Lipid droplets in the nervous system:involvement in cell metabolic homeostasis

Lipid droplets serve as primary storage organelles for neutral lipids in neurons,glial cells,and other cells in the nervous system.Lipid droplet formation begins with the synthesis of neutral lipids in the endoplasmic reticulum.Previously,lipid droplets were recognized for their role in maintaining lipid metabolism and energy homeostasis;however,recent research has shown that lipid droplets are highly adaptive organelles with diverse functions in the nervous system.In addition to their role in regulating cell metabolism,lipid droplets play a protective role in various cellular stress responses.Furthermore,lipid droplets exhibit specific functions in neurons and glial cells.Dysregulation of lipid droplet formation leads to cellular dysfunction,metabolic abnormalities,and nervous system diseases.This review aims to provide an overview of the role of lipid droplets in the nervous system,covering topics such as biogenesis,cellular specificity,and functions.Additionally,it will explore the association between lipid droplets and neurodegenerative disorders.Understanding the involvement of lipid droplets in cell metabolic homeostasis related to the nervous system is crucial to determine the underlying causes and in exploring potential therapeutic approaches for these diseases.

Lipid droplets imaging with three-photon microscopy

Lipid droplets(LDs)participate in many physiological processes,the abnormality of which will cause chronic diseases and pathologies such as diabetes and obesity.It is crucial to monitor the distribution of LDs at high spatial resolution and large depth.Herein,we carried three-photon imaging of LDs in fat liver.Owing to the large three-photon absorption cross-section of the luminogen named NAP-CF_(3)(1:67×10^(-79) cm^(6) s^(2)),three-photon fluorescence fat liver imaging reached the largest depth of 80μm.Fat liver diagnosis was successfully carried out with excellent performance,providing great potential for LDs-associated pathologies research.

Effect of testicular capsulotomy on lipid droplets in the seminiferous tubules of rats

Aim: In order to reveal the histochemical alteration that might occur during the processes of the spermatogenic dis-ruption induced by testicular capsulotomy, the location and alteration of lipid droplets in the seminiferous tubules wereobserved in the present study. Methods: Osmium tetroxide was used to demonstrate the lipid droplets in the semi-niferous tubules of capsulotomized and sham-operated control testes. Results: In the seminiferous tubules of thesham-operated rat testes, many small lipid droplets were located close to the basement membrane of the seminiferoustubules. But for the capsulotomized testes, the lipid droplets in the seminiferous tubules had increased in size and num-ber, with many lipid droplets migrated towards the lumen of the tubules. Conclusion: The results indicated that aprogressive fatty degeneration occurred in the seminiferous tubules after testicular capsulotomy.

Lipid droplets as metabolic determinants for stemness and chemoresistance in cancer

Previously regarded as simple fat storage particles,new evidence suggests thatlipid droplets(LDs)are dynamic and functional organelles involved in keycellular processes such as membrane biosynthesis,lipid metabolism,cellsignalling and inflammation.Indeed,an increased LD content is one of the mostapparent features resulting from lipid metabolism reprogramming necessary tosupport the basic functions of cancer cells.LDs have been associated to differentcellular processes involved in cancer progression and aggressiveness,such astumorigenicity,invasion and metastasis,as well as chemoresistance.Interestingly,all of these processes are controlled by a subpopulation of highly aggressivetumoral cells named cancer stem cells(CSCs),suggesting that LDs may befundamental elements for stemness in cancer.Considering the key role of CSCs onchemoresistance and disease relapse,main factors of therapy failure,the design ofnovel therapeutic approaches targeting these cells may be the only chance forlong-term survival in cancer patients.In this sense,their biology and functionalproperties render LDs excellent candidates for target discovery and design ofcombined therapeutic strategies.In this review,we summarise the currentknowledge identifying LDs and CSCs as main contributors to cancer aggressiveness,metastasis and chemoresistance.

Anti-lipid droplets accumulation effect of Annona montana(mountain soursop)leaves extract on differentiation of preadipocytes

The Annona genus is a member of Annonaceae,one of the largest families of plants across tropical and subtropical regions.This family has been used in several ethnomedicinal practices to treat a multitude of human diseases.However,the molecular mechanism underlying its effect on the lipid droplet formation and on the expression of adipogenic markers of this plant remain to be investigated.In this study,we examined whether the extracts from the aerial part of Annona montana affect in vitro differentiation of preadipocytes.For our investigations,both mouse embryo fibroblast 3T3-L1 and normal human primary subcutaneous preadipocytes were incubated with Annona montana extracts(-and its subfractions-)and then analyzed on preadipocyte differentiation,lipid content,lipid droplet size and number,the expression of adipogenic-specific transcriptional factors,as well as cell survival.From our examinations,we found the Annona montana ethyl acetate extract to exhibit a potent inhibitory effect on adipogenesis,without affecting cell survival,in a dose-dependent manner.Such inhibitory effects included a significant decrease in the accumulation of lipid content by both a dramatic reduction of size and number of lipid droplets.This extract strongly attenuated the expression of PPARγand HMGB2.It also inhibited the expression of CEBPα,FAS,and Akt without influencing Erk1/2 activities.Our findings suggest that specifically,the Annona montana ethyl acetate extract has a prominent inhibitory effect in cellular pathways of adipocyte differentiation by modulating specific gene expression,which is known to perform a pivotal role during adipogenesis.

Dynamics of Micelle Formation from Mixed Lipid Droplets

Amphiphilic lipid molecules can form various micelles depending on not only their molecular composition but also their self-assembly pathway. In this work, coarse-grained molecular dynamics simulations have been applied to study the micellization behaviors of mixed dipalmitoylphosphatidylcholine （DPPC）/hexadecylphosphocholine （HPC） droplets. By vary- ing DPPC/HPC composition and the size of lipid droplets, various micelles such as spherical and nonspherical （oblate or prolate） vesicles, disk-like micelles, double or single ring-like and worm-like micelles were observed. It is found that the lipid droplet as an initial state favors forming vesicles and ring-like micelles due to in situ micellization. Our simulation results demonstrate that using special initial conditions combined with various molecular compositions is an effective way to tune lipid micellar structure.

Mesoscale Simulation of Vesiculation of Lipid Droplets

An implicit solvent coarse-grained （CG） lipid model using three beads to reflect the basically molecular structure of two-tailed lipid is developed. In this model, the nonbonded interaction employs a variant MIE potential and the bonded interaction utilizes a Harmonic potential form. The CG force field parameters are achieved by matching the structural and mechan-ical properties of dipalmitoylphosphatidylcholine （DPPC） bilayers. The model successfully reproduces the formation of lipid bilayer from a random initial state and the spontaneous vesiculation of lipid bilayer from a disk-like structure. After that, the model is used to sys-tematically study the vesiculation processes of spherical and cylindrical lipid droplets. The results show that the present CG model can effectively simulate the formation and evolution of mesoscale complex vesicles.

Oleic acid reduces steroidogenesis by changing the lipid type stored in lipid droplets of ovarian granulosa cells

Background:Oleic acid is an abundant free fatty acid present in livestock that are in a negative energy-balance state,and it may have detrimental effects on female reproduction and fertility.Oleic acid induces lipid accumulation in bovine granulosa cells,which leads to a foam cell-like morphology and reduced steroidogenesis.However,why oleic acid increases lipid accumulation but decreases steroidogenesis remains unclear.This study focused on oleic acid’s effects on lipid type and steroidogenesis.Results:Oleic acid increased the lipid accumulation in a concentration-dependent manner and mainly increased the triglyceride level and decreased the cholesterol ester level.Oleic acid also led to a decline in estradiol and progesterone production in porcine granulosa cells in vitro.In addition,oleic acid up-regulated the expression of CD36 and diacylglycerol acyltransferase 2,but down-regulated the expression of 3-hydroxy-3-methylglutarylcoenzyme A reductase,scavenger receptor class B member 1 and acetyl-Coenzyme A acetyltransferase 2,as well as steroidogenesis-related genes,including cytochrome P450 family 11 subfamily A member 1,cytochrome P450family 19 subfamily A member 1 and 3 as well as steroidogenic acute regulatory protein at the mRNA and protein levels.An oleic acid-rich diet also enhanced the triglyceride levels and reduced the cholesterol levels in ovarian tissues of female mice,which resulted in lower estradiol levels than in control-fed mice.Compared with the control,decreases in estrus days and the numbers of antral follicles and corpora lutea,as well as an increase in the numbers of the atretic follicles,were found in the oleic acid-fed female mice.Conclusions:Oleic acid changed the lipid type stored in lipid droplets of ovarian granulosa cells,and led to a decrease in steroidogenesis.These results improve our understanding of fertility decline in livestock that are in a negative energy-balance state.

A lipid droplets-targetable fluorescent probe for polarity detection in cells of iron death,inflammation and fatty liver tissue

Abnormal accumulation and metabolism of lipid droplets can lead to a variety of diseases.Polarity,a key parameter of the microenvironment,is closely associated with many diseases and dysfunctions in the body.It is important to elucidate the relationship between the physiological activity of lipid droplets(LDs)and the polarity of the microenvironment.In this work,based on push-pull mechanism,a fluorescent probe(E)-3-(5-(4-(diphenylamino)phenyl)thiophen-2-yl)-1-(2-hydroxyphenyl)prop-2-en-1-one(PPTH)with aggregation-induced emission(AIE)properties for the detection of polarity changes in cells was synthesized.PPTH not only visualize intracellular polarity fluctuation of iron death and inflammation but also distinguish between normal and fatty liver tissue.

Facile synthesis of ultrabright luminogens with specific lipid droplets targeting feature for in vivo two-photon fluorescence retina imaging

The application of fluorescent probes for in vivo retinal imaging is of great importance,which could provide direct and crucial imaging evidence for a better understanding of common eye diseases.Herein,a group of bright organic luminogens with typical electron-donating(D)and electron-accepting(A)structures(abbreviated as LDs-BDM,LDs-BTM,and LDs-BHM)was synthesized through a simple single-step reaction.They were found to be efficient solid-state emitters with high fluorescence quantum yields of above 70%(e.g.,83.7%for LDs-BTM).Their light-emission properties could be tuned by the modulation ofπ-conjugation effect with methoxy groups at different substituent positions.Their resulting fluorescent nanoparticles(NPs)were demonstrated as specific lipid droplets(LDs)targeting probes with high brightness,good biocompatibility,and satisfactory photostability.LDs-BTM NPs with a large two-photon absorption cross section(σ2=249 GM)were further utilized as ultrabright two-photon fluorescence(2PF)nanoprobes for in vivo retina imaging of live zebrafish by NIR excitation at an ultralow concentration(0.5μmol/L).Integrated histological structures at the tissue level and corresponding fine details at the cellular level of the embryonic retina of live zebrafish were clearly demonstrated.This is the first report of using ultrabright LDs-targeting nanoprobes to accurately measure fine details in the retina with 2PF microscopic technique.These good results are anticipated to open up a new avenue in the development of efficient 2PF emitters for non-invasive bioimaging of living animals.

Installing the neurospora carotenoid pathway in plants enables cytosolic formation of provitamin A and its sequestration in lipid droplets

Vitamin A deficiency remains a severe global health issue,which creates a need to biofortify crops with provitamin A carotenoids(PACs).Expanding plant cell capacity for synthesis and storing of PACs outside the plastids is a promising biofortification strategy that has been little explored.Here,we engineered PAC formation and sequestration in the cytosol of Nicotiana benthamiana leaves,Arabidopsis seeds,and citrus callus cells,using a fungal(Neurospora crassa)carotenoid pathway that consists of only three enzymes converting C5 isopentenyl building blocks formed from mevalonic acid into PACs,including β-carotene.This strategy led to the accumulation of significant amounts of phytoene and γ-and β-carotene,in addition to fungal,health-promoting carotenes with 13 conjugated double bonds,such as the PAC torulene,in the cytosol.Increasing the isopentenyl diphosphate pool by adding a truncated Arabidopsis hydroxymethylglutaryl-coenzyme A reductase substantially increased cytosolic carotene production.Engineered carotenes accumulate in cytosolic lipid droplets(CLDs),which represent a novel sequestering sink for storing these pigments in plant cytosol.Importantly,β-carotene accumulated in the cytosol of citrus callus cells was more light stable compared to compared with plastidialβ-carotene.Moreover,engineering cytosolic carotene formation increased the number of large-sized CLDs and the levels of β-apocarotenoids,including retinal,the aldehyde corresponding to vitamin A.Collectively,our study opens up the possibility of exploiting the high-flux mevalonic acid pathway for PAC biosynthesis and enhancing carotenoid sink capacity in green and non-green plant tissues,especially in lipid-storing seeds,and thus paves the way for further optimization of carotenoid biofortification in crops.

Hydrogen-bond locked purine chromophores with high photostability for lipid droplets imaging in cells and tissues

Recently, hydrogen-bonding has attracted extensive attention in the design of chromophores. Here, a new class of hydrogen-bond locked purine chromophores(HOPs) were reported by introducing a hydroxyphenyl group into the C(6) position of purine. The intramolecular hydrogen bond plays a dominant role to light up these probes. As a bonus, HOPs show high photostability. Moreover, HOPs exhibit remarkable capability for the specific lipid droplets imaging in living cells with excellent biocompatibility and are also potential for diagnosing fatty liver diseases. These results bring important new insights into the photophysics of the purine-based chromophores and provide a new scaffold with high photostability for bioimaging.

Manifesting viscosity changes in lipid droplets during iodined CT contrast media treatment by the real-time and in situ fluorescence imaging

Computed tomography(CT) is one of the most commonly used non-invasive clinical imaging modalities to predict, diagnose and treat the disease. Iodinated contrast media(ICM) is a form of intravenous radiocontrast agent containing iodine, which enhances the visibility of hollow tissue structures in medical CT imaging. ICM may cause allergic reactions, contrast-induced nephropathy, hyperthyroidism and possibly metformin accumulation. It is significant to find out the risk factors, pathogenesis, diagnosis, prevention, and treatment of adverse reactions caused by ICM. Revealing the changes of the lipid droplets(LDs)viscosity in pathophysiological processes such as cancer and iodined contrast media induced adverse reaction is not only important for monitoring the occurrence and development of some pathophysiological processes but also vital for the deep insight of the biological effects of LDs in these pathophysiological processes. A lipid droplets targeted fluorescent probe DN-1 was devised to sense cellular viscosity alteration with high selectivity and sensitivity, which was applied to distinguish cancer cells and normal cells and reveal viscosity changes during iodined CT contrast media treatment.

Autophagic Clearance of Lipid Droplets Alters Metabolic Phenotypes in a Genetic Obesity–Diabetes Mouse Model

Lipid droplets(LDs)are intracellular organelles that store neutral lipids,and their aberrant accumulation is associated with many diseases including metabolic disorders such as obesity and diabetes.Meanwhile,the potential pathological contribu-tions of LDs in these diseases are unclear,likely due to a lack of chemical biology tools to clear LDs.We recently developed LD-clearance small molecule compounds,Lipid Droplets·AuTophagy TEthering Compounds(LD·ATTECs),that are able to induce autophagic clearance of LDs in cells and in the liver of db/db(C57BL/6J Leprdb/Leprdb)mouse model,which is a widely used genetic model for obesity–diabetes.Meanwhile,the potential effects on the metabolic phenotype remain to be elucidated.Here,using the metabolic cage assay and the blood glucose assay,we performed phenotypic characteriza-tion of the effects of the autophagic degradation of LDs by LD·ATTECs in the db/db mouse model.The study reveals that LD·ATTECs increased the oxygen uptake of mice and the release of carbon dioxide,enhanced the heat production of animals,partially enhanced the exercise during the dark phase,decreased the blood glucose level and improved insulin sensitivity.Collectively,the study characterized the metabolic phenotypes induced by LD·ATTECs in an obesity–diabetes mouse model,revealing novel functional impacts of autophagic clearance of LDs and providing insights into LD biology and obesity–dia-betes pathogenesis from the phenotypic perspective.

RNA sequencing-based optimization of biological lipid droplets for sonodynamic therapy to reverse tumor hypoxia and elicit robust immune response

Mitochondria-targeted sonodynamic therapy(SDT)is a promising strategy to inhibit tumor growth and activate the anti-tumor immune responses.Identifying the mechanisms underlying mitochondria-targeted SDT,further optimizing its efficacy,developing novel sonosensitizer carriers with good biocompatibility pose major challenges to the clinical practice of SDT.In this study,we investigated the mechanisms of mitochondria-targeted SDT and demonstrated that it suppressed the mitochondrial electron transport chain(ETC)in pancreatic cancer cells through RNA-sequencing analysis.Based on these findings,we constructed the functional lipid droplets(LDs)(CPI-613/IR780@LDs),which combined mitochondria-targeted SDT with the tricarboxylic acid(TCA)cycle inhibitor CPI-613.CPI-613/IR780@LDs synergistically inhibited the TCA cycle and the ETC of mitochondrial aerobic respiration to reduce oxygen consumption and increase reactive oxygen species(ROS)generation at the tumor site,thus enhancing the efficacy of SDT in hypoxic pancreatic cancer.Moreover,the combination of mitochondria-targeted SDT and anti-PD-1 antibody exhibited excellent tumor inhibition and activated anti-tumor immune responses by increasing tumorinfiltrating CD8+T cells and reducing regulatory T cells,synergistically arresting the growth of both primary and metastatic pancreatic tumors.Meanwhile,lipid droplets are cell-derived biological carriers with natural mitochondrial targeting ability and can achieve efficient hydrophobic drug loading through active phagocytosis.Therefore,the functional lipid droplet-based SDT combined with anti-PD-1 antibody holds great potential in the clinical treatment of hypoxic pancreatic cancer.

Porcine enteric alphacoronavirus infection increases lipid droplet accumulation to facilitate the virus replication

Coronaviruses are widely transmissible between humans and animals, causing diseases of varying severity. Porcine enteric alphacoronavirus(PEAV) is a newly-discovered pathogenic porcine enteric coronavirus in recent years, which causes watery diarrhea in newborn piglets. The host inflammatory responses to PEAV and its metabolic regulation mechanisms remain unclear, and no antiviral studies have been reported. Therefore, we investigated the pathogenic mechanism and antiviral drugs of PEAV. The transcriptomic analysis of PEAV-infected host cells revealed that PEAV could upregulate lipid metabolism pathways. In lipid metabolism, steady-state energy processes, which can be mediated by lipid droplets(LDs), are the main functions of organelles. LDs are also important in viral infection and inflammation. In infected cells, PEAV increased LD accumulation, upregulated NF-κB signaling, promoted the production of the inflammatory cytokines IL-1β and IL-8, and induced cell death. Inhibiting LD accumulation with a DGAT-1 inhibitor significantly inhibited PEAV replication, downregulated the NF-κB signaling pathway, reduced the production of IL-1β and IL-8, and inhibited cell death. The NF-κB signaling pathway inhibitor BAY11-7082 significantly inhibited LD accumulation and PEAV replication. Metformin hydrochloride also exerted anti-PEAV effects and significantly inhibited LD accumulation, downregulated the NF-κB signaling pathway, reduced the production of IL-1β and IL-8, and inhibited cell death. LD accumulation in the lipid metabolism pathway therefore plays an important role in the replication and pathogenesis of PEAV, and metformin hydrochloride inhibits LD accumulation and the inflammatory response to exert anti-PEAV activity and reducing pathological injury. These findings contribute new targets for developing treatments for PEAV infections.

OPA3 overexpression modulates lipid droplet production and sensitizes colorectal cancer cells to bevacizumab treatment

Background:Colorectal cancer(CRC)represents a substantial risk to public health.Bevacizumab,thefirst US FDA-approved antiangiogenic drug(AAD)for human CRC treatment,faces resistance in patients.The role of lipid metabolism,particularly through OPA3-regulated lipid droplet production,in overcoming this resistance is under investigation.Methods:The protein expression pattern of OPA3 in CRC primary/normal tissues was evaluated by bioinformatics analysis.OPA3-overexpressed SW-480 and HCT-116 cell lines were established,and bevacizumab resistance and OPA3 effects on cell malignancy were examined.OPA3 protein/mRNA expression and lipid droplet-related genes were measured with Western blot and qRT-PCR.OPA3 subcellular localization was detected using immunofluorescence.Proliferation and apoptosis were assessed via colony formation andflow cytometry.Tube formation assays were conducted to assess the angiogenic potential of human umbilical vein endothelial cells(HUVECs).Lipid analysis was used to measure the phosphatidylcholine(PC)and lysophosphatidylcholine(LPC)levels in CRC cells.Results:Bioinformatics analysis revealed that OPA3 was downregulated in CRC.Overexpression of OPA3 inhibited CRC cell proliferation,stimulated apoptosis,and suppressed the angiogenic ability of HUVECs.OPA3 effectively reversed the resistance of CRC cells to bevacizumab and decreased lipid droplet production in CRC cells.Additionally,OPA3 reversed the bevacizumab-induced lipid droplet production in CRC cells,thereby increasing CRC cell sensitivity to bevacizumab treatment.Conclusion:This study suggests that OPA3 modulates lipid metabolism in CRC cells and reduces resistance to bevacizumab in CRC cells.Therefore,OPA3 may be a potential therapeutic target against the AAD resistance in CRC.

Localization of Seed Oil Body Proteins in Tobacco Protoplasts Reveals Specific Mechanisms of Protein Targeting to Leaf Lipid Droplets

Oleosin, caleosin and steroleosin are normally expressed in developing seed cells and are targeted to oil bodies. In the present work, the cDNA of each gene tagged with fluorescent proteins was transiently expressed into tobacco protoplasts and the fluorescent patterns observed by confocal laser scanning microscopy. Our results indicated clear differences in the endocellular localization of the three proteins. Oleosin and caleosin both share a common structure consisting of a central hydrophobic domain flanked by two hydrophilic domains and were correctly targeted to lipid droplets （LD）, whereas steroleosin, characterized by an N-terminal oil body anchoring domain, was mainly retained in the endoplasmic reticulum （ER）. Protoplast fractionation on sucrose gradients indicated that both oleosin and caleosin- green fluorescent protein （GFP） peaked at different fractions than where steroleosin-GFP or the ER marker binding immunoglobulin protein （BiP）, were recovered. Chemical analysis confirmed the presence of triacylglycerols in one of the fractions where oleosin-GFP was recovered. Finally, only oleosin- and caleosin-GFP were able to reconstitute artificial oil bodies in the presence of triacylglycerols and phospholipids. Taken together, our results pointed out for the first time that leaf LDs can be separated by the ER and both oleosin or caleosin are selectively targeted due to the existence of selective mechanisms controlling protein association with these organelles.

3D imaging lipidometry in single cell by in-flow holographic tomography

The most recent discoveries in the biochemical field are highlighting the increasingly important role of lipid droplets(LDs)in several regulatory mechanisms in living cells.LDs are dynamic organelles and therefore their complete characterization in terms of number,size,spatial positioning and relative distribution in the cell volume can shed light on the roles played by LDs.Until now,fluorescence microscopy and transmission electron microscopy are assessed as the gold standard methods for identifying LDs due to their high sensitivity and specificity.However,such methods generally only provide 2D assays and partial measurements.Furthermore,both can be destructive and with low productivity,thus limiting analysis of large cell numbers in a sample.Here we demonstrate for the first time the capability of 3D visualization and the full LD characterization in high-throughput with a tomographic phase-contrast flow-cytometer,by using ovarian cancer cells and monocyte cell lines as models.A strategy for retrieving significant parameters on spatial correlations and LD 3D positioning inside each cell volume is reported.The information gathered by this new method could allow more in depth understanding and lead to new discoveries on how LDs are correlated to cellular functions.

Light-up lipid droplets for the visualization of lipophagy and atherosclerosis by coumarin-derived bioprobe

Lipid droplets(LDs) are intracellular lipid-metabolism organelles that involved in many physiological processes,metabolic disorders as well as diseases such as atherosclerosis.However,the specific probes that can visually locate abnormal LDs-rich tissues and track LDs-associated behavior to the naked eye with adequate biosafety still are rare.Herein,we develop a new design strategy of LDs-targeted probe based on the solvatochromism of coumarin derivatives.The results revealed that the emission wavelength of coumarin fluorophores gradually red shift in different solvents with increasing polarity,while absorption wavelength almost unchanged.As a result,the enlarged stokes shift of coumarin was emerged from oil to water.Furthermore,properly reducing water solubility and adding electronic donor at the structure of coumarins can enlarge this type of solvatochromism.This discovery was utilized to develop suitable probe for the image of LDs and LDs-rich tissues with high resolution and biosafety.Therefore,LDs-associated behavior was visible to the naked eye during the process of lipophagy and atherosclerosis.We deem that the developed probe here offers a new possibility to accurately diagnosis and analyse LDs-related diseases in clinic and preclinical study.