Neutral Lipid Accumulation Research Articles

Alterations in cellular lipid metabolism produce neutral lipid accumulation following exposure to the organochlorine compound trans-nonachlor in rat primary hepatocytes.

Recent epidemiological studies have revealed significant positive associations between exposure to organochlorine (OC) pesticides and occurrence of the metabolic syndrome and there are a growing number of animal-based studies to support causality. However, the cellular mechanisms linking OC compound exposure and metabolic dysfunction remain elusive. Therefore, the present study was designed to determine if direct exposure to three highly implicated OC compounds promoted hepatic steatosis, the hepatic ramification of the metabolic syndrome. First, the steatotic effect of p,p'-dichlorodiphenyldichloroethylene (DDE), oxychlordane, and trans-nonachlor was determined in freshly isolated rat primary hepatocytes. Exposure to trans-nonachlor significantly increased neutral lipid accumulation as opposed to DDE and oxychlordane. To determine possible mechanisms governing increased fatty acid availability, the effects of trans-nonachlor exposure on fatty acid uptake, de novo lipogenesis, triglyceride secretion, and fatty acid oxidation were explored. Trans-nonachlor did not significantly alter fatty acid uptake. However, insulin-stimulated de novo lipogenesis as well as basal expression of fatty acid synthase, a major regulator of lipogenesis were significantly increased following trans-nonachlor exposure. Interestingly, there was a significant decrease in fatty acid oxidation following trans-nonachlor exposure. This decrease in fatty acid oxidation was accompanied by a slight, but significant increase in oleic acid-induced cellular triglyceride secretion. Therefore, taken together, the present data indicate direct exposure to trans-nonachlor has a more potent pro-steatotic effect than exposure to DDE or oxychlordane. This pro-steatotic effect of trans-nonachlor appears to be predominately mediated via increased de novo lipogenesis and decreased fatty acid oxidation.

Systems-level analysis of metabolic mechanism following nitrogen limitation in benthic dinoflagellate Prorocentrum lima

Prorocentrum lima is a typical diarrheic shellfish poisoning (DSP) toxin-producing benthic dinoflagellate usually found attached to or associated with macrophytes, floating detritus, debris, or other substrates. In this paper, to gain a deeper understanding of the molecular and cellular responses to N limitation in P. lima, transcriptome profiling was performed. We found that most genes related to photosynthesis, porphyrin and chlorophyll metabolism were down-regulated following N limitation, while some genes concerning fatty acid biosynthesis, starch synthesis and ubiquitin pathway were up-regulated. Meanwhile, ABC transports ABCB1 and ABCG2 and glutathione S-transferase were up-regulated under N-limited conditions. These transcriptomic data were further corroborated by the biochemical analyses which revealed that starch, lipid content was increased, while photosynthetic efficiency was decreased. In addition, TEM analysis demonstrated that chloroplasts appeared smaller and were less abundant, while a major fraction of the cell volume was occupied by lipid bodies and starch granules, and thylakoid was twisted in the cells. Taking together, we proposed that N limitation could induce the accumulation of neutral lipid and starch in P. lima cells for carbon fixation trough recycling chloroplast membranes by autophagy. ABCB and ABCG transporters might be involved in the transport of DSP toxins. Our findings might provide global information for the response or adaptation of P. lima to N limitation.

NDRG1 regulates neutral lipid metabolism in breast cancer cells

BackgroundAltered lipid metabolism is an emerging hallmark of aggressive breast cancers. The N-myc downstream regulated gene (NDRG1) gene plays a critical role in peripheral nervous system myelination, as inactivating mutations cause severe demyelinating neuropathy. In breast cancer, elevated NDRG1 expression has been linked to clinical outcomes, but its functional role in breast cancer physiology has remained unclear.MethodsA meta-analysis of NDRG1 expression in multiple large publicly available genomic databases was conducted. Genome-wide expression correlation and Cox proportional hazards and Kaplan-Meier modeling of clinical outcomes associated with elevated expression were assessed. To study NDRG1 function, gene silencing and overexpression phenotypic studies were carried out in a panel of cell lines representing all major breast cancer molecular subtypes. Changes in cell proliferation, morphology, and neutral lipid accumulation due to altered NDRG1 expression were assessed by high throughput, quantitative microscopy. Comprehensive lipidomics mass spectrometry was applied to characterize global changes in lipid species due to NDRG1 silencing. Labeled fatty acids were used to monitor cellular fatty acid uptake and subcellular distribution under nutrient replete and starvation culture conditions.ResultsNDRG1 overexpression correlated with glycolytic and hypoxia-associated gene expression, and was associated with elevated rates of metastasis and patient mortality. Silencing NDRG1 reduced cell proliferation rates, causing lipid metabolism dysfunction including increased fatty acid incorporation into neutral lipids and lipid droplets. Conversely, NDRG1 expression minimized lipid droplet formation under nutrient replete and starvation conditions.ConclusionsHere we report that NDRG1 contributes to breast cancer aggressiveness by regulating the fate of lipids in cells that exhibit an altered lipid metabolic phenotype. In line with its role in promoting myelination and its association with altered metabolism in cancer, our findings show that NDRG1 is a critical regulator of lipid fate in breast cancer cells. The association between NDRG1 and poor prognosis in breast cancer suggests it should play a more prominent role in patient risk assessment. The function of NDRG1 in breast cancer lipid metabolism may represent a promising therapeutic approach in the future.

Clinical and genetic characterization of a Chanarin Dorfman Syndrome patient born to diseased parents

BackgroundChanarin Dorfman Syndrome (CDS) is a rare autosomal recessive disorder characterized by ichthyosiform non-bullous erythroderma and variable involvement of the liver and the neuromuscular system. In CDS patients, the accumulation of neutral lipids inside cytoplasmic lipid droplets has been demonstrated in different tissues. To date, ninety families with this disease have been described worldwide; most of them are from Mediterranean countries.Case presentationIn this report, we describe a consanguineous Turkish family with typical features of CDS. The parents are first cousins and are both diseased. At the age of eight, their child presented CDS with non-bullous congenital ichthyosiform erythroderma, hepatosteatosis, hepatomegaly and ectropion. Electromyographic examination is compatible with myopathy. A five-year-old cousin of the child is also affected by CDS. She was born to non-affected consanguineous parents. Mutation analysis of the ABHD5 gene revealed the previously reported mutation, N209X, which is the most frequent in Turkish patients. Lipid vacuoles, also known as Jordan’s anomaly, are detectable in their leucocytes.ConclusionsTo the best of our knowledge, this is the first report of a CDS family in which both parents and their child are affected by CDS. To date, the child does not present a more severe clinical phenotype compared with those of his relatives or other CDS patients of the same age. These findings suggest that high levels of triacylglycerol accumulation, that may be supposed to be present in high amount inside the ooplasm, did not affect embryo development and foetal growth.

Effects of fundamental nutrient stresses on the lipid accumulation profiles in two diatom species Thalassiosira weissflogii and Chaetoceros muelleri.

Microalgae are considered as attractive feedstocks for biofuel production nowadays because of their high lipid contents and easy cultivation. In the present study, two diatoms, Thalassiosira weissflogii and Chaetoceros muelleri, were cultured under various nutrient-limitation conditions to explore their comprehensive lipid accumulation profiles for further commercialization. In T. weissflogii, the highest neutral lipid accumulation and highest lipid productivity (14.28mgL-1day-1) were both recorded under P-limitation. In C. muelleri, the highest lipid content (35.03% of dry cell weight), highest neutral lipid accumulation, and highest lipid productivity (29.07mgL-1day-1) were all recorded under N-limitation. Besides, the predominant fatty acids of T. weissflogii and C. muelleri were myristic acid (C14:0), palmitic acid (C16:0), and palmitoleic acid (C16:1), with the amounts of 58.4-74.4 and 74.1-87.7% of the total fatty acids, respectively. Moreover, nutrient limitations led to a lower proportion of polyunsaturated fatty acids (PUFA) than that of saturated fatty acid (SFA) and monounsaturated fatty acid (MUFA) in both species. The ratios of (SFA + MUFA) to PUFA were from 1.65 to 3.01 in T. weissflogii, and up to 3.61 to 8.59 in C. muelleri. Our results suggested the feasibility of C. muelleri as biodiesel feedstock due to its more suitable fatty acid composition and higher lipid productivity compared to T. weissflogii.

Molecular characterization of freshwater microalgae and nutritional exploration to enhance their lipid yield.

Biomass and lipid yield under nutrient depleted and supplemented conditions of N and Mg in two freshwater isolates, Chlorella sp. NC-MKM and Scenedesmus acutus NC-M2 from Meghalaya were investigated for biodiesel production. Both the strains, Chlorella sp. NC-MKM and Scenedesmus acutus NC-M2 are oleaginous in nature having lipid content of 40.2 ± 1.4 and 37.3 ± 2.6% DCW, respectively. The significant increase (92.8%) in lipid content was found in N-depleted condition while an increase (46.65%) in biomass yield was observed under Mg-supplemented condition in Chlorella sp. NC-MKM. Studying the interactive effects of nutrient depletion and supplementation, combination of N-depleted and Mg-supplemented condition was selected for further investigation to check enhanced lipid yield in Chlorella sp. NC-MKM. The results showed a significant increase in biomass yield, lipid yield and lipid content (30, 66.8, and 28.66%, respectively). Under this condition, accumulation of neutral lipid was also enhanced (47.17% M2 gated cells) compared to control (21.37% M2 gated cells). Further, FAMEs revealed that the relative percentage of saturated and mono-unsaturated fatty acids increased (66.16%) in Chlorella sp. NC-MKM compared to control that improves biodiesel properties.

F41. Expanding the phenotype-genotype correlations of PNPLA2 mutations

Introduction Neutral lipid storage disease with myopathy (NLSD-M), due to diverse PNPLA2 mutations, usually presents with an asymmetrical weakness, proximal involvement of upper limbs and posterior compartment of lower limbs. NLSD-M can be associated with other co-morbidities, like diabetes mellitus, hyperlipidemia, and most commonly, cardiomyopathy, which can be detected in almost 50% of patients. Although clinical manifestations begin around the third decade, late onset disease with unusual symptoms, such as fatigue and exercise intolerance, has been reported. Magnetic resonance imaging (MRI) of skeletal muscles reveals fatty degeneration, with a predilection for shoulder girdle and lower limb muscles. Electromyographic (EMG) features have been reported in one patient, where complex repetitive discharges (CRD) and myotonic discharges were detected in a few muscles. Methods Description of the clinical, neurophysiological, muscle MRI, pathological and genetic findings of a novel mutation in the PNPLA2 gene. This work received ethics board approval. Results Two sisters (aged 31 years at first examination), born 3 years apart from consanguineous parents, presented with a 2 year history of progressive motor deficits, proximal/asymmetric in upper limbs and distal/symmetric in lower limbs. The most severely affected muscles were: deltoids, biceps, external cuff rotators, gastrocnemius and tibialis anterior muscles. EMG recordings showed diffuse short-duration motor unit potentials, with asymmetrical distribution; some long duration, polyphasic potentials in distal muscles; profuse CRD in the biceps and tibialis anterior and sparse CRD in paravertebral muscles. MRI revealed an asymmetrical fatty replacement in the posterior compartment of thigs and legs, paravertebral musculature, deltoids and biceps. Jordans’ anomaly, characterized by lipid accumulation in leukocytes, was detected in peripheral blood smears in both patients. Muscle biopsy showed cytoplasmic vacuoles, suggestive of neutral lipid accumulation. Genetic analysis revealed a homozygous deletion in exon 6 of PNPLA2 gene (c.995delG), not yet described in the literature. Five years after the initial symptoms, the elder sister was completely dependent for upper limb activities, although she had independent walking, while the younger sister had only minor fine motor skills impairment. Creatine phosphokinase levels increased in a stepwise manner over the years in the elder sister (97–2840 UI/L), while stable levels were found in the younger one (621–681 UI/L). Cardiomyopathy was not present in those patients. Conclusion NLSD-M due to PNPLA2 gene mutations should be suspected in patients presenting with an asymmetrical myopathy with upper limb predominance; profuse and also asymmetrical EMG and MRI abnormalities, even without cardiomyopathy. Although some mutations have been described, genotypical variability seems to be common, so novel mutations should be sought for genetic counselling purposes.

“Trans-nonachlor increases extracellular free fatty acid accumulation and de novo lipogenesis to produce hepatic steatosis in McArdle-RH7777 cells”

Recent studies suggest there may be an environmental exposure component to the development and progression of non-alcoholic fatty liver disease (NAFLD) involving the organochlorine (OC) pesticides or their metabolites. However, the roles of OC compounds in the development of NAFLD has not been fully elucidated. Therefore, the current study was designed to determine if exposure to trans-nonachlor, a prevalent OC compound, could promote hepatocyte lipid accumulation and determine potential pro-steatotic mechanisms. McArdle-RH7777 (McA) hepatoma cells were incubated with trans-nonachlor for 24 h then neutral lipid accumulation was determined by Oil Red O staining. Exposure to trans-nonachlor produced a concentration dependent increase in neutral lipid accumulation. Trans-nonachlor also increased extracellular free fatty acid-induced neutral lipid accumulation which appears to be due at least in part to increased free fatty acid accumulation as evident by increased accumulation of Bodipy labeled dodecanoic acid. Additionally, 14C-acetate incorporation into total cellular lipids was increased by trans-nonachlor implicating increased de novo lipogenesis (DNL) as a potential mediator of trans-nonachlor-induced neutral lipid accumulation. Taken together, the present data indicate exposure to trans-nonachlor has a direct, pro-steatotic effect on hepatocytes to increase lipid accumulation through the combinatorial actions of extracellular free fatty acid accumulation and increased DNL.

Dietary Calcium and Phosphate levels affect bone development and mesenchymal stem cell lineage allocation in neonatal pigs

Deficits in bone deposition during early‐life have been associated with increased risk of osteoporosis later in life. Bone marrow mesenchymal stem cells (MSC) are a subset of multipotent stem cells residing within the bone which provide the life‐long supply of bone forming cells. Molecular mechanisms affecting the activity and differentiation potential of MSC are still poorly understood. Our previous work has revealed that the activity and differentiation potential of MSC are subject to alteration based on neonatal nutrition. Dietary calcium (Ca) and phosphate (PO4) levels play a critical role in regulation of bone development. With increasing interest in MSC therapies and an increasing incidence of bone diseases, we aimed to elucidate the impact of marginal dietary Ca and PO4 deficiency and excess on the activity and differentiation potential of MSC during neonatal development. During a 16‐day study, 27 neonatal pigs (n= 27, 24±6h old) were fed milk replacers that were isonitrogenous and isocaloric, but either 25% deficient (D), adequate (A), or 25% in excess (E) of Ca and PO4 requirements. Piglets were fed 8 times/d at a rate to match the growth rate of sow‐reared piglets. Body weight and feed intake were recorded daily. Blood was collected on d8 and d16 for analysis. Humeri were collected for the isolation of MSC and radial/ulna bones were collected for physical measurement and determination of bone mineral content. Serum PO4 concentrations were significantly higher in the A group compared to the D group on day 8, however at day 16 the D group had the highest serum PO4 concentrations with the E group having the lowest PO4 concentrations (P<0.05). Serum FGF23 was greater in the D group than the E group at 16d (77.21±1.90 pg/mL vs. 70.38±1.90 pg/mL, P<0.05), with values for the A group being intermediate to these groups. The D group had lower bone mineral content and bone volume as compared to the E group. Among the MSC, those isolated from D pigs had significantly lower rates of in vivo proliferation of than those isolated from A pigs, as determined by BrdU labelling (0.063±0.007 vs. 0.089±0.007), but significantly higher in vitro proliferation than those isolated from E pigs at both 12 and 24 h post‐plating (12 h: 0.40 ±0.04 vs 0.23±0.04; 24h: 0.28±0.04 vs 0.10±004). Ca and PO4 deficiency appears, based on the gene expression of key adipogenic factors as well as cellular accumulation of neutral lipids, to have predisposed MSC towards adipogenic differentiation. MSC from D pigs had increased expression of PPARγ2, FABP4, and LPL (P<0.05) and greater accumulation of neutral lipids as indicated by Oil Red O staining. Coupled with this apparent predisposition for adipocytic differentiation, a decrease in osteogenic capabilities were seen among MSC isolated from D pigs. This was determined based on reduced expression of the osteogenic regulators osteocalcin and fibronectin at d6 post differentiation. These findings demonstrated that even minor dietary deficiencies of Ca and PO4 during the neonatal period can have a profound impact on bone development. These dietary deficiencies may also program future bone health by altering the differentiation capabilities of MSC.Support or Funding InformationThis project is support by USDA grantThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Developing a piggyBac Transposon System and Compatible Selection Markers for Insertional Mutagenesis and Genome Engineering in Yarrowia lipolytica.

Yarrowia lipolytica is a non-conventional yeast of interest to the biotechnology industry. However, the physiology, metabolism, and genetic regulation of Y. lipolytica diverge significantly from more well-studied and characterized yeasts such as Saccharomyces cerevisiae. To develop additional genetic tools for this industrially relevant host, the piggyBac transposon system to enable efficient generation of genome-wide insertional mutagenesis libraries and introduction of scarless, footprint-free genomic modifications in Y. lipolytica. Specifically, we demonstrate piggyBac transposition in Y. lipolytica, and then use the approach to screen transposon insertion libraries for rapid isolation of mutations that confer altered canavanine resistance, pigment formation, and neutral lipid accumulation. We also develop a variety of piggyBac compatible selection markers for footprint-free genome engineering, including a novel dominant marker cassette (Escherichia coli guaB) for effective Y. lipolytica selection using mycophenolic acid. We utilize these marker cassettes to construct a piggyBac vector set that allows for auxotrophic selection (uracil or tryptophan biosynthesis) or dominant selection (hygromycin, nourseothricin, chlorimuron ethyl, or mycophenolic acid resistance) and subsequent marker excision. These new genetic tools and techniques will help to facilitate and accelerate the engineering of Y. lipolytica strains for efficient and sustainable production of a wide variety of small molecules and proteins.

Lysosomal acid lipase promotes cholesterol ester metabolism and drives clear cell renal cell carcinoma progression.

Clear cell renal cell carcinoma (ccRCC) is characterized histologically by accumulation of cholesterol esters, cholesterol and other neutral lipids. Lysosomal acid lipase (LAL) is a critical enzyme involved in the cholesterol ester metabolism. Here, we sought to determine whether LAL could orchestrate metabolism of cholesterol esters in order to promote ccRCC progression. Quantitative reverse-transcription PCR and western blots were conducted to assess the expression of LAL in human ccRCC tissues. We analysed the relationship between LAL levels and patient survival using tissue microarrays. We used cell proliferation assays, colony formation assays, cell death assays, metabolic assays and xenograft tumour models to evaluate the biological function and underlying mechanisms. LAL was up-regulated in ccRCC tissue. Tissue microarray analysis revealed higher levels of LAL in advanced grades of ccRCC, and high LAL expression indicated lower patient survival. Suppressing LAL expression not only blocked the utilization of cholesterol esters but also impaired proliferation and cellular survival. Furthermore, immunohistochemistry staining showed that LAL expression was correlated with Akt phosphorylation. Suppressing LAL expression decreased the phosphorylation level of Akt and Src and reduced the level of 14,15-epoxyeicosatrienoic acids in ccRCC cells. Supplement of 14,15-epoxyeicosatrienoic acids rescued proliferation in vitro and in vivo. LAL promoted cell proliferation and survival via metabolism of epoxyeicosatrienoic acids and activation of the Src/Akt pathway.

Effects of dietary Pb and Cd and their combination on lysosomal and tissue-level biomarkers and histopathology in digestive gland of the land snail, Cantareus apertus (Born, 1778)

The present study was aimed at determining cell and tissue-level biomarkers and histopathological alterations in the green garden snail, Cantareus apertus (Born, 1778), exposed to different nominal dietary concentrations of Pb (25, 100 and 2500 mg Pb/kg), Cd (5, 10 and 100 mg Cd/kg) and their combination (25 mg Pb + 5 mg Cd/Kg, 100 mg Pb + 10 mg Cd/kg and 2500 mg Pb + 100 mg Cd/ kg) for 1 and 8 weeks. Lead and Cd exerted histopathological effects on the digestive gland in a dose-dependent manner and related to lysosomal and tissue-level biomarkers. The biological responses observed included digestive cell vacuolisation and numerical atrophy, calcium cell hydropic degeneration, excretory cell hypertrophy, inflammatory responses, blood vessel congestion, and disruption of the blood vessel wall and the interstitial connective tissue. Lysosomal enlargement and transient intracellular accumulation of neutral lipids and lipofuscins were also observed, together with alterations in the cell type composition and thinning of the digestive gland epithelium and with diverticular distortion. This response profile fits well with the biological effects reported after metal exposure in gastropods from other regions, as well as with data obtained in parallel studies dealing with metal bioaccumulation and intralysosomal accumulation, mortality, feeding, growth, oxidative stress and neurotoxicity exerted elicited by Pb, Cd and their mixture in green garden snails under the present experimental conditions. Consequently, C. apertus seems to be a suitable model species for the biomarker-based assessment of the biological effects of Pb and Cd, alone or in combinations, thus providing a challenging opportunity to advance in identifying suitable sentinel species for metal pollution biomonitoring and ecosystem health assessment in soil ecosystems in Northern Africa.

De novo transcriptome analysis and gene expression profiling of an oleaginous microalga Scenedesmus acutus TISTR8540 during nitrogen deprivation-induced lipid accumulation

Nitrogen deprivation (−N) has been used as a technique to promote lipid accumulation in various microalgae. Scenedesmus acutus is a promising oleaginous green microalga that can be cultivated in organic wastewater for biodiesel production. Nevertheless, the molecular mechanisms controlling S. acutus lipid accumulation in response to −N remain unidentified. Physiological study determined that −N reduced cell growth and photosynthetic pigments. On the other hand, it promoted carbohydrate and neutral lipid accumulation. To find the mechanisms underlying lipid accumulation, we performed de novo transcriptome profiling of the non-model S. acutus in response to −N. The transcriptome analysis revealed that glycolysis and starch degradation were up-regulated; on the contrary, gluconeogenesis, photosynthesis, triacylglycerol (TAG) degradation and starch synthesis were down-regulated by −N. Under −N, the carbon flux was shifted toward fatty acid and TAG synthesis, and the down regulation of TAG lipase genes may contribute to TAG accumulation. A comparative analysis of the −N transcriptomes of oleaginous microalgae identified that the down-regulation of multiple lipase genes was a specific mechanism found only in the −N transcriptome of S. acutus. Our study unraveled the mechanisms controlling −N-induced lipid accumulation in S. acutus, and provided new perspectives for the genetic manipulation of biodiesel-producing microalgae.

Effect of brimonidine, an α2 adrenergic agonist, on human meibomian gland epithelial cells

We recently discovered that the anti-glaucoma pharmaceuticals timolol, a β adrenergic antagonist, and pilocarpine, a cholinergic compound, negatively influence the morphology, proliferative capacity and survival of human meibomian gland epithelial cells (HMGECs). We hypothesize that another class of anti-glaucoma drugs, the α2 adrenergic agonists, also acts directly on HMGECs to affect their structure and function. We tested this hypothesis. Immortalized (i) HMGECs were cultured with brimonidine, as well as clonidine (α2 agonist), phenylephrine (α1 agonist), RX821002 (inverse α2 agonist) and MK912 (neutral α2 agonist) for up to 7 days. Cells were counted with a hemocytometer, and evaluated for morphology, signaling pathway activity, protein biomarker expression, and the accumulation of neutral lipids, phospholipids and lysosomes. Our findings demonstrate that brimondine treatment induces a dose-dependent decrease in Akt signaling and proliferation of iHMGECs. In contrast, brimonidine also promotes a dose-dependent differentiation of iHMGECs, including an increase in neutral lipid, phospholipid and lysosome levels. These effects were paralleled by an inhibition of p38 signaling, and duplicated by cellular exposure to clonidine, but not phenylephrine. Brimonidine also enhanced the cellular content of sterol regulatory binding protein-1, a master regulator of lipid synthesis. Of particular interest, the putative α2 antagonists, RX821002 and MK912, did not interfere with brimonidine action, but rather stimulated IHMGEC differentiation. Our results support our hypothesis and demonstrate that α2 adrenergic agonists act directly on iHMGECs. However, these compounds do not elicit an overall negative effect. Rather, the α2 agonists promote the differentiation of iHMGECs.

Lipid accumulation and CO2 utilization of two marine oil-rich microalgal strains in response to CO2 aeration

Biological CO2 sequestration by microalgae is a promising and environmentally friendly technology applied to sequester CO2. The characteristics of neutral lipid accumulation by two marine oil-rich microalgal strains, namely, Isochrysis galbana and Nannochloropsis sp., through CO2 enrichment cultivation were investigated in this study. The optimum culture conditions of the two microalgal strains are 10% CO2 and f medium. The maximum biomass productivity, total lipid content, maximum lipid productivity, carbon content, and CO2 fixation ability of the two microalgal strains were obtained. The corresponding parameters of the two strains were as follows: ((142.42±4.58) g/(m2·d), (149.92±1.80) g/(m2·d)), ((39.95±0.77)%, (37.91±0.58)%), ((84.47±1.56) g/(m2·d), (89.90±1.98) g/(m2·d)), ((45.98±1.75)%, (46.88±2.01)%), and ((33.74±1.65) g/(m2·d), (34.08±1.32) g/(m2·d)). Results indicated that the two marine microalgal strains with high CO2 fixation ability are potential strains for marine biodiesel development coupled with CO2 emission reduction.

Homologous expression of lipid droplet protein-enhanced neutral lipid accumulation in the marine diatom Phaeodactylum tricornutum

Lipid droplets are ubiquitous cellular compartments that store neutral lipids and specific proteins localize on their surface. These proteins work as a scaffold in maintaining the lipid droplet structure or as regulators of lipogenesis or lipolysis. Previously, the most abundant lipid droplet protein, namely stramenopile-type lipid droplet protein (StLDP), was identified in the marine diatom Phaeodactylum tricornutum; however, its function remains unclear because StLDP does not reveal homology with known lipid droplet proteins and lacks a predictable domain. In this study, P. tricornutum was transformed to express a homologous StLDP gene under an fcpA promoter in order to determine its function. StLDP expression was strongly enhanced in the mutant (H8), especially in nitrogen-sufficient conditions; however, it was attenuated in nitrogen-deficient conditions. Despite the strong expression, no significant difference was observed in the lipid composition between the wild type (WT) and H8 under nitrogen-sufficient conditions. After cultivation in nitrogen-free medium for 6 days, neutral lipid content significantly increased in H8 than in WT. After 2 days of cultivation in nitrogen-free medium, 97.0% of single cells in WT formed one or two lipid droplets, whereas in H8, this proportion decreased to 78.8%, and the proportion of cells forming three or four lipid droplets increased. Thus, the function of StLDP was speculated to sequester triacylglycerol on the initial lipid droplet formation.

Mechanistic insight into ROS and neutral lipid alteration induced toxicity in the human model with fins (Danio rerio) by industrially synthesized titanium dioxide nanoparticles.

The toxicological impact of TiO2 nanoparticles on the environment and human health has been extensively studied in the last few decades, but the mechanistic details were unknown. In this study, we evaluated the impact of industrially prepared TiO2 nanoparticles on the biological system using zebrafish embryo as an in vivo model. The industrial synthesis of TiO2 nanoparticles was mimicked on the lab scale using the high energy ball milling (HEBM) method by milling bulk TiO2 particles for 5 h, 10 h, and 15 h in an ambient environment. The physiochemical properties were characterized by standard methods like field emission scanning electron microscopy (FESEM), dynamic light scattering (DLS), X-ray diffraction (XRD) and UV-Visible spectroscopy. In vivo cytotoxicity was assessed on zebrafish embryos by the evaluation of their mortality rate and hatching rate. Experimental and computational analysis of reactive oxygen species (ROS) induction, apoptosis, and neutral lipid alteration was done to study the effects on the cellular level of zebrafish larvae. The analysis depicted the change in size and surface charge of TiO2 nanoparticles with respect to the increase in milling time. In silico investigations revealed the significant role of ROS quenching and altered neutral lipid accumulation functionalised by the molecular interaction of respective metabolic proteins in the cytotoxicity of TiO2 nanoparticles with zebrafish embryos. The results reveal the hidden effect of industrially synthesized TiO2 nanoparticle exposure on the alteration of lipid accumulation and ROS in developing zebrafish embryos. Moreover, the assessment provided a detailed mechanistic analysis of in vivo cytotoxicity at the molecular level.

Zanthoxylum ailanthoides Suppresses Oleic Acid-Induced Lipid Accumulation through an Activation of LKB1/AMPK Pathway in HepG2 Cells.

Zanthoxylum ailanthoides (ZA) has been used as folk medicines in East Asian and recently reported to have several bioactivity; however, the studies of ZA on the regulation of triacylglycerol (TG) biosynthesis have not been elucidated yet. In this study, we examined whether the methanol extract of ZA (ZA-M) could reduce oleic acid- (OA-) induced intracellular lipid accumulation and confirmed its mode of action in HepG2 cells. ZA-M was shown to promote the phosphorylation of AMPK and its upstream LKB1, followed by reduction of lipogenic gene expressions. As a result, treatment of ZA-M blocked de novo TG biosynthesis and subsequently mitigated intracellular neutral lipid accumulation in HepG2 cells. ZA-M also inhibited OA-induced production of reactive oxygen species (ROS) and TNF-α, suggesting that ZA-M possess the anti-inflammatory feature in fatty acid over accumulated condition. Taken together, these results suggest that ZA-M attenuates OA-induced lipid accumulation and inflammation through the activation of LKB1/AMPK signaling pathway in HepG2 cells.

Metabolic Heterogeneity Evidenced by MRS among Patient-Derived Glioblastoma Multiforme Stem-Like Cells Accounts for Cell Clustering and Different Responses to Drugs.

Clustering of patient-derived glioma stem-like cells (GSCs) through unsupervised analysis of metabolites detected by magnetic resonance spectroscopy (MRS) evidenced three subgroups, namely clusters 1a and 1b, with high intergroup similarity and neural fingerprints, and cluster 2, with a metabolism typical of commercial tumor lines. In addition, subclones generated by the same GSC line showed different metabolic phenotypes. Aerobic glycolysis prevailed in cluster 2 cells as demonstrated by higher lactate production compared to cluster 1 cells. Oligomycin, a mitochondrial ATPase inhibitor, induced high lactate extrusion only in cluster 1 cells, where it produced neutral lipid accumulation detected as mobile lipid signals by MRS and lipid droplets by confocal microscopy. These results indicate a relevant role of mitochondrial fatty acid oxidation for energy production in GSCs. On the other hand, further metabolic differences, likely accounting for different therapy responsiveness observed after etomoxir treatment, suggest that caution must be used in considering patient treatment with mitochondria FAO blockers. Metabolomics and metabolic profiling may contribute to discover new diagnostic or prognostic biomarkers to be used for personalized therapies.

The Effect of Solithromycin, a Cationic Amphiphilic Drug, on the Proliferation and Differentiation of Human Meibomian Gland Epithelial Cells

ABSTRACTPurpose: We previously discovered that azithromycin (AZM) acts directly on immortalized human meibomian gland epithelial cells (IHMGECs) to stimulate their lipid and lysosome accumulation and overall differentiation. We hypothesize that this phospholipidosis-like effect is due to AZM’s cationic amphiphilic drug (CAD) nature. If our hypothesis is correct, then other CADs (e.g., solithromycin [SOL]) should be able to duplicate AZM’s action on IHMGECs. Our purpose was to test this hypothesis.Materials and Methods: IHMGECs were cultured in the presence of vehicle or SOL (2, 10, or 20 µg/ml) for up to 7 days under proliferating or differentiating conditions. Positive (epidermal growth factor and bovine pituitary extract for proliferation; AZM for differentiation) and negative (vehicle) controls were included with the experiments. IHMGECs were evaluated for cell number, neutral lipid content, and lysosome accumulation.Results: Our results demonstrate that SOL induces a rapid and dose-dependent increase in the accumulation of neutral lipids and lysosomes in HMGECs. The lysosomal effects were most prominent with the 10 and 20 µg/ml doses, and occurred earlier (i.e., 1 day) with SOL than with the AZM (10 µg/ml) control. The effects of SOL and AZM on IHMGEC differentiation were essentially the same after 3 days of culture. SOL did not influence the proliferation of HMGECs during a 7-day time period.Conclusions: Our results support our hypothesis that SOL, a CAD, is able to reproduce AZM’s impact on lysosome and lipid accumulation, as well as the differentiation, of HMGECs. The effect of SOL on lysosome appearance was faster than that of AZM.