RESUMO

Investigations indicated that sn-2 palmitate have positive effects on brain development, although its mechanism remains largely unexamined. This research delved into how a diet abundant in sn-2 palmitate influenced the cognitive behavior of mice and elucidated the associated mechanisms using metabolomics and lipidomics. The study demonstrated that dietary sn-2 palmitate led to improved working memory and cognition in mice, as well as an increase in brain BDNF concentration when compared to those fed blend vegetable oil (BVO). This was because sn-2 palmitate feeding promoted the synthesis of very long-chain fatty acids (VLCPUFAs) for the lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE) in the liver. This led to more efficient delivery of VLCPUFAs to the brain, as indicated by elevated concentration of LPC/LPE-VLCPUFAs in the liver and heightened expression of the major facilitator superfamily domain containing 2a (MFSD2A). In essence, this paper offered a potential mechanism by which sn-2 palmitate enhanced mouse neurodevelopment.

Assuntos

Encéfalo , Cognição , Fígado , Lisofosfatidilcolinas , Palmitatos , Animais , Lisofosfatidilcolinas/metabolismo , Camundongos , Fígado/metabolismo , Encéfalo/metabolismo , Encéfalo/crescimento & desenvolvimento , Encéfalo/efeitos dos fármacos , Masculino , Palmitatos/metabolismo , Cognição/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Ácidos Graxos/metabolismo , Ácidos Graxos/química , Humanos

RESUMO

Metabolic reprogramming, a key mechanism regulating the growth and recurrence of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), still lacks effective clinical strategies for its integration into the precise screening of primary liver cancer. This study utilized ultra-high-performance liquid chromatography with quadrupole time-of-flight mass spectrometry to conduct a comprehensive, non-targeted metabolomics analysis, revealing significant upregulation of lipid metabolites such as phosphatidylcholine and lysophosphatidylcholine in patients with HCC and CCA, particularly within the glycerophospholipid metabolic pathway. Hematoxylin and eosin and immunohistochemical staining demonstrated marked upregulation of phospholipase A2 in tumor tissues, further emphasizing the potential of lipid metabolism as a therapeutic target and its important part in the course of cancer. This work provides a new viewpoint for addressing the clinical challenges associated with HCC and CCA, laying the groundwork for the broad application of early diagnosis and personalized treatment strategies, and ultimately aiming to provide tailored and precise therapeutic options for patients.

Assuntos

Carcinoma Hepatocelular , Colangiocarcinoma , Glicerofosfolipídeos , Metabolismo dos Lipídeos , Neoplasias Hepáticas , Humanos , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Glicerofosfolipídeos/metabolismo , Masculino , Pessoa de Meia-Idade , Feminino , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Metabolômica/métodos , Progressão da Doença , Fosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/metabolismo , Idoso , Fosfolipases A2/metabolismo , Reprogramação Metabólica

RESUMO

This study examined the effect of dietary lysolecithin on growth performance, nutrient digestibility, haematobiochemistry, and oxidative status in finisher broiler chickens. In a completely randomized design (CRD), 300 21-day-old Chikun strain chicks were randomly allocated to 30 pens in which they were allotted to 5 dietary treatments as follows TI (0 g /100kg), T2 (100 g /100kg), T3 (200 g /100kg), T4 (300 g /100kg), T5 (400 g /100kg) of feed with 6 replicates pens of 10 during the finisher phase (28 days). Results showed that dietary supplementation with lysolecithin increased the final body weight (FBW) (quadratic P = 0.0178), body weight gain (BWG) (quadratic P = 0.0232), whilst it decreased the total feed intake (TFI) (linear P = 0.0104). Similarly, it linearly increased the retention of dry matter (P = 0.0324); crude protein (P = 0.0029), crude fibre (P = 0.0147), and crude fat (P = 0.0002). Furthermore, it increased the superoxide dismutase (linear P < 0.001), glutathione peroxidase (quadratic P < 0.001), glutathione (linear P < 0.001), whilst it decreased malondialdehyde (linear P = 0.003), without affecting (P > 0.05) the haematobiochemistry parameters. Therefore, dietary lysolecithin could be supplemented up to 400 g /100 kg without compromising performance, nutrient retention, haemato-biochemistry, and oxidative status in finisher broiler diets.

Assuntos

Ração Animal , Fenômenos Fisiológicos da Nutrição Animal , Galinhas , Dieta , Suplementos Nutricionais , Digestão , Lisofosfatidilcolinas , Estresse Oxidativo , Animais , Galinhas/crescimento & desenvolvimento , Galinhas/fisiologia , Galinhas/metabolismo , Ração Animal/análise , Digestão/efeitos dos fármacos , Suplementos Nutricionais/análise , Dieta/veterinária , Estresse Oxidativo/efeitos dos fármacos , Fenômenos Fisiológicos da Nutrição Animal/efeitos dos fármacos , Lisofosfatidilcolinas/administração & dosagem , Lisofosfatidilcolinas/metabolismo , Distribuição Aleatória , Masculino , Nutrientes/metabolismo

RESUMO

Neutrophil extracellular traps (NETs) are three-dimensional reticular structures that release chromatin and cellular contents extracellularly upon neutrophil activation. As a novel effector mechanism of neutrophils, NETs possess the capacity to amplify localized inflammation and have been demonstrated to contribute to the exacerbation of various inflammatory diseases, including cardiovascular diseases and tumors. It is suggested that lysophosphatidylcholine (LPC), as the primary active component of oxidized low-density lipoprotein, represents a significant risk factor for various inflammatory diseases, such as cardiovascular diseases and neurodegenerative diseases. However, the specific mechanism of NETs formation induced by LPC remains unclear. Quercetin has garnered considerable attention due to its anti-inflammatory properties, serving as a prevalent flavonoid in daily diet. However, little is currently known about the underlying mechanisms by which quercetin inhibits NETs formation and alleviates associated diseases. In our study, we utilized LPC-treated primary rat neutrophils to establish an in vitro model of NETs formation, which was subsequently subjected to treatment with a combination of quercetin or relevant inhibitors/activators. Compared to the control group, the markers of NETs and the expression of P2X7R/P38MAPK/NOX2 pathway-associated proteins were significantly increased in cells treated with LPC alone. Quercetin intervention decreased the LPC-induced upregulation of the P2X7R/P38MAPK/NOX2 pathway and effectively reduced the expression of NETs markers. The results obtained using a P2X7R antagonist/activator and P38MAPK inhibitor/activator support these findings. In summary, quercetin reversed the upregulation of the LPC-induced P2X7R/P38MAPK/NOX2 pathway, further mitigating NETs formation. Our study investigated the potential mechanism of LPC-induced NETs formation, elucidated the inhibitory effect of quercetin on NETs formation, and offered new insights into the anti-inflammatory properties of quercetin.

Assuntos

Armadilhas Extracelulares , Lisofosfatidilcolinas , NADPH Oxidase 2 , Neutrófilos , Quercetina , Receptores Purinérgicos P2X7 , Proteínas Quinases p38 Ativadas por Mitógeno , Quercetina/farmacologia , Lisofosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/farmacologia , Armadilhas Extracelulares/metabolismo , Armadilhas Extracelulares/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Ratos , Neutrófilos/metabolismo , Neutrófilos/efeitos dos fármacos , Receptores Purinérgicos P2X7/metabolismo , NADPH Oxidase 2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Masculino

RESUMO

We reported that lysophosphatidic acid (LPA) is present at 0.8 µM in mixed human saliva (MS). In this study, we examined the distribution, origin, and enzymatic generation pathways of LPA in MS. LPA was distributed in the medium and cell pellet fraction; a true level of soluble LPA in MS was about 150 nM. The soluble LPA was assumed to be generated by ecto-type lysophospholipase D on exfoliated cells in MS from LPC that originated mainly from the major salivary gland saliva. Our results with the albumin-back extraction procedures suggest that a significant pool of LPA is kept in the outer layer of the plasma membranes of detached oral mucosal cells. Such pool of LPA may contribute to wound healing in upper digestive organs including oral cavity. We obtained evidence that the choline-producing activity in MS was mainly due to Ca2+-activated lysophospholipase D activity of glycerophosphodiesterase 7.

Assuntos

Lisofosfatidilcolinas , Lisofosfolipídeos , Mucosa Bucal , Diester Fosfórico Hidrolases , Saliva , Adulto , Feminino , Humanos , Masculino , Lisofosfatidilcolinas/metabolismo , Lisofosfolipídeos/metabolismo , Mucosa Bucal/metabolismo , Mucosa Bucal/citologia , Mucosa Bucal/enzimologia , Diester Fosfórico Hidrolases/metabolismo , Saliva/metabolismo , Saliva/enzimologia , Adulto Jovem

RESUMO

Lysophosphatidylcholine (LPC) is a bioactive lipid present at high concentrations in inflamed and injured tissues where it contributes to the initiation and maintenance of pain. One of its important molecular effectors is the transient receptor potential canonical 5 (TRPC5), but the explicit mechanism of the activation is unknown. Using electrophysiology, mutagenesis and molecular dynamics simulations, we show that LPC-induced activation of TRPC5 is modulated by xanthine ligands and depolarizing voltage, and involves conserved residues within the lateral fenestration of the pore domain. Replacement of W577 with alanine (W577A) rendered the channel insensitive to strong depolarizing voltage, but LPC still activated this mutant at highly depolarizing potentials. Substitution of G606 located directly opposite position 577 with tryptophan rescued the sensitivity of W577A to depolarization. Molecular simulations showed that depolarization widens the lower gate of the channel and this conformational change is prevented by the W577A mutation or removal of resident lipids. We propose a gating scheme in which depolarizing voltage and lipid-pore helix interactions act together to promote TRPC5 channel opening.

Assuntos

Lisofosfatidilcolinas , Simulação de Dinâmica Molecular , Canais de Cátion TRPC , Humanos , Canais de Cátion TRPC/metabolismo , Canais de Cátion TRPC/genética , Canais de Cátion TRPC/química , Lisofosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/farmacologia , Animais , Ativação do Canal Iônico/efeitos dos fármacos , Células HEK293 , Mutação , Lisofosfolipídeos/metabolismo , Lisofosfolipídeos/farmacologia , Potenciais da Membrana/efeitos dos fármacos

RESUMO

Lysosomal function is impaired in Niemann-Pick disease type C1 (NPC1), a rare and inherited neurodegenerative disorder, resulting in late endosomal/lysosomal accumulation of unesterified cholesterol. The precise pathogenic mechanism of NPC1 remains incompletely understood. In this study, we employed metabolomics to uncover secondary accumulated substances in NPC1. Our findings unveiled a substantial elevation in the levels of three alkyl-lysophosphatidylcholine [alkyl-LPC, also known as lyso-platelet activating factor (PAF)] species in NPC1 compared to controls across various tissues, including brain tissue from individuals with NPC1, liver, spleen, cerebrum, cerebellum, and brain stem from NPC1 mice, as well as in both brain and liver tissue from NPC1 cats. The three elevated alkyl-LPC species were as follows: LPC O-16:0, LPC O-18:1, and LPC O-18:0. However, the levels of PAF 16:0, PAF 18:1, and PAF 18:0 were not altered in NPC1. In the NPC1 feline model, the brain and liver alkyl-LPC levels were reduced following 2-hydroxypropyl-ß-cyclodextrin (HPßCD) treatment, suggesting that alkyl-LPCs are secondary storage metabolites in NPC1 disease. Unexpectedly, cerebrospinal fluid (CSF) levels of LPC O-16:0 and LPC O-18:1 were decreased in individuals with NPC1 compared to age-appropriate comparison samples, and their levels were increased in 80% of participants 2 years after intrathecal HPßCD treatment. The fold increases in CSF LPC O-16:0 and LPC O-18:1 levels were more pronounced in responders compared to nonresponders. This study identified alkyl-LPC species as secondary storage metabolites in NPC1 and indicates that LPC O-16:0 and LPC O-18:1, in particular, could serve as potential biomarkers for tracking treatment response in NPC1 patients.

Assuntos

Lisofosfatidilcolinas , Doença de Niemann-Pick Tipo C , Doença de Niemann-Pick Tipo C/metabolismo , Doença de Niemann-Pick Tipo C/patologia , Animais , Gatos , Camundongos , Humanos , Lisofosfatidilcolinas/metabolismo , Masculino , Feminino , Encéfalo/metabolismo , 2-Hidroxipropil-beta-Ciclodextrina , Criança , Adulto , Fígado/metabolismo , Adolescente , Pré-Escolar , beta-Ciclodextrinas/farmacologia

RESUMO

Platelets are essential component of circulation that plays a major role in hemostasis and thrombosis. During activation and its demise, platelets release platelet-derived microvesicles, with lysophosphatidylcholine (LPC) being a prominent component in their lipid composition. LPC, an oxidized low-density lipoprotein, is involved in cellular metabolism, but its higher level is implicated in pathologies like atherosclerosis, diabetes, and inflammatory disorders. Despite this, its impact on platelet function remains relatively unexplored. To address this, we studied LPC's effects on washed human platelets. A multimode plate reader was employed to measure reactive oxygen species and intracellular calcium using H2DCF-DA and Fluo-4-AM, respectively. Flow cytometry was utilized to measure phosphatidylserine expression, mitochondrial membrane potential (ΔΨm), and mitochondrial permeability transition pore (mPTP) formation using FITC-Annexin V, JC-1, and CoCl2/calcein-AM, respectively. Additionally, platelet morphology and its ultrastructure were observed via phase contrast and electron microscopy. Sonoclot and light transmission aggregometry were employed to examine fibrin formation and platelet aggregation, respectively. The findings demonstrate that LPC induced oxidative stress and increased intracellular calcium in platelets, resulting in increased phosphatidylserine expression and reduced ΔΨm. LPC triggered caspase-independent platelet death and mPTP opening via cytosolic and mitochondrial calcium, along with microvesiculation and reduced platelet counts. LPC increased the platelet's size, adopting a balloon-shaped morphology, causing membrane fragmentation and releasing its cellular contents, while inducing a pro-coagulant phenotype with increased fibrin formation and reduced integrin αIIbß3 activation. Conclusively, this study reveals LPC-induced oxidative stress and calcium-mediated platelet death, necrotic in nature with pro-coagulant properties, potentially impacting inflammation and repair mechanisms during vascular injury.

Assuntos

Plaquetas , Cálcio , Morte Celular , Lisofosfatidilcolinas , Estresse Oxidativo , Espécies Reativas de Oxigênio , Humanos , Estresse Oxidativo/efeitos dos fármacos , Lisofosfatidilcolinas/farmacologia , Lisofosfatidilcolinas/metabolismo , Cálcio/metabolismo , Plaquetas/metabolismo , Plaquetas/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Morte Celular/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Agregação Plaquetária/efeitos dos fármacos , Poro de Transição de Permeabilidade Mitocondrial/metabolismo

RESUMO

BACKGROUND: Recent studies indicated that bioactive lipids of phosphatidylcholines (PCs) and lysophosphatidylcholines (LysoPCs) predict unhealthy metabolic phenotypes, but results remain inconsistent. To fill this knowledge gap, we investigated whether essential trace elements affect PC-Lyso PC remodeling pathways and the risk of insulin resistance (IR). METHODS: Anthropometric and blood biochemical data (glucose, insulin, and lipoprotein-associated phospholipase A2 (Lp-PLA2)) were obtained from 99 adults. Blood essential/probably essential trace elements and lipid metabolites were respectively measured by inductively coupled plasma mass spectrometry (ICP-MS), and ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). RESULT AND CONCLUSION: Except for LysoPC (O-18:0/0:0), an inverse V shape was observed between body weight and PC and LysoPC species. A Pearson correlation analysis showed that essential/probably-essential metals (Se, Cu, and Ni: r=-0.4∼-0.7) were negatively correlated with PC metabolites but positively correlated with LysoPC (O-18:0/0:0) (Se, Cu, and Ni: r=0.85-0.64). Quantile-g computation showed that one quantile increase in essential metals was associated with a 2.16-fold increase in serum Lp-PLA2 (ß=2.16 (95 % confidence interval (CI): 0.34, 3.98), p=0.023), which are key enzymes involved in PC/Lyso PC metabolism. An interactive analysis showed that compared to those with the lowest levels (reference), individuals with the highest levels of serum PCs (pooled, M2) and the lowest essential/probably essential metals (M1) were associated with a healthier body composition and had a 76 % decreased risk of IR (odds ratio (OR)=0.24 (95 % CI: 0.06, 0.90), p<0.05). In contrast, increased exposure to LysoPC(O-18:0/0:0) (M2) and essential metals (M2) exhibited an 8.22-times highest risk of IR (OR= 8.22 (2.07, 32.57), p<0.05) as well as an altered body composition. In conclusion, overexposure to essential/probably essential trace elements may promote an unhealthy body weight and IR through modulating PC/LysoPC remodeling pathways.

Assuntos

Composição Corporal , Resistência à Insulina , Fosfatidilcolinas , Oligoelementos , Humanos , Masculino , Fosfatidilcolinas/sangue , Fosfatidilcolinas/metabolismo , Feminino , Oligoelementos/sangue , Oligoelementos/metabolismo , Adulto , Pessoa de Meia-Idade , Lisofosfatidilcolinas/sangue , Lisofosfatidilcolinas/metabolismo

RESUMO

Obesity and metabolic syndrome alter serum lipid profiles. They also increase vulnerability to viral infections and worsen the survival rate and symptoms after infection. How serum lipids affect influenza virus proliferation is unclear. Here, we investigated the effects of lysophosphatidylcholines on influenza A virus (IAV) proliferation. IAV particles in the culture medium were titrated using extraction-free quantitative PCR, and viral RNA and protein levels were assessed using real-time PCR and Western blot, respectively. RNA sequencing data were analyzed using PCA and heatmap analysis, and pathway analysis was performed using the KEGG mapper and PathIN tools. Statistical analysis was conducted using SPSS21.0. LPC treatment of THP-1 cells significantly increased IAV proliferation and IAV RNA and protein levels, and saturated LPC was more active in IAV RNA expression than unsaturated LPC was. The functional analysis of genes affected by LPCs showed that the expression of genes involved in IAV signaling, such as suppressor of cytokine signaling 3 (SOCS3), phosphoinositide-3-kinase regulatory subunit 3 (PI3K) and AKT serine/threonine kinase 3 (AKT3), Toll-like receptor 7 (TKR7), and interferon gamma receptor 1 (IFNGR1), was changed by LPC. Altered influenza A pathways were linked with MAPK and PI3K/AKT signaling. Treatment with inhibitors of MAPK or PI3K attenuated viral gene expression changes induced by LPCs. The present study shows that LPCs stimulated virus reproduction by modifying the cellular environment to one in which viruses proliferated better. This was mediated by the MAPK, JNK, and PI3K/AKT pathways. Further animal studies are needed to confirm the link between LPCs from serum or the respiratory system and IAV proliferation.

Assuntos

Vírus da Influenza A , Lisofosfatidilcolinas , Sistema de Sinalização das MAP Quinases , Replicação Viral , Humanos , Lisofosfatidilcolinas/farmacologia , Lisofosfatidilcolinas/metabolismo , Replicação Viral/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Vírus da Influenza A/fisiologia , Macrófagos/metabolismo , Macrófagos/virologia , Macrófagos/efeitos dos fármacos , Células THP-1 , Diferenciação Celular/efeitos dos fármacos , Influenza Humana/virologia , Influenza Humana/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais

RESUMO

BACKGROUND: Lipid droplet (LD)-laden microglia is a key pathological hallmark of multiple sclerosis. The recent discovery of this novel microglial subtype, lipid-droplet-accumulating microglia (LDAM), is notable for increased inflammatory factor secretion and diminished phagocytic capability. Lipophagy, the autophagy-mediated selective degradation of LDs, plays a critical role in this context. This study investigated the involvement of microRNAs (miRNAs) in lipophagy during demyelinating diseases, assessed their capacity to modulate LDAM subtypes, and elucidated the potential underlying mechanisms involved. METHODS: C57BL/6 mice were used for in vivo experiments. Two weeks post demyelination induction at cervical level 4 (C4), histological assessments and confocal imaging were performed to examine LD accumulation in microglia within the lesion site. Autophagic changes were observed using transmission electron microscopy. miRNA and mRNA multi-omics analyses identified differentially expressed miRNAs and mRNAs under demyelinating conditions and the related autophagy target genes. The role of miR-223 in lipophagy under these conditions was specifically explored. In vitro studies, including miR-223 upregulation in BV2 cells via lentiviral infection, validated the bioinformatics findings. Immunofluorescence staining was used to measure LD accumulation, autophagy levels, target gene expression, and inflammatory mediator levels to elucidate the mechanisms of action of miR-223 in LDAM. RESULTS: Oil Red O staining and confocal imaging revealed substantial LD accumulation in the demyelinated spinal cord. Transmission electron microscopy revealed increased numbers of autophagic vacuoles at the injury site. Multi-omics analysis revealed miR-223 as a crucial regulatory gene in lipophagy during demyelination. It was identified that cathepsin B (CTSB) targets miR-223 in autophagy to integrate miRNA, mRNA, and autophagy gene databases. In vitro, miR-223 upregulation suppressed CTSB expression in BV2 cells, augmented autophagy, alleviated LD accumulation, and decreased the expression of the inflammatory mediator IL-1ß. CONCLUSION: These findings indicate that miR-223 plays a pivotal role in lipophagy under demyelinating conditions. By inhibiting CTSB, miR-223 promotes selective LD degradation, thereby reducing the lipid burden and inflammatory phenotype in LDAM. This study broadens the understanding of the molecular mechanisms of lipophagy and proposes lipophagy induction as a potential therapeutic approach to mitigate inflammatory responses in demyelinating diseases.

Assuntos

Autofagia , Catepsina B , Doenças Desmielinizantes , Gotículas Lipídicas , Lisofosfatidilcolinas , Camundongos Endogâmicos C57BL , MicroRNAs , Microglia , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Microglia/metabolismo , Microglia/patologia , Camundongos , Gotículas Lipídicas/metabolismo , Doenças Desmielinizantes/metabolismo , Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/genética , Doenças Desmielinizantes/patologia , Catepsina B/metabolismo , Catepsina B/genética , Lisofosfatidilcolinas/metabolismo , Modelos Animais de Doenças , Masculino , Regulação da Expressão Gênica , Linhagem Celular

RESUMO

The NLRP3 inflammasome functions as an inflammatory driver, but its relationship with lipid metabolic changes in early sepsis remains unclear. Here, we found that GITR expression in monocytes/macrophages was induced by lysophosphatidylcholine (LPC) and was positively correlated with the severity of sepsis. GITR is a costimulatory molecule that is mainly expressed on T cells, but its function in macrophages is largely unknown. Our in vitro data showed that GITR enhanced LPC uptake by macrophages and specifically enhanced NLRP3 inflammasome-mediated macrophage pyroptosis. Furthermore, in vivo studies using either cecal ligation and puncture (CLP) or LPS-induced sepsis models demonstrated that LPC exacerbated sepsis severity/lethality, while conditional knockout of GITR in myeloid cells or NLRP3/caspase-1/IL-1ß deficiency attenuated sepsis severity/lethality. Mechanistically, GITR specifically enhanced inflammasome activation by regulating the posttranslational modification (PTM) of NLRP3. GITR competes with NLRP3 for binding to the E3 ligase MARCH7 and recruits MARCH7 to induce deacetylase SIRT2 degradation, leading to decreasing ubiquitination but increasing acetylation of NLRP3. Overall, these findings revealed a novel role of macrophage-derived GITR in regulating the PTM of NLRP3 and systemic inflammatory injury, suggesting that GITR may be a potential therapeutic target for sepsis and other inflammatory diseases. GITR exacerbates LPC-induced macrophage pyroptosis in sepsis via posttranslational regulation of NLRP3. According to the model, LPC levels increase during the early stage of sepsis, inducing GITR expression on macrophages. GITR not only competes with NLRP3 for binding to the E3 ligase MARCH7 but also recruits MARCH7 to induce the degradation of the deacetylase SIRT2, leading to decreasing ubiquitination but increasing acetylation of NLRP3 and therefore exacerbating LPC-induced NLRP3 inflammasome activation, macrophage pyroptosis and systemic inflammatory injury.

Assuntos

Proteína Relacionada a TNFR Induzida por Glucocorticoide , Lisofosfatidilcolinas , Macrófagos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR , Processamento de Proteína Pós-Traducional , Piroptose , Sepse , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Animais , Sepse/imunologia , Macrófagos/metabolismo , Macrófagos/imunologia , Lisofosfatidilcolinas/metabolismo , Camundongos , Proteína Relacionada a TNFR Induzida por Glucocorticoide/metabolismo , Inflamassomos/metabolismo , Masculino , Camundongos Knockout , Humanos , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Sirtuína 2/metabolismo , Sirtuína 2/genética , Acetilação

RESUMO

Bone fracture healing is a complex process in which specific molecular knowledge is still lacking. The citrulline-arginine-nitric oxide metabolism is one of the involved pathways, and its enrichment via citrulline supplementation can enhance fracture healing. This study investigated the molecular effects of citrulline supplementation during the different fracture healing phases in a rat model. Microcomputed tomography (µCT) was applied for the analysis of the fracture callus formation. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) and liquid-chromatography tandem mass spectrometry (LC-MS/MS) were used for lipid and protein analyses, respectively. µCT analysis showed no significant differences in the fracture callus volume and volume fraction between the citrulline supplementation and control group. The observed lipid profiles for the citrulline supplementation and control group were distinct for the different fracture healing stages. The main contributing lipid classes were phosphatidylcholines (PCs) and lysophosphatidylcholines (LPCs). The changing effect of citrulline supplementation throughout fracture healing was indicated by changes in the differentially expressed proteins between the groups. Pathway analysis showed an enhancement of fracture healing in the citrulline supplementation group in comparison to the control group via improved angiogenesis and earlier formation of the soft and hard callus. This study showed the molecular effects on lipids, proteins, and pathways associated with citrulline supplementation during bone fracture healing, even though no effect was visible with µCT.

Assuntos

Citrulina , Consolidação da Fratura , Ratos Sprague-Dawley , Espectrometria de Massas em Tandem , Microtomografia por Raio-X , Animais , Consolidação da Fratura/efeitos dos fármacos , Ratos , Citrulina/análise , Citrulina/metabolismo , Citrulina/farmacologia , Espectrometria de Massas em Tandem/métodos , Microtomografia por Raio-X/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Suplementos Nutricionais/análise , Modelos Animais de Doenças , Masculino , Calo Ósseo/efeitos dos fármacos , Calo Ósseo/diagnóstico por imagem , Calo Ósseo/metabolismo , Cromatografia Líquida/métodos , Lisofosfatidilcolinas/metabolismo , Lisofosfatidilcolinas/análise , Fosfatidilcolinas/metabolismo , Fosfatidilcolinas/análise , Fosfatidilcolinas/farmacologia

RESUMO

The human malaria parasite Plasmodium falciparum requires exogenous fatty acids to support its growth during the pathogenic, asexual erythrocytic stage. Host serum lysophosphatidylcholine (LPC) is a significant fatty acid source, yet the metabolic processes responsible for the liberation of free fatty acids from exogenous LPC are unknown. Using an assay for LPC hydrolysis in P. falciparum-infected erythrocytes, we have identified small-molecule inhibitors of key in situ lysophospholipase activities. Competitive activity-based profiling and generation of a panel of single-to-quadruple knockout parasite lines revealed that two enzymes of the serine hydrolase superfamily, termed exported lipase (XL) 2 and exported lipase homolog (XLH) 4, constitute the dominant lysophospholipase activities in parasite-infected erythrocytes. The parasite ensures efficient exogenous LPC hydrolysis by directing these two enzymes to distinct locations: XL2 is exported to the erythrocyte, while XLH4 is retained within the parasite. While XL2 and XLH4 were individually dispensable with little effect on LPC hydrolysis in situ, loss of both enzymes resulted in a strong reduction in fatty acid scavenging from LPC, hyperproduction of phosphatidylcholine, and an enhanced sensitivity to LPC toxicity. Notably, growth of XL/XLH-deficient parasites was severely impaired when cultured in media containing LPC as the sole exogenous fatty acid source. Furthermore, when XL2 and XLH4 activities were ablated by genetic or pharmacologic means, parasites were unable to proliferate in human serum, a physiologically relevant fatty acid source, revealing the essentiality of LPC hydrolysis in the host environment and its potential as a target for anti-malarial therapy.

Assuntos

Malária Falciparum , Parasitos , Animais , Humanos , Plasmodium falciparum , Lisofosfatidilcolinas/metabolismo , Lisofosfolipase/genética , Lisofosfolipase/metabolismo , Malária Falciparum/parasitologia , Eritrócitos/metabolismo , Parasitos/metabolismo , Ácidos Graxos/metabolismo , Lipase/metabolismo , Proteínas de Protozoários/metabolismo

RESUMO

INTRODUCTION: Placental phospholipid synthesis is critical for the expansion of the placental exchange surface area and for production of signaling molecules. Despite their importance, it is not yet established which enzymes involved in the de novo synthesis and remodeling of placental phospholipids are expressed and active in the human placenta. METHODS: We identified phospholipid synthesis enzymes by immunoblotting in placental homogenates and immunofluorescence in placenta tissue sections. Primary human trophoblast (PHT) cells from term healthy placentas (n = 10) were cultured and exposed to 13C labeled fatty acids (16:0, 18:1 and 18:2 n-6, 22:6 n-3) for 2 and 24 h. Three phospholipid classes; phosphatidic acid, phosphatidylcholine, and lysophosphatidylcholine containing 13C fatty acids were quantified by Liquid Chromatography with tandem mass spectrometry (LC/MS-MS). RESULTS: Acyl transferase and phospholipase enzymes were detected in human placenta homogenate and primarily expressed in the syncytiotrophoblast. Three representative 13C fatty acids (16:0, 18:1 and 18:2 n-6) were incorporated rapidly into phosphatidic acid in trophoblasts, but 13C labeled docosahexaenoic acid (DHA; 22:6 n-3) incorporation was not detected. 13C DHA was incorporated into phosphatidylcholine. Lysophosphatidylcholine containing all four 13C labeled fatty acids were found in high abundance. CONCLUSIONS: Phospholipid synthesis and remodeling enzymes are present in the syncytiotrophoblast. 13C labeled fatty acids were rapidly incorporated into cellular phospholipids. 13C DHA was incorporated into phospholipids through the remodeling pathway rather than by de novo synthesis. These understudied pathways are highly active and critical for structure and function of the placenta.

Assuntos

Fosfolipídeos , Placenta , Humanos , Gravidez , Feminino , Placenta/metabolismo , Fosfolipídeos/metabolismo , Lisofosfatidilcolinas/metabolismo , Ácidos Graxos/metabolismo , Fosfatidilcolinas/metabolismo

RESUMO

Lipids and their modifying enzymes regulate diverse features of the tumor microenvironment and cancer progression. The secreted enzyme autotaxin (ATX) hydrolyzes extracellular lysophosphatidylcholine to generate the multifunctional lipid mediator lysophosphatidic acid (LPA) and supports the growth of several tumor types, including pancreatic ductal adenocarcinoma (PDAC). Here we show that ATX suppresses the accumulation of eosinophils in the PDAC microenvironment. Genetic or pharmacologic ATX inhibition increased the number of intratumor eosinophils, which promote tumor cell apoptosis locally and suppress tumor progression. Mechanistically, ATX suppresses eosinophil accumulation via an autocrine feedback loop, wherein ATX-LPA signaling negatively regulates the activity of the AP-1 transcription factor c-Jun, in turn suppressing the expression of the potent eosinophil chemoattractant CCL11 (eotaxin-1). Eosinophils were identified in human PDAC specimens, and rare individuals with high intratumor eosinophil abundance had the longest overall survival. Together with recent findings, this study reveals the context-dependent, immune-modulatory potential of ATX-LPA signaling in cancer.

Assuntos

Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Eosinófilos/metabolismo , Quimiocina CCL11 , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Processos Neoplásicos , Lisofosfatidilcolinas/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Microambiente Tumoral

RESUMO

Lysosomes are acidic organelles responsible for lipid catabolism, and their functions can be disrupted by cationic amphiphilic drugs that neutralize lumenal pH and thereby inhibit most lysosomal hydrolases. These drugs can also induce lysosomal membrane permeabilization and cancer cell death, but the underlying mechanism remains elusive. Here, we uncover that the cationic amphiphilic drugs induce a substantial accumulation of cytolytic lysoglycerophospholipids within the lysosomes of cancer cells, and thereby prevent the recycling of lysoglycerophospholipids to produce common membrane glycerophospholipids. Using quantitative mass spectrometry-based shotgun lipidomics, we demonstrate that structurally diverse cationic amphiphilic drugs, along with other types of lysosomal pH-neutralizing reagents, elevate the amounts of lysoglycerophospholipids in MCF7 breast carcinoma cells. Lysoglycerophospholipids constitute ∼11 mol% of total glycerophospholipids in lysosomes purified from MCF7 cells, compared with ∼1 mol% in the cell lysates. Treatment with cationic amphiphilic drug siramesine further elevates the lysosomal lysoglycerophospholipid content to ∼24 mol% of total glycerophospholipids. Exogenously added traceable lysophosphatidylcholine is rapidly acylated to form diacylphosphatidylcholine, but siramesine treatment sequesters the lysophosphatidylcholine in the lysosomes and prevents it from undergoing acylation. These findings shed light on the unexplored role of lysosomes in the recycling of lysoglycerophospholipids and uncover the mechanism of action of promising anticancer agents.

Assuntos

Glicerofosfolipídeos , Indóis , Neoplasias , Compostos de Espiro , Humanos , Glicerofosfolipídeos/metabolismo , Lisofosfatidilcolinas/metabolismo , Lisossomos/metabolismo , Morte Celular , Neoplasias/metabolismo

RESUMO

Lysophosphatidylcholine (LPC) is a bioactive lipid that has been shown to attenuate endothelium-dependent vasorelaxation contributing to endothelial dysfunction; however, the underlying mechanisms are not well understood. In this study, we investigated the molecular mechanisms involved in the development of LPC-evoked impairment of endothelium-dependent vasorelaxation. In aortic rings isolated from wild-type (WT) mice, a 20-min exposure to LPC significantly reduced the acetylcholine chloride (ACh)-induced vasorelaxation indicating the impairment of normal endothelial function. Interestingly, pharmacological inhibition of autotaxin (ATX) by GLPG1690 partially reversed the endothelial dysfunction, suggesting that lysophosphatidic acid (LPA) derived from LPC may be involved in the effect. Therefore, the effect of LPC was also tested in aortic rings isolated from different LPA receptor knock-out (KO) mice. LPC evoked a marked reduction in ACh-dependent vasorelaxation in Lpar1, Lpar2, and Lpar4 KO, but its effect was significantly attenuated in Lpar5 KO vessels. Furthermore, addition of superoxide dismutase reduced the LPC-induced endothelial dysfunction in WT but not in the Lpar5 KO mice. In addition, LPC increased H2O2 release from WT vessels, which was significantly reduced in Lpar5 KO vessels. Our findings indicate that the ATX-LPA-LPA5 receptor axis is involved in the development of LPC-induced impairment of endothelium-dependent vasorelaxation via LPA5 receptor-mediated reactive oxygen species production. Taken together, in this study, we identified a new pathway contributing to the development of LPC-induced endothelial dysfunction.

Assuntos

Peróxido de Hidrogênio , Receptores de Ácidos Lisofosfatídicos , Animais , Camundongos , Endotélio/metabolismo , Lisofosfatidilcolinas/farmacologia , Lisofosfatidilcolinas/metabolismo , Lisofosfolipídeos/farmacologia , Lisofosfolipídeos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo

RESUMO

α-synuclein (α-Syn) is a presynaptic protein that is involved in Parkinson's and other neurodegenerative diseases and binds to negatively charged phospholipids. Previously, we reported that α-Syn clusters synthetic proteoliposomes that mimic synaptic vesicles. This vesicle-clustering activity depends on a specific interaction of α-Syn with anionic phospholipids. Here, we report that α-Syn surprisingly also interacts with the neutral phospholipid lysophosphatidylcholine (lysoPC). Even in the absence of anionic lipids, lysoPC facilitates α-Syn-induced vesicle clustering but has no effect on Ca2+-triggered fusion in a single vesicle-vesicle fusion assay. The A30P mutant of α-Syn that causes familial Parkinson disease has a reduced affinity to lysoPC and does not induce vesicle clustering. Taken together, the α-Syn-lysoPC interaction may play a role in α-Syn function.

Assuntos

Doença de Parkinson , alfa-Sinucleína , Humanos , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo , Vesículas Sinápticas/metabolismo , Lisofosfatidilcolinas/metabolismo , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Fosfolipídeos/metabolismo

RESUMO

Synovial fluid (SF) from human knee joints with osteoarthritis (OA) has elevated levels of lysophosphatidylcholine (LPC) species, but their functional role is not well understood. This in vitro study was designed to test the hypothesis that various LPCs found elevated in OA SF and their metabolites, lysophosphatidic acids (LPAs), modulate the abundance of proteins and phospholipids (PLs) in human fibroblast-like synoviocytes (FLSs), with even minute chemical variations in lysophospholipids determining the extent of regulation. Cultured FLSs (n = 5-7) were treated with one of the LPC species, LPA species, IL-1ß, or a vehicle. Tandem mass tag peptide labeling coupled with LC-MS/MS/MS was performed to quantify proteins. The expression of mRNA from regulated proteins was analyzed using RT-PCR. PL synthesis was determined via ESI-MS/MS, and the release of radiolabeled PLs was determined by means of liquid scintillation counting. In total, 3960 proteins were quantified using multiplexed MS, of which 119, 8, and 3 were significantly and reproducibly regulated by IL-1ß, LPC 16:0, and LPC 18:0, respectively. LPC 16:0 significantly inhibited the release of PLs and the synthesis of phosphatidylcholine, LPC, and sphingomyelin. Neither LPC metabolite-LPA 16:0 nor LPA 18:0-had any reproducible effect on the levels of each protein. In conclusion, small chemical variations in LPC species can result in the significantly altered expression and secretion of proteins and PLs from FLSs. IL-1ß influenced all proteins that were reproducibly regulated by LPC 16:0. LPC species are likely to modulate FLS protein expression only in more advanced OA stages with low IL-1ß levels. None of the eight proteins being significantly regulated by LPC 16:0 have been previously reported in OA. However, our in vitro findings show that the CD81 antigen, calumenin, and B4E2C1 are promising candidates for further study, focusing in particular on their potential ability to modulate inflammatory and catabolic mechanisms.

Assuntos

Osteoartrite , Sinoviócitos , Humanos , Sinoviócitos/metabolismo , Espectrometria de Massas em Tandem , Lipidômica , Proteômica , Cromatografia Líquida , Lisofosfolipídeos/metabolismo , Osteoartrite/metabolismo , Lisofosfatidilcolinas/metabolismo , Fibroblastos/metabolismo