of pMLKL destabilizes the protein at the membrane interface and causes its degradation, ameliorating the necroptotic activity. At a broader level, our findings shed light on the effect of S -acylation on MLKL functioning in necroptosis and MLKL-membrane interactions mediated by its acylation.

We have predicted acid dissociation constants (p K a ), octanol-water partition coefficients ( K OW ), and DMPC lipid membrane-water partition coefficients ( K lipid-w ) of 150 different eight-carbon-containing poly-/perfluoroalkyl carboxylic acids (C8-PFCAs) utilizing the COnductor-like Screening MOdel for Realistic Solvents (COSMO-RS) theory. Different trends associated with functionalization, degree of fluorination, degree of saturation, degree of chlorination, and branching are discussed on the basis of the predicted values for the partition coefficients. In general, functionalization closest to the carboxylic headgroup had the greatest impact on the value of the predicted physicochemical properties., Competing Interests: The authors declare no competing financial interest.

The SH-SY5Y, neuroblastoma cell line, is a common in vitro model used to study physiological neuronal function and the neuronal response to different stimuli, including exposure to toxic chemicals. These cells can be differentiated to neuron-like cells by administration of various reagents, including retinoic acid or phorbol-12-myristate-13-acetate. Despite their common use, there is an incomplete understanding of the molecular changes that occur during differentiation. Therefore, there is a critical need to fully understand the molecular changes that occur during differentiation to properly study neurotoxicity in response to various environmental exposures. Previous studies have investigated the proteome and transcriptome during differentiation; however, the regulation of the cellular lipidome in this process is unexplored. In this work, we conducted liquid chromatography-mass spectrometry (LC-MS)-based untargeted lipidomics in undifferentiated and differentiated SH-SY5Y cells, induced by retinoic acid. We show that there are global differences between the cellular lipidomes of undifferentiated and differentiated cells. Out of thousands of features detected in positive and negative electrospray ionization modes, 44 species were identified that showed significant differences ( p -value ≤0.05, fold change ≥2) in differentiated cells. Identification of these features combined with targeted lipidomics highlighted the accumulation of phospholipids, sterols, and sphingolipids during differentiation while triacylglycerols were depleted. These results provide important insights into lipid-related changes that occur during cellular differentiation of SH-5YSY cells and emphasize the need for the detailed characterization of biochemical differences that occur during differentiation while using this in vitro model for assessing ecological impacts of environmental pollutants., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Co-published by Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, and American Chemical Society.)

Necroptosis is a type of programed cell death characterized by an inflammatory phenotype due to extensive membrane permeabilization and rupture. Initiation of necroptosis involves activation of tumor necrosis factor receptors by tumor necrosis factor alpha (TNFα) followed by coordinated activities of receptor-interacting protein kinases and mixed lineage kinase-like protein (MLKL). Subsequently, MLKL undergoes phosphorylation and translocates to the plasma membrane, leading to permeabilization. Such permeabilization results in the release of various cytokines and causes extensive inflammatory activity at the organismal level. This inflammatory activity is one of the major differences between apoptosis and necroptosis and links necroptosis to several human pathologies that exhibit inflammation, in addition to the ultimate cell death phenotype. Given the crosstalk between the activation of cell death pathway and inflammatory activity, approaches that provide insights on the regulation of transcripts, proteins and their processing at the global level have substantially improved our understanding of necroptosis and its involvement in different disease states. In this review, we highlight recent omic studies probing the transcriptome, proteome and lipidome which elucidate potential new mechanisms and signaling pathways during necroptosis and the necroptosis-associated inflammatory activity observed in various diseases. We specifically focus on studies investigating the transcriptome and intracellular and released proteome that contribute to inflammatory nature of necroptotic cells. We also highlight different lipids that have been implicated in necroptosis and lipidomic studies identifying lipid players in necroptosis. Finally, we review studies which suggest certain necroptosis-related genes as potential prognosis markers for different cancers and discuss their translational implications.

Necroptosis is a type of programmed cell death. It is characterized by membrane permeabilization and is associated with the release of intracellular components due to compromised membrane integrity which induces a strong inflammatory response. We recently showed that the accumulation of very long chain fatty acids (VLCFAs) contributes to membrane permeabilization during necroptosis. However, the mechanisms that result in the accumulation of these cytotoxic lipids remain unknown. Using comparative transcriptomics and digital PCR validations, we found that several target genes of sterol regulatory element-binding proteins (SREBPs) were upregulated during necroptosis, suggesting that they might be responsible for the accumulation of VLCFA in this process. We demonstrated that activation of SREBPs during necroptosis exacerbates the permeability of the plasma membrane and cell death. Consistent with these observations, targeting sterol regulatory element-binding protein cleavage-activating protein (SCAP), a protein involved in SREBP activation, reversed the accumulation of VLCFAs, and restored cell death and membrane permeabilization during necroptosis. Collectively, our results highlight a role for SREBP in regulating lipid changes during necroptosis and suggest SREBP-mediated lipid remodeling as a potential target for therapeutics to reduce membrane permeabilization during necroptosis., Competing Interests: The authors declare no conflict of interest., (This journal is © The Royal Society of Chemistry.)

Lipids, the structural part of membranes, play important roles in biological functions. However, our understanding of their implication in key cellular processes such as cell division and protein-lipid interaction is just emerging. This is the case for molecular interactions in mechanisms of cell death, where the role of lipids for protein localization and subsequent membrane permeabilization is key. For example, during the last stage of necroptosis, the mixed lineage kinase domain-like (MLKL) protein translocates and, eventually, permeabilizes the plasma membrane (PM). This process results in the leakage of cellular content, inducing an inflammatory response in the microenvironment that is conducive to oncogenesis and metastasis, among other pathologies that exhibit inflammatory activity. This work presents insights from long all-atom molecular dynamics (MD) simulations of complex membrane models for the PM of mammalian cells with an MLKL protein monomer. Our results show that the binding of the protein is initially driven by the electrostatic interactions of positively charged residues. The protein bound conformation modulates lipid recruitment to the binding site, which changes the local lipid environment recruiting PIP lipids and cholesterol, generating a unique fingerprint. These results increase our knowledge of protein-lipid interactions at the membrane interface in the context of molecular mechanisms of the necroptotic pathway, currently under investigation as a potential treatment target in cancer and inflamatory diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Ramirez, Campbell, Pradhan, Atilla-Gokcumen and Monje-Galvan.)

One of the classic hallmarks of cancer is the imbalance between elevated cell proliferation and reduced cell death. Ceramide, a bioactive sphingolipid that can regulate this balance, has long been implicated in cancer. While the effects of ceramide on cell death and therapeutic efficacy are well established, emerging evidence indicates that ceramide turnover to downstream sphingolipids, such as sphingomyelin, hexosylceramides, sphingosine-1-phosphate, and ceramide-1-phosphate, is equally important in driving pro-tumorigenic phenotypes, such as proliferation, survival, migration, stemness, and therapy resistance. The complex and dynamic sphingolipid network has been extensively studied in several cancers, including breast cancer, to find key sphingolipidomic alterations that can be exploited to develop new therapeutic strategies to improve patient outcomes. Here, we review how the current literature shapes our understanding of how ceramide synthesis and turnover are altered in breast cancer and how these changes offer potential strategies to improve breast cancer therapy.

Antifouling (AF) coatings containing booster biocides are used worldwide as one of the most cost-effective ways to prevent the attachment of marine organisms to submerged structures. Nevertheless, many of the commercial biocides, such as Econea ® (tralopyril), are toxic in marine environments. For that reason, it is of extreme importance that new efficient AF compounds that do not cause any harm to non-target organisms and humans are designed. In this study, we measured the half-maximal inhibitory concentration (IC 50 ) of a promising nature-inspired AF compound, a triazolyl glycosylated chalcone (compound 1 ), in an immortalized human retinal pigment epithelial cell line (hTERT-RPE-1) and compared the results with the commercial biocide Econea ® . We also investigated the effects of these biocides on the cellular lipidome following an acute (24 h) exposure using liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF/MS). Our results showed that compound 1 did not affect viability in hTERT-RPE-1 cells at low concentrations (1 μM), in contrast to Econea ® , which caused a 40% reduction in cell viability. In total, 71 lipids were found to be regulated upon exposure to 10 µM of both compounds. Interestingly, both compounds induced changes in lipids involved in cell death, membrane modeling, lipid storage, and oxidative stress, but often in opposing directions. In general, Econea ® exposure was associated with an increase in lipid concentrations, while compound 1 exposure resulted in lipid depletion. Our study showed that exposure to human cells at sublethal Econea ® concentrations results in the modulation of several lipids that are linked to cell death and survival.

Per- and poly-fluorinated substances (PFASs) are organic pollutants that have been linked to numerous health effects, including diabetes, cancers, and dysregulation of the endocrine system. This study aims to develop a liquid chromatography with tandem mass spectrometry (LC-MS/MS) assay to measure changes in 17 hormones in H295R cell line (a steroid producing adrenocortical cells) upon exposure to PFASs. Due to the challenges in the analysis of steroid hormones using electrospray ionization MS, a chemical derivatization method was employed to achieve 0.07-2 μg/L detection limits in LC-MS/MS. Furthermore, a 10-fold concentration factor through solid-phase extraction (SPE) allows for consistent sub-parts per billion detections. Optimization of the derivatization conditions showed doubly-derivatized products in some hormone analytes, including progesterone, corticosterone, and cortisol, and gave improved ionization efficiency up to 20-fold higher signal than the singly-derivatized product. The use of SPE for sample cleanup to analyze hormones from cellular media using weak anion exchange sorbent yielded 80-100% recovery for the 17 targeted hormones. The method was validated by exposing H295R cells to two known endocrine disruptors, forskolin and prochloraz, which showed expected changes in hormones. An initial exposure of H295R cells with various PFAS standards and their mixtures at 1 μM showed significant increases in progestogens with some PFAS treatments, which include PFBS, PFHxA, PFOS, PFDA, and PFDS. In addition, modest changes in hormone levels were observed in cells treated with other sulfonated or carboxylated headgroup PFASs. This sensitive LC-MS/MS method for hormone analysis in H295R cells will allow for the investigations of the alterations in the hormone production caused by exposure to various environmental insults in cell-based assays and other in vitro models.

ET resistance is a critical problem for estrogen receptor-positive (ER+) breast cancer. In this study, we have investigated how alterations in sphingolipids promote cell survival in ET-resistant breast cancer. We have performed LC-MS-based targeted sphingolipidomics of tamoxifen-sensitive and -resistant MCF-7 breast cancer cell lines. Follow-up studies included treatments of cell lines and patient-derived xenograft organoids (PDxO) with small molecule inhibitors; cytometric analyses to measure cell death, proliferation, and apoptosis; siRNA-mediated knockdown; RT-qPCR and Western blot for gene and protein expression; targeted lipid analysis; and lipid addback experiments. We found that tamoxifen-resistant cells have lower levels of ceramides and hexosylceramides compared to their tamoxifen-sensitive counterpart. Upon perturbing the sphingolipid pathway with small molecule inhibitors of key enzymes, we identified that CERK is essential for tamoxifen-resistant breast cancer cell survival, as well as a fulvestrant-resistant PDxO. CERK inhibition induces ceramide-mediated cell death in tamoxifen-resistant cells. Ceramide-1-phosphate (C1P) partially reverses CERK inhibition-induced cell death in tamoxifen-resistant cells, likely through lowering endogenous ceramide levels. Our findings suggest that ET-resistant breast cancer cells maintain lower ceramide levels as an essential pro-survival mechanism. Consequently, ET-resistant breast cancer models have a unique dependence on CERK as its activity can inhibit de novo ceramide production.

Per- and polyfluorinated alkyl substances (PFAS) are a class of widely used compounds in an array of commercial and industrial applications. Due to their extensive use and chemical stability, PFAS persist in the environment and bioaccumulate in humans and wildlife. PFAS exposure have been linked to several negative health effects, including the formation of various cancers, disruption of the endocrine system, and obesity. However, there is a major gap in understanding how structural differences in PFAS impact their interactions within a biological system. In this study, we examined the toxicity of PFAS with differences in chain length, head group, and degree of fluorination in human retinal epithelial cells. We focused on fluorotelomeric and fully fluorinated sulfonates and carboxylates and measured their uptake. Our results showed that sulfonates are taken up at higher levels as compared to their fluorotelomer and carboxylate counterparts. Furthermore, PFAS with 8 and 10 carbons (C8 and C10) are taken up at a higher level compared to those with six carbons (C6). We also investigated the role of the fatty acid transporter CD36 in PFAS uptake and found that increased CD36 levels result in higher levels of PFAS in cells. Overall, our results suggest that the head group structure of PFAS impacts toxicity, with sulfonates inducing a higher decrease in cell viability (∼50%) than carboxylates. Our results also link the activity of CD36 to PFAS uptake into cells.

Sphingolipids are key signaling lipids and their dysregulation has been associated with various cellular processes. We have previously shown significant changes in sphingolipids in therapy-induced senescence, a state of cell cycle arrest as a response to chemotherapy, including the accumulation of ceramides, and provided evidence suggesting that ceramide processing is important for this process. Herein, we conducted a focused small molecule inhibitor screen targeting the sphingolipid pathway, which highlighted a new lipid regulator of therapy-induced senescence. Among the inhibitors tested, the inhibition of ceramide kinase by NVP-231 reduced the levels of senescent cells. Ceramide kinase knockdown exhibited similar effects, strongly supporting the involvement of ceramide kinase during this process. We showed that ceramide-1-phosphate was upregulated in therapy-induced senescence and that NVP-231 reduced ceramide-1-phosphate levels in different cell line models of therapy-induced senescence. Finally, ceramide-1-phosphate addition to NVP-231-treated cells reversed the effects of NVP-231 during senescence. Overall, our results identify a previously unknown lipid player in therapy-induced senescence and highlight a potential targetable enzyme to reduce the levels of therapy-induced senescent cells.

Sphingolipids are highly bioactive lipids. Sphingolipid metabolism produces key membrane components (e.g. sphingomyelin) and a variety of signaling lipids with different biological functions (e.g. ceramide, sphingosine-1-phosphate). The coordinated activity of tens of different enzymes maintains proper levels and localization of these lipids with key roles in cellular processes. In this review, we highlight the signaling roles of sphingolipids in cell death and survival. We discuss recent findings on the role of specific sphingolipids during these processes, enabled by the use of lipidomics to study compositional and spatial regulation of these lipids and synthetic sphingolipid probes to study subcellular localization and interaction partners of sphingolipids to understand the function of these lipids., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The common deletion of the third exon of the growth hormone receptor gene ( GHRd3 ) in humans is associated with birth weight, growth after birth, and time of puberty. However, its evolutionary history and the molecular mechanisms through which it affects phenotypes remain unresolved. We present evidence that this deletion was nearly fixed in the ancestral population of anatomically modern humans and Neanderthals but underwent a recent adaptive reduction in frequency in East Asia. We documented that GHRd3 is associated with protection from severe malnutrition. Using a novel mouse model, we found that, under calorie restriction, Ghrd3 leads to the female-like gene expression in male livers and the disappearance of sexual dimorphism in weight. The sex- and diet-dependent effects of GHRd3 in our mouse model are consistent with a model in which the allele frequency of GHRd3 varies throughout human evolution as a response to fluctuations in resource availability.

Necroptosis is a form of cell death characterized by receptor-interacting protein kinase activity and plasma membrane permeabilization via mixed-lineage kinase-like protein (MLKL). This permeabilization is responsible for the inflammatory properties of necroptosis. We previously showed that very long chain fatty acids (VLCFAs) are functionally involved in necroptosis, potentially through protein fatty acylation. Here, we define the scope of protein acylation by saturated VLCFAs during necroptosis. We show that MLKL and phosphoMLKL, key for membrane permeabilization, are exclusively acylated during necroptosis. Reducing the levels of VLCFAs decreases their membrane recruitment, suggesting that acylation by VLCFAs contributes to their membrane localization. Acylation of phosphoMLKL occurs downstream of phosphorylation and oligomerization and appears to be, in part, mediated by ZDHHC5 (a palmitoyl transferase). We also show that disruption of endosomal trafficking increases cell viability during necroptosis, possibly by preventing recruitment, or removal, of phosphoMLKL from the plasma membrane., Competing Interests: Declaration of interests The authors declare no competing interest., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Liposomes containing small amounts of porphyrin-phospholipid (PoP) have been shown to encapsulate small molecular weight cargos and then release them upon exposure to red light. A putative mechanism involves transient pore formation in the bilayer induced by PoP-mediated photo-oxidation of unsaturated lipids. However, little is known about the properties of such pores. This study assesses whether large carbohydrate and protein molecules could be released from PoP liposomes upon red light exposure. A small fluorophore with ∼0.5 kDa in molecular weight, fluorescently labeled dextrans of ∼5 and ∼500 kDa, and a ∼240 kDa fluorescent protein were passively entrapped in PoP liposomes. When exposed to 665 nm irradiation, liposomes containing PoP, but not liposomes lacking it, released all these cargos in a size-dependent manner that occurred with oxidation of unsaturated lipids included in the bilayer. Thus, this study demonstrates the feasibility of light-triggered release of large biomacromolecules from liposomes.

Research in fundamental cell biology and pathology could be revolutionized by developing the capacity for quantitative molecular analysis of subcellular structures. To that end, we introduce the Ramanomics platform, based on confocal Raman microspectrometry coupled to a biomolecular component analysis algorithm, which together enable us to molecularly profile single organelles in a live-cell environment. This emerging omics approach categorizes the entire molecular makeup of a sample into about a dozen of general classes and subclasses of biomolecules and quantifies their amounts in submicrometer volumes. A major contribution of our study is an attempt to bridge Raman spectrometry with big-data analysis in order to identify complex patterns of biomolecules in a single cellular organelle and leverage discovery of disease biomarkers. Our data reveal significant variations in organellar composition between different cell lines. We also demonstrate the merits of Ramanomics for identifying diseased cells by using prostate cancer as an example. We report large-scale molecular transformations in the mitochondria, Golgi apparatus, and endoplasmic reticulum that accompany the development of prostate cancer. Based on these findings, we propose that Ramanomics datasets in distinct organelles constitute signatures of cellular metabolism in healthy and diseased states.

We report short ceramide analogs that can be activated with light and further functionalized using azide-alkyne click chemistry. These molecules, termed scaCers , exhibit increased cell permeability compared to their long-chain analogs as demonstrated using mass spectrometry and imaging. Notably, scaCers enable optical control of apoptosis, which is not observed with long-chain variants. Additionally, they function as photoswitchable substrates for sphingomyelin synthase 2 (SMS2), exhibiting inverted light-dependence compared to their extended analogs.

Lipids are emerging as key players of senescence. Here, we review the exciting new findings on the diverse roles of lipids in cellular senescence, most of which are enabled by the advancements in omics approaches. Senescence is a cellular process in which the cell undergoes growth arrest while retaining metabolic activity. At the organismal level, senescence contributes to organismal aging and has been linked to numerous diseases. Current research has documented that senescent cells exhibit global alterations in lipid composition, leading to extensive morphological changes through membrane remodeling. Moreover, senescent cells adopt a secretory phenotype, releasing various components to their environment that can affect the surrounding tissue and induce an inflammatory response. All of these changes are membrane and, thus, lipid related. Our work, and that of others, has revealed that fatty acids, sphingolipids, and glycerolipids are involved in the initiation and maintenance of senescence and its associated inflammatory components. These studies opened up an exciting frontier to investigate the deeper mechanistic understanding of the regulation and function of these lipids in senescence. In this review, we will provide a comprehensive snapshot of the current state of the field and share our enthusiasm for the prospect of potential lipid-related protein targets for small-molecule therapy in pathologies involving senescence and its related inflammatory phenotypes.

In Barth syndrome (BTHS) mutations in tafazzin leads to changes in both the quantities and the molecular species of cardiolipin (CL), which are the hallmarks of BTHS. Contrary to the well-established alterations in CL associated with BTHS; recently a marked decrease in the plasmalogen levels in Barth specimens has been identified. To restore the plasmalogen levels, the present study reports the effect of promotion of plasmalogen biosynthesis on the lipidome of lymphoblasts derived from Barth patients as well as on cell viability, mitochondria biogenesis, and mitochondrial membrane potential. High resolution 31 P NMR phospholipidomic analysis showed an increase in the levels of plasmenylethanolamine (the major plasmalogen in lymphoblasts), which reached values comparable to the control and a compensatory decrease in the levels of its diacyl-PE counterpart. Importantly, 31 P NMR showed a significant increase in the levels of CL, while not altering the levels of monolysocardiolipin. Mass spectrometry measurements showed that the promotion of plasmalogen biosynthesis did not change the molecular species profile of targeted phospholipids. In addition, promotion of plasmalogen biosynthesis did not impact on cellular viability, although it significantly decrease mitochondria copy number and restored mitochondrial membrane potential. Overall, the results showed the efficacy of the promotion of plasmalogen biosynthesis on increasing the CL levels in a BTHS cell model and highlight the potential beneficial effect of a diet supplemented with plasmalogen precursors to BTHS patients., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest with the content of this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Coessentiality mapping has been useful to systematically cluster genes into biological pathways and identify gene functions 1-3 . Here, using the debiased sparse partial correlation (DSPC) method 3 , we construct a functional coessentiality map for cellular metabolic processes across human cancer cell lines. This analysis reveals 35 modules associated with known metabolic pathways and further assigns metabolic functions to unknown genes. In particular, we identify C12orf49 as an essential regulator of cholesterol and fatty acid metabolism in mammalian cells. Mechanistically, C12orf49 localizes to the Golgi, binds membrane-bound transcription factor peptidase, site 1 (MBTPS1, site 1 protease) and is necessary for the cleavage of its substrates, including sterol regulatory element binding protein (SREBP) transcription factors. This function depends on the evolutionarily conserved uncharacterized domain (DUF2054) and promotes cell proliferation under cholesterol depletion. Notably, c12orf49 depletion in zebrafish blocks dietary lipid clearance in vivo, mimicking the phenotype of mbtps1 mutants. Finally, in an electronic health record (EHR)-linked DNA biobank, C12orf49 is associated with hyperlipidaemia through phenome analysis. Altogether, our findings reveal a conserved role for C12orf49 in cholesterol and lipid homeostasis and provide a platform to identify unknown components of other metabolic pathways.

Therapy-induced senescence is a state of cell cycle arrest that occurs as a response to various chemotherapeutic reagents, especially ones that cause DNA damage. Senescent cells display resistance to cell death and can impair the efficacy of chemotherapeutic strategies. Since lipids can exhibit pro-survival activity, it is envisioned in this article that probing the lipidome could provide insights into novel lipids that are involved in senescence. Therefore, a tissue culture model system is established and the cellular lipidomes of senescent and proliferating cells are comparatively analyzed. Out of thousands of features detected, 17 species are identified that show significant changes in senescent cells. The majority of these species (11 out of 17) are atypical sphingolipids, 1-deoxyceramides/dihydroceramides, which are produced as a result of the utilization of alanine, instead of serine during sphingolipid biosynthesis. These lipids are depleted in senescent cells. Elevating the levels of deoxyceramides by supplementing the growth medium with metabolic precursors or by directly adding deoxyceramide result in decreased senescence, suggesting that these species might play a key role in this process., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Siderophores are small molecule metal chelators secreted in sparse quantities by their native microbial hosts but can be engineered for enhanced production from heterologous hosts like Escherichia coli. These molecules have been proved to be capable of binding heavy metals of commercial and/or environmental interest. In this work, we incorporated, as needed, the appropriate pathways required to produce several siderophores (anguibactin, vibriobactin, bacillibactin, pyoverdine, and enterobactin) into the base E. coli K-12 MG1655 metabolic network model to computationally predict, via flux balance analysis methodologies, gene knockout targets, gene over-expression targets, and media modifications capable of improving siderophore reaction flux. E. coli metabolism proved supportive for the underlying production mechanisms of various siderophores. Within such a framework, the gene deletion and over-expression targets identified, coupled with complementary insights from medium optimization predictions, portend experimental implementation to both enable and improve heterologous siderophore production. Successful production of siderophores would then spur novel metal-binding applications., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Mass-spectrometry based metabolomics has recently emerged as a valuable technique in understanding the ecotoxicity and mode of action of a wide range of xenobiotics in the environment, including engineered nanomaterials (ENMs). However, the applications of metabolomics in elucidating the biochemical pathways affected by xenobiotics have been mostly performed using targeted analysis. In this study, the effects of copper oxide nanoparticles (CuO NPs) on Arabidopsis thaliana, a model plant, was investigated using untargeted metabolite profiling based on two platforms of high-resolution mass spectrometry (MS): (1) liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QToF-MS) and (2) LC Q Exactive™ Hybrid Quadrupole-Orbitrap™-MS (LC-Orbitrap-MS). This approach was performed to identify specific features (mass-to-charge ratios, m/z's) that are significantly changed in a reproducible manner regardless of the MS platform used in metabolomics. In addition, the total copper concentrations taken up in plant tissues were quantified using inductively coupled plasma mass spectrometry (ICP-MS), which provided evidence of translocation of CuO NPs from roots to leaves and flowering shoots. Results from untargeted metabolomics showed that there were 65 plant metabolites that were altered commonly in both LC/MS platforms resulting from CuO NPs exposure of Arabidopsis thaliana. These metabolites belong to the jasmonic acid and glucosinolates pathways, suggesting the stress response induced by CuO NPs in Arabidopsis. This study demonstrated the effectiveness of high-resolution LC/MS in providing insight on the mechanism of nanotoxicity of CuO NPs in plants., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Necroptosis is a form of regulated cell death which results in loss of plasma membrane integrity, release of intracellular contents, and an associated inflammatory response. We previously found that saturated very long chain fatty acids (VLCFAs), which contain ≥20 carbons, accumulate during necroptosis. Here, we show that genetic knockdown of Fatty Acid (FA) Elongase 7 (ELOVL7) reduces accumulation of specific very long chain FAs during necroptosis, resulting in reduced necroptotic cell death and membrane permeabilization. Conversely, increasing the expression of ELOVL7 increases very long chain fatty acids and membrane permeabilization. In vitro , introduction of the VLCFA C24 FA disrupts bilayer integrity in liposomes to a greater extent than a conventional C16 FA. To investigate the microscopic origin of these observations, atomistic Molecular Dynamics (MD) simulations were performed. MD simulations suggest that fatty acids cause clear differences in bilayers based on length and that it is the interdigitation of C24 FA between the individual leaflets that results in disorder in the region and, consequently, membrane disruption. We synthesized clickable VLCFA analogs and observed that many proteins were acylated by VLCFAs during necroptosis. Taken together, these results confirm the active role of VLCFAs during necroptosis and point to multiple potential mechanisms of membrane disruption including direct permeabilization via bilayer disruption and permeabilization by targeting of proteins to cellular membranes by fatty acylation.

The use of cultured cells has been instrumental in studying biochemical, molecular, and cellular processes. The composition of serum that cells are maintained in can have a profound impact on important cellular checkpoints. Cell growth and apoptosis are analyzed in an estrogen receptor positive breast cancer cell line in the presence of serum that have been treated to remove steroids or lipids, as well-described in the literature. It is shown that maintaining cells in the presence of charcoal-dextran-treated serum causes reduced growth rate, which can be reversed by the addition of estradiol. Silica-treated-serum also slows down cell growth and induces apoptosis. In order to investigate the role of lipids in these phenotypes, the levels of a wide range of lipids in different sera are investigated. It is shown that silica-treatment significantly depletes phosphatidylcholines and cholesterol. It is also shown that lipogenesis is stimulated when cells are cultured with silica-treated-serum and this is reversed by the addition of exogenous lipids, which also restores growth rate and apoptosis. The results show that cultured cells are sensitive to different serum, most likely due to the differences in levels of structural and signaling metabolites present in their growth environment., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Metabolomics, the global profiling of metabolite composition, is a powerful technique that can be applied to answer a diverse set of research questions concerning effects of toxicants on organisms. It has recently emerged as a tool to understand complex environmental perturbations in biological systems, especially at sub-lethal concentrations. Organisms can be affected by different stressors such as xenobiotics or increase in concentration of natural compounds such as nitrogen, phosphorous, and sulfur. Metabolomics has facilitated a better understanding of the effects of these perturbations on organisms such as plants, animals, and humans providing phenotypic and biological information in a high throughput manner. In this review, we will discuss recent applications of metabolomics to study the ecological effects of different environmental perturbations, including nanoparticles, pharmaceuticals and personal care products, pesticides, as well as the changes in natural compounds found in the environment with a focus on plant systems., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Lipids are emerging as key regulators of apoptosis. Specific lipid species are associated with apoptosis with important functional roles, but the understanding of the regulation of these lipid species is still limited. It has been previously shown by our laboratory that polyunsaturated triacylglycerols accumulate and get stored within lipid droplets during apoptosis via activated glycerolipid biosynthesis. In this work, the biochemical mechanisms that result in the activation of glycerolipid biosynthesis and, consequently, triacylglycerol and lipid droplet accumulation during apoptosis are investigated. The transcriptomes of control and apoptotic HCT-116 cells are compared and gene enrichment analysis revealed the upregulation of p38 mitogen-activated protein kinase (MAPK). It is shown that p38 MAPK regulates triacylglycerol biosynthesis through diacylglycerol acyltransferase1 during apoptosis. Perilipin 2 and cytosolic phospholipase A2delta are also shown to be involved in lipid droplet and polyunsaturated triacylglycerol accumulation in this process. Overall, the results provide new insights into the upregulation of glycerolipid synthesis during apoptosis., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Engineered nanomaterials have unique properties compared to their bulk counterparts. Copper oxide nanoparticles (CuO NPs) are one example of nanomaterials used in a wide range of consumer products due to their conductivity and biocidal properties. While CuO NPs can induce toxicity in various organisms, their interactions with different organisms and how they affect cellular homeostasis is yet to be fully understood. In this work, the toxicity of CuO NPs was evaluated in different human cell lines (colorectal carcinoma, cervical cancer, embryonic kidney, and lung fibroblast), showing a dose-dependent toxicity. An untargeted lipidomics approach using liquid chromatography-quadrupole time of flight mass spectrometry was employed in a human colon carcinoma cell line to investigate the impact of CuO NP exposure at the cellular level. A 24 h CuO NP exposure at 2.5 and 5 μg mL-1 resulted in upregulation of different metabolites: triacylglycerols, phosphatidylcholines, and ceramides accumulated. The most profound increase in a dose-dependent manner was observed in ceramides, specifically in C18:0, C18:1, and C22:0 species, with up to ∼10 fold accumulations. Further experiments suggested that activation of autophagy and oxidative stress could be responsible for the toxicity observed in these cell lines. Increases in the level of glutathione oxide (∼7 fold) also supported the activation of oxidative stress upon CuO NP treatment. Based on the changes in different metabolites induced by CuO NP exposure and previous studies from our laboratory, we propose that autophagy and oxidative stress could play a role in CuO NP-induced toxicity.

The identification of myriad of chemicals in the environment that mimic hormones and affect the endocrine functions of exposed organism is a daunting analytical challenge for environmental scientists and engineers. Many of these endocrine disrupting chemicals (EDCs) are present at very low concentrations in the aquatic systems, but yet affect the metabolic, developmental, and reproductive functions in exposed fish and wildlife. Vitellogenin (VTG) protein is a widely used biomarker in fish for assessing exposure to EDCs, and is commonly measured using species-specific immunochemical techniques. In this study, we developed a liquid chromatography tandem mass spectrometry (LC-MS/MS) method that can measure common peptides from digested VTG in multiple fish species. In the initial experiments using high resolution mass spectrometry, two peptides (ALHPELR and FIELIQLLR) were identified as common fragments in the digested VTG protein isolated from three different fish species (Pimephales promelas, Micropterus salmoides, and Fundulus heteroclitus). Then, a quantitative analysis using LC-MS/MS under selected reaction monitoring mode was developed for the detection of these two peptides in trypsin-digested plasma from female fish (positive control), estrogen-exposed male fish (test sample), and unexposed male fish (negative control) using two of the same species used for identifying the common peptides (P. promelas, and M. salmoides) and one new species (Ameiurus nebulosus) that was not included during the selection of peptides. Results from this study demonstrate the potential of LC-MS/MS as an effective cross-species method to detect VTG in fish, which can be an alternative analytical technique for assessing endocrine disruption in multiple fish species., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Metabolic decoys are synthetic analogs of naturally occurring biosynthetic acceptors. These compounds divert cellular biosynthetic pathways by acting as artificial substrates that usurp the activity of natural enzymes. While O-linked glycosides are common, they are only partially effective even at millimolar concentrations. In contrast, we report that N-acetylglucosamine (GlcNAc) incorporated into various thioglycosides robustly truncate cell surface N- and O-linked glycan biosynthesis at 10-100 μM concentrations. The >10-fold greater inhibition is in part due to the resistance of thioglycosides to hydrolysis by intracellular hexosaminidases. The thioglycosides reduce β-galactose incorporation into lactosamine chains, cell surface sialyl Lewis-X expression, and leukocyte rolling on selectin substrates including inflamed endothelial cells under fluid shear. Treatment of granulocytes with thioglycosides prior to infusion into mouse inhibited neutrophil homing to sites of acute inflammation and bone marrow by ∼80%-90%. Overall, thioglycosides represent an easy to synthesize class of efficient metabolic inhibitors or decoys. They reduce N-/O-linked glycan biosynthesis and inflammatory leukocyte accumulation., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Given the dependence of cancers on de novo lipogenesis, we tested the effect of fatostatin, a small molecule thought to target this pathway by blocking activation of SREBP transcription factors, in breast cancer cell lines and xenograft tumors. We found that estrogen receptor (ER) positive cells were more sensitive to fatostatin than ER negative cells and responded with cell cycle arrest and apoptosis. Surprisingly, we found that rather than inhibiting lipogenesis, fatostatin caused an accumulation of lipids as a response to endoplasmic reticulum stress rather than inhibition of SREBP activity. In particular, ceramide and dihydroceramide levels increased and contributed to the apoptotic effects of fatostatin. In addition, an accumulation of triacylglycerides (TAGs), particularly those containing polyunsaturated fatty acids (PUFAs), was also observed as a result of elevated diacylglycerol transferase activity. Blocking PUFA-TAG production enhanced the apoptotic effect of fatostatin, suggesting that these lipids play a protective role and limit fatostatin response. Together, these findings indicate that the ability of breast cancer cells to respond to fatostatin depends on induction of endoplasmic reticulum stress and subsequent ceramide accumulation, and that limiting production of PUFA-TAGs may be therapeutically beneficial in specific tumor subtypes.

Cellular senescence, the irreversible ceasing of cell division, has been associated with organismal aging, prevention of cancerogenesis, and developmental processes. As such, the evolutionary basis and biological features of cellular senescence remain a fascinating area of research. In this study, we conducted comparative RNAseq experiments to detect genes associated with replicative senescence in two different human fibroblast cell lines and at different time points. We identified 841 and 900 genes (core senescence-associated genes) that are significantly up- and downregulated in senescent cells, respectively, in both cell lines. Our functional enrichment analysis showed that downregulated core genes are primarily involved in cell cycle processes while upregulated core gene enrichment indicated various lipid-related processes. We further demonstrated that downregulated genes are significantly more conserved than upregulated genes. Using both transcriptomics and genetic variation data, we identified one of the upregulated, lipid metabolism genes, CD36, as an outlier. We found that overexpression of CD36 induces a senescence-like phenotype and, further, the media of CD36-overexpressing cells alone can induce a senescence-like phenotype in proliferating young cells. Moreover, we used a targeted lipidomics approach and showed that phosphatidylcholines accumulate during replicative senescence in these cells, suggesting that upregulation of CD36 could contribute to membrane remodeling during senescence. Overall, these results contribute to the understanding of evolution and biology of cellular senescence and identify several targets and questions for future studies.

Loxoprofen is an anti-inflammatory drug that requires bioactivation into the trans-OH metabolite to exert pharmacological activity. Evidence suggests that carbonyl reductase 1 (CBR1) is important during the bioactivation of loxoprofen. This study examined the impact of the functional single nucleotide polymorphism CBR1 rs9024 on the bioactivation of loxoprofen in a collection of human liver samples. The synthesis ratios of trans-OH loxoprofen/cis-OH loxoprofen were 33% higher in liver cytosols from donors homozygous for the CBR1 rs9024 G allele in comparison with the ratios in samples from donors with heterozygous GA genotypes. Complementary studies examined the impact of CBR1 rs9024 on the bioactivation of loxoprofen in lymphoblastoid cell lines. CBR1 rs9024 genotype status impacts the synthesis of the bioactive trans-OH metabolite of loxoprofen in human liver., (© 2018 John Wiley & Sons, Ltd.)

Although apoptosis has long dominated the spotlight, studies in the past two decades have expanded the repertoire of programmed cell death (PCD). Several forms of non-apoptotic regulated cell death have been identified, with important links to organismal homeostasis and different disease pathologies. Necroptosis, ferroptosis, pyroptosis, and NETosis are the major forms of PCD that have attracted attention. Clear biochemical distinctions differentiate these forms of non-apoptotic PCD at the protein and membrane levels. For instance, pore formation at the plasma membrane is a hallmark of necroptosis and pyroptosis; however, different proteins facilitate pore formation in these processes. Here, we will highlight the role of lipids in different forms of non-apoptotic PCD. In particular, we discuss how lipids can trigger or facilitate the membrane-related changes that result in cell death. We also highlight the use of small molecules in elucidating the mechanisms of non-apoptotic PCD and the potential of lipid biosynthetic pathways to perturb these processes for therapeutic applications as a future avenue of research.

Background: Microalgae are uniquely advantageous organisms cultured and harvested for several value-added biochemicals. A majority of these compounds are lipid-based, such as triacylglycerols (TAGs), which can be used for biofuel production, and their accumulation is most affected under nutrient stress conditions. As such, the balance between cellular homeostasis and lipid metabolism becomes more intricate to achieve efficiency in bioproduct synthesis. Lipidomics studies in microalgae are of great importance as biochemical diversity also plays a major role in lipid regulation among oleaginous species., Methods: The aim of this study was to analyze time-series changes in lipid families produced by microalga under different nutrient conditions and growth phases to gain comprehensive information at the cellular level. For this purpose, we worked with a highly adaptable, oleaginous, non-model green microalga species, Ettlia oleoabundans (a.k.a. Neochloris oleoabundans ). Using a mass spectrometry-based untargeted and targeted metabolomics' approach, we analyzed the changes in major lipid families under both replete and deplete nitrogen and phosphorus conditions at four different time points covering exponential and stationary growth phases., Results: Comprehensive analysis of the lipid metabolism highlighted the accumulation of TAGs, which can be utilized for the production of biodiesel via transesterification, and depletion of chlorophylls and certain structural lipids required for photosynthesis, under nutrient deprived conditions. We also found a correlation between the depletion of digalactosyldiacylglycerols (DGDGs) and sulfoquinovosyldiacylglycerols (SQDGs) under nutrient deprivation., Conclusions: High accumulation of TAGs under nutrient limitation as well as a depletion of other lipids of interest such as phosphatidylglycerols (PGs), DGDGs, SQDGs, and chlorophylls seem to be interconnected and related to the microalgal photosynthetic efficiency. Overall, our results provided key biochemical information on the lipid regulation and physiology of a non-model green microalga, along with optimization potential for biodiesel and other value-added product synthesis.

Lipids are a diverse class of biomolecules. The biosynthesis and transport of these molecules are controlled by a considerable number of proteins, which facilitate spatiotemporal regulation of lipids during different fundamental cellular processes. Although lipids are traditionally considered as molecules for energy storage and as structural components of membranes, they are being increasingly recognized for their signaling roles. There is a growing appreciation of lipids' chemical diversity, which approaches that of proteins. In this Perspective, we discuss recent studies that suggest novel functions for distinct lipid species during different cellular processes. In particular, we discuss findings from our laboratory that illuminate the involvement of ceramides, polyunsaturated triacylglycerols, and very long chain fatty acids in different cellular fates. We also highlight recent innovative methods that have enabled the recognition of previously unknown lipid classes and/or roles of these molecules in different biological processes. We envision that advances in lipid identification, visualization, and perturbation will pave the way for broader investigations into this fascinating and influential class of biomolecules.

Triacylglycerols (TAGs) are one of the major constituents of the glycerolipid family. Their main role in cells is to store excess fatty acids, and they are mostly found within lipid droplets. TAGs contain acyl chains that vary in length and degree of unsaturation, resulting in hundreds of chemically distinct species. We have previously reported that TAGs containing polyunsaturated fatty acyl chains (PUFA-TAGs) accumulate via activation of diacylglycerol acyltransferases during apoptosis. In this work, we show that accumulation of PUFA-TAGs is a general phenomenon during this process. We further show that the accumulated PUFA-TAGs are stored in lipid droplets. Because membrane-residing PUFA phospholipids can undergo oxidation and form reactive species under increased levels of oxidative stress, we hypothesized that incorporation of PUFAs into PUFA-TAGs and their localization within lipid droplets during apoptosis limit the toxicity during this process. Indeed, exogenous delivery of a polyunsaturated fatty acid resulted in a profound accumulation of PUFA phospholipids and rendered cells more sensitive to oxidative stress, causing reduced viability. Overall, our results support the concept that activation of TAG biosynthesis protects cells from lipid peroxide-induced membrane damage under increased levels of oxidative stress during apoptosis. As such, targeting triacylglycerol biosynthesis in cancer cells might represent a new approach to promoting cell death during apoptosis.

Necroptosis is a form of regulated cell death that is linked to various human diseases. Distinct membrane-related, thus lipid-dependent, alterations take place during necroptosis. However, little is known about the roles of specific lipids in this process. We used an untargeted LC-MS-based approach to reveal that distinct lipid species are regulated at the molecular level during necroptosis. We found that ceramides and very long chain fatty acids accumulate during this process. Intrigued by the specificity of very long chain fatty acid accumulation, we focused on characterizing their involvement during necroptosis. Biochemical characterizations suggested that activated fatty acid biosynthesis and elongation could be responsible for these accumulations. We further showed that inhibition of fatty acid biosynthesis and depletion of very long chain fatty acids prevented loss of plasma membrane integrity and cell death, strongly suggesting that very long chain fatty acids are functionally involved in necroptosis., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Cationic liposomes have been used for targeted drug delivery to tumor blood vessels, via mechanisms that are not fully elucidated. Doxorubicin (Dox)-loaded liposomes were prepared that incorporate a cationic lipid; 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), along with a small amount of porphyrin-phospholipid (PoP). Near-infrared (NIR) light caused release of entrapped Dox via PoP-mediated DOTAP photo-oxidation. The formulation was optimized to enable extremely rapid NIR light-triggered Dox release (i.e., in 15 seconds), while retaining reasonable serum stability. In vitro , cationic PoP liposomes readily bound to both MIA PaCa-2 human pancreatic cancer cells and human vascular endothelial cells. When administered intravenously, cationic PoP liposomes were cleared from circulation within minutes, with most accumulation in the liver and spleen. Fluorescence imaging revealed that some cationic PoP liposomes also localized at the tumor blood vessels. Compared with analogous neutral liposomes, strong tumor photoablation was induced with a single treatment of cationic PoP liposomes and laser irradiation (5 mg/kg Dox and 100 J/cm 2 NIR light). Unexpectedly, empty cationic PoP liposomes (lacking Dox) induced equally potent antitumor phototherapeutic effects as the drug loaded ones. A more balanced chemo- and phototherapeutic response was subsequently achieved when antitumor studies were repeated using higher drug dosing (7 mg/kg Dox) and a low fluence phototreatment (20 J/cm 2 NIR light). These results demonstrate the feasibility of vessel-targeted chemophototherapy using cationic PoP liposomes and also illustrate synergistic considerations. Mol Cancer Ther; 16(11); 2452-61. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Intratumoral (IT) drug injections reduce systemic toxicity, but delivered volumes and distribution can be inconsistent. To improve IT delivery paradigms, porphyrin-phospholipid (PoP) liposomes are passively loaded with three hydrophilic cargos: sulforhodamine B, a fluorophore; gadolinium-gadopentetic acid, a magnetic resonance (MR) agent; and oxaliplatin, a colorectal cancer chemotherapeutic. Liposome composition is optimized so that cargo is retained in serum and storage, but is released in less than 1 min with exposure to near infrared light. Light-triggered release occurs with PoP-induced photooxidation of unsaturated lipids and all cargos release concurrently. In subcutaneous murine colorectal tumors, drainage of released cargo is delayed when laser treatment occurs 24 h after IT injection, at doses orders of magnitude lower than systemic ones. Delayed light-triggering results in substantial tumor shrinkage relative to controls a week following treatment, although regrowth occurs subsequently. MR imaging reveals that over this time frame, pools of liposomes within the tumor migrate to adjacent regions, possibly leading to altered spatial distribution during triggered drug release. Although further characterization of cargo loading and release is required, this proof-of-principle study suggests that multimodal theranostic IT delivery approaches hold potential to both guide injections and interpret outcomes, in particular when combined with chemo-phototherapy., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Cellular replicative senescence, a state of permanent cell-cycle arrest, has been linked to organismal aging, tissue repair and tumorigenesis. In this study, we comparatively investigated the global lipid profiles and mRNA content of proliferating and senescent-state BJ fibroblasts. We found that both expression levels of lipid-regulating genes and the abundance of specific lipid families, are actively regulated. We further found that 19 specific polyunsaturated triacylglycerol species constituted the most prominent changes in lipid composition during replicative senescence. Based on the transcriptome analysis, we propose that the activation of CD36-mediated fatty acid uptake and diversion to glycerolipid biosynthesis could be responsible for the accumulation of triacylglycerols during replicative senescence. This, in turn, could be a cellular mechanism to prevent lipotoxicity under increased oxidative stress conditions observed in this process. Our results indicate that regulation of specific lipid species has a central role during replicative senescence.

Lipids are emerging as key regulators of fundamental cellular processes including cell survival, division, and death. Apoptosis, a form of programmed cell death, is accompanied by numerous membrane-related phenotypic changes. However, we have an incomplete understanding of the involvement of specific lipid structures during this process. Here, we report that triacylglycerols are regulated at the molecular level during 5-fluorouracil-induced apoptosis in HCT-116. Mass-spectrometry-based global lipid profiling shows that specific triacylglycerols accumulate during apoptosis. Expression levels and activities of enzymes that are responsible for the biosynthesis and metabolic processing of triacylglycerols suggest that triacylglycerol biosynthesis is responsible for these accumulations. Based on our data, we propose that regulation of triacylglycerols at the molecular level happens downstream of p53 activation and potentially is a mechanism to prevent lipid oxidation during apoptosis.

Prompt membrane permeabilization is a requisite for liposomes designed for local stimuli-induced intravascular release of therapeutic payloads. Incorporation of a small amount (i.e., 5 molar percent) of an unsaturated phospholipid, such as dioleoylphosphatidylcholine (DOPC), accelerates near infrared (NIR) light-triggered doxorubicin release in porphyrin-phospholipid (PoP) liposomes by an order of magnitude. In physiological conditions in vitro, the loaded drug can be released in a minute under NIR irradiation, while liposomes maintain serum stability otherwise. This enables rapid laser-induced drug release using remarkably low amounts of PoP (i.e., 0.3 molar percent). Light-triggered drug release occurs concomitantly with DOPC and cholesterol oxidation, as detected by mass spectrometry. In the presence of an oxygen scavenger or an antioxidant, light-triggered drug release is inhibited, suggesting that the mechanism is related to singlet oxygen mediated oxidization of unsaturated lipids. Despite the irreversible modification of lipid composition, DOPC-containing PoP liposome permeabilization is transient. Human pancreatic xenograft growth in mice is significantly delayed with a single chemophototherapy treatment following intravenous administration of 6 mg kg(-1) doxorubicin, loaded in liposomes containing small amounts of DOPC and PoP., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)