RESUMEN

Mitochondria are involved in various cellular processes, such as energy production, inflammatory responses and cell death. Mitochondrial dysfunction is associated with many age-related diseases, including neurological disorders and heart failure. Mitochondrial quality is strictly maintained by mitochondrial dynamics linked to an adequate supply of phospholipids and other substances from the endoplasmic reticulum (ER). The outer mitochondrial membrane-localized E3 ubiquitin ligase MITOL/MARCHF5 is responsible for mitochondrial quality control through the regulation of mitochondrial dynamics, formation of mitochondria-ER contacts and mitophagy. MITOL deficiency has been shown to impair mitochondrial function, cause an excessive inflammatory response and increase vulnerability to stress, resulting in the exacerbation of the disease. In this study, we overview the ubiquitin-mediated regulation of mitochondrial function by MITOL and the relationship between MITOL and diseases.

Asunto(s)

Ubiquitina-Proteína Ligasas , Ubiquitina , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Retículo Endoplásmico/metabolismo , Muerte Celular , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo

RESUMEN

Introduction: Although Staphylococcus aureus is the leading cause of biofilm-related infections, the lipidomic distributions within these biofilms is poorly understood. Here, lipidomic mapping of S. aureus biofilm cross-sections was performed to investigate heterogeneity between horizontal biofilm layers. Methods: S. aureus biofilms were grown statically, embedded in a mixture of carboxymethylcellulose/gelatin, and prepared for downstream matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS). Trapped ion mobility spectrometry (TIMS) was also applied prior to mass analysis. Results: Implementation of TIMS led to a âˆ¼ threefold increase in the number of lipid species detected. Washing biofilm samples with ammonium formate (150 mM) increased signal intensity for some bacterial lipids by as much as tenfold, with minimal disruption of the biofilm structure. MALDI TIMS IMS revealed that most lipids localize primarily to a single biofilm layer, and species from the same lipid class such as cardiolipins CL(57:0) - CL(66:0) display starkly different localizations, exhibiting between 1.5 and 6.3-fold intensity differences between layers (n = 3, p < 0.03). No horizontal layers were observed within biofilms grown anaerobically, and lipids were distributed homogenously. Conclusions: High spatial resolution analysis of S. aureus biofilm cross-sections by MALDI TIMS IMS revealed stark lipidomic heterogeneity between horizontal S. aureus biofilm layers demonstrating that each layer was molecularly distinct. Finally, this workflow uncovered an absence of layers in biofilms grown under anaerobic conditions, possibly indicating that oxygen contributes to the observed heterogeneity under aerobic conditions. Future applications of this workflow to study spatially localized molecular responses to antimicrobials could provide new therapeutic strategies.

RESUMEN

Lysophosphatidylcholine (LPC) has been widely used as emulsifier in animal feeds to enhance the lipid utilization. However, the effects of LPC on fillet quality has rarely been known. The present study was the first time to investigate the response of fish muscle lipidomics to dietary LPC supplementation. Turbot muscle samples were collected after a 56-day feeding trial where the experimental diet contained 0 or 0.25% LPC. Targeted tandem mass spectrometry was used in the lipidomic analysis. A total of 62 individual lipids (58 up-regulated and 7 down-regulated by LPC) showed significant difference in concentration in response to dietary LPC. Most of these differentially abundant lipids were diacylglycerol, free fatty acid and cardiolipin, and they all were up-regulated by dietary LPC. However, LPC exerted only marginal effects on muscle fatty acid composition and lipid content. The effects of dietary LPC on fillet lipid composition cannot be neglected in fish product evaluation.

RESUMEN

Detailed knowledge on tissue-specific metabolic reprogramming in diabetic nephropathy (DN) is vital for more accurate understanding the molecular pathological signature and developing novel therapeutic strategies. In the present study, a spatial-resolved metabolomics approach based on air flow-assisted desorption electrospray ionization (AFADESI) and matrix-assisted laser desorption ionization (MALDI) integrated mass spectrometry imaging (MSI) was proposed to investigate tissue-specific metabolic alterations in the kidneys of high-fat diet-fed and streptozotocin (STZ)-treated DN rats and the therapeutic effect of astragaloside IV, a potential anti-diabetic drug, against DN. As a result, a wide range of functional metabolites including sugars, amino acids, nucleotides and their derivatives, fatty acids, phospholipids, sphingolipids, glycerides, carnitine and its derivatives, vitamins, peptides, and metal ions associated with DN were identified and their unique distribution patterns in the rat kidney were visualized with high chemical specificity and high spatial resolution. These region-specific metabolic disturbances were ameliorated by repeated oral administration of astragaloside IV (100 mg/kg) for 12 weeks. This study provided more comprehensive and detailed information about the tissue-specific metabolic reprogramming and molecular pathological signature in the kidney of diabetic rats. These findings highlighted the promising potential of AFADESI and MALDI integrated MSI based metabolomics approach for application in metabolic kidney diseases.

RESUMEN

Phosphatidylglycerol is a crucial phospholipid found ubiquitously in biological membranes of prokaryotic and eukaryotic cells. The phosphatidylglycerol phosphate (PGP) synthase (PgsA), a membrane-embedded enzyme, catalyzes the primary reaction of phosphatidylglycerol biosynthesis. Mutations in pgsA frequently correlate with daptomycin resistance in Staphylococcus aureus and other prevalent infectious pathogens. Here we report the crystal structures of S. aureus PgsA (SaPgsA) captured at two distinct states of the catalytic process, with lipid substrate (cytidine diphosphate-diacylglycerol, CDP-DAG) or product (PGP) bound to the active site within a trifurcated amphipathic cavity. The hydrophilic head groups of CDP-DAG and PGP occupy two different pockets in the cavity, inducing local conformational changes. An elongated membrane-exposed surface groove accommodates the fatty acyl chains of CDP-DAG/PGP and opens a lateral portal for lipid entry/release. Remarkably, the daptomycin resistance-related mutations mostly cluster around the active site, causing reduction of enzymatic activity. Our results provide detailed mechanistic insights into the dynamic catalytic process of PgsA and structural frameworks beneficial for development of antimicrobial agents targeting PgsA from pathogenic bacteria.

RESUMEN

BACKGROUND: Failure of fascial healing in the abdominal wall can result in incisional hernia, which is one of the most common complications after laparotomy. Understanding the molecular healing process of abdominal fascia may provide lipid markers of incisional hernia or therapeutic targets that allow prevention or treatment of incisional hernias. PURPOSE: This study aims to investigate temporal and in situ changes of lipids during the normal healing process of abdominal fascia in the first postoperative week. METHODS: Open hemicolectomy was performed in a total of 35 Wistar rats. The midline fascia was closed identically for all rats using a single continuous suturing technique. These animals were sacrificed with equal numbers (n = 5) at each of 7-time points (6, 12, 24, 48, 72, 120, and 168 h. The local and temporal changes of lipids were examined with mass spectrometry imaging and correlated to histologically scored changes during healing using hematoxylin and eosin staining. RESULTS: Two phosphatidylcholine lipid species (PC O-38:5 and PC 38:4) and one phosphatidylethanolamine lipid (PE O-16:1_20:4) were found to significantly correlate with temporal changes of inflammation. A phosphatidylcholine (PC 32:0) and a monosialodihexosylganglioside (GM3 34:1;2) were found to correlate with fibroblast cell growth. CONCLUSION: Glycerophospholipids and gangliosides are strongly involved in the normal healing process of abdominal fascia and their locally fluctuating concentrations are considered as potential lipid markers and therapeutic targets of fascial healing.

RESUMEN

Inhibition of animal cell phospholipid biosynthesis has been proposed for anticancer and antiviral therapies. Using CHO-K1 derived cell lines, we have developed and used a cell-based high-throughput procedure to screen a 1280 compound, small molecule library for inhibitors of phospholipid biosynthesis. We identified tyrphostin AG 879 (AG879), which inhibited phospholipid biosynthesis by 85-90% at a concentration of 10 µM, displaying an IC50 of 1-3 µM. The synthesis of all phospholipid head group classes was heavily affected. Fatty acid biosynthesis was also dramatically inhibited (90%). AG879 inhibited phospholipid biosynthesis in all additional cell lines tested, including MDCK, HUH7, Vero, and HeLa cell lines. In CHO cells, AG879 was cytostatic; cells survived for at least four days during exposure and were able to divide following its removal. AG879 is an inhibitor of receptor tyrosine kinases (RTK) and inhibitors of signaling pathways known to be activated by RTK's also inhibited phospholipid biosynthesis. We speculate that inhibition of RTK by AG879 results in an inhibition of fatty acid biosynthesis with a resulting decrease in phospholipid biosynthesis and that AG879's effect on fatty acid synthesis and/or phospholipid biosynthesis may contribute to its known capacity as an effective antiviral/anticancer agent.

RESUMEN

Mitochondria are functionally versatile organelles. In addition to their conventional role of meeting the cell's energy requirements, mitochondria also actively regulate innate immune responses against infectious and sterile insults. Components of mitochondria, when released or exposed in response to dysfunction or damage, can be directly recognized by receptors of the innate immune system and trigger an immune response. In addition, despite initiation that may be independent from mitochondria, numerous innate immune responses are still subject to mitochondrial regulation as discrete steps of their signaling cascades occur on mitochondria or require mitochondrial components. Finally, mitochondrial metabolites and the metabolic state of the mitochondria within an innate immune cell modulate the precise immune response and shape the direction and character of that cell's response to stimuli. Together, these pathways result in a nuanced and very specific regulation of innate immune responses by mitochondria.

Asunto(s)

Inmunidad Innata , Mitocondrias/metabolismo , Transducción de Señal , Alarminas/metabolismo , Animales , ADN Mitocondrial/genética , Humanos , Modelos Biológicos

RESUMEN

The gene encoding the Saccharomyces cerevisiae phospholipid deacylation enzyme, phospholipase B (ScPLB1), was successfully expressed in E. coli. The enzyme (Scplb1p) was engineered to have a histidine-tag at the C-terminal end and was purified by metal (Ni) affinity chromatography. Enzymatic properties, optimal pH, and substrate specificity were similar to those reported previously. For example, deacylation activity was observed in acidic pH in the absence of Ca2+ and was additive in neutral pH in the presence of Ca2+, and the enzyme had the same substrate priority as reported previously, with the exception of PE, suggesting that yeast phospholipase B could be produced in its native structure in bacterial cells. Scplb1p retained transacylation activity in aqueous medium, and esterified lysophosphatidylcholine with free fatty acid to form phosphatidylcholine in a non-aqueous, glycerin medium. We propose that phospholipase B could serve as an additional tool for in vitro enzyme-mediated phospholipid synthesis.

RESUMEN

The incidence of obesity and its metabolic complications are rapidly increasing and become a major public health issue. This trend is associated with an increase in the prevalence of non-alcoholic fatty liver disease (NAFLD), insulin resistance and diabetes. The sequence of events leading to NAFLD progression and mitochondrial dysfunction and their interrelation remains to be elucidated. This study aimed to explore the installation and progression of NAFLD and its association with the liver mitochondrial structure and activity changes in rats fed an obesogenic diet up to 20 weeks. Male Wistar rats were fed either a standard or high-fat-high-fructose (HFHFR) diet and killed on 4, 8, 12, 16 and 20 weeks of diet intake. Rats fed the HFHFR diet developed mildly overweight, associated with increased adipose tissue weight, hepatic steatosis, hyperglycaemia and hyperinsulinaemia after 8 weeks of HFHFR diet. Hepatic steatosis and many biochemical modifications plateaued at 8-12 weeks of HFHFR diet with slight amelioration afterwards. Interestingly, several biochemical and physiological parameters of mitochondrial function, as well as its phospholipid composition, in particular cardiolipin content, were tightly related to hepatic steatosis installation. These results showed once again the interrelation between hepatic steatosis development and mitochondrial activity alterations without being able to say whether the mitochondrial alterations preceded or followed the installation/progression of hepatic steatosis. Because both hepatic steatosis and mitochondrial alterations occurred as early as 4 weeks of diet, future studies should consider these four 1st weeks to reveal the exact interconnection between these major consequences of obesogenic diet intake.

Asunto(s)

Dieta Alta en Grasa/efectos adversos , Hígado Graso/etiología , Fructosa/administración & dosificación , Fructosa/efectos adversos , Mitocondrias Hepáticas/patología , Tejido Adiposo/crecimiento & desarrollo , Análisis de Varianza , Animales , Respiración de la Célula , Carbohidratos de la Dieta/administración & dosificación , Carbohidratos de la Dieta/efectos adversos , Intolerancia a la Glucosa/diagnóstico , Hiperglucemia/etiología , Hiperinsulinismo/etiología , Lípidos/análisis , Hígado/química , Masculino , Potencial de la Membrana Mitocondrial , Mitocondrias Hepáticas/química , Mitocondrias Hepáticas/fisiología , Sobrepeso/etiología , Fosfolípidos/química , Fosfolípidos/clasificación , Fosfolípidos/aislamiento & purificación , Fosfolípidos/metabolismo , Distribución Aleatoria , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo

RESUMEN

The quality of dietary lipids in the maternal diet can programme the offspring to diseases in later life. We investigated whether the maternal intake of palm oil or interesterified fat, substitutes for trans-unsaturated fatty acids (FA), induces metabolic changes in the adult offspring. During pregnancy and lactation, C57BL/6 female mice received normolipidic diets containing partially hydrogenated vegetable fat rich in trans-unsaturated fatty acids (TG), palm oil (PG), interesterified fat (IG) or soyabean oil (CG). After weaning, male offspring from all groups received the control diet until day 110. Plasma glucose and TAG and liver FA profiles were ascertained. Liver mitochondrial function was accessed with high-resolution respirometry by measuring VO2, fluorimetry for detection of hydrogen peroxide (H2O2) production and mitochondrial Ca2+ uptake. The results showed that the IG offspring presented a 20 % increase in plasma glucose and both the IG and TG offspring presented a 2- and 1·9-fold increase in TAG, respectively, when compared with CG offspring. Liver MUFA and PUFA contents decreased in the TG and IG offspring when compared with CG offspring. Liver MUFA content also decreased in the PG offspring. These modifications in FA composition possibly affected liver mitochondrial function, as respiration was impaired in the TG offspring and H2O2 production was higher in the IG offspring. In addition, mitochondrial Ca2+ retention capacity was reduced by approximately 40 and 55 % in the TG and IG offspring, respectively. In conclusion, maternal consumption of trans-unsaturated and interesterified fat affected offspring health by compromising mitochondrial bioenergetics and lipid metabolism in the liver.

Asunto(s)

Metabolismo Energético , Ácidos Grasos/efectos adversos , Lactancia , Hígado/metabolismo , Fenómenos Fisiologicos Nutricionales Maternos , Mitocondrias/metabolismo , Ácidos Grasos trans/efectos adversos , Fenómenos Fisiológicos Nutricionales de los Animales , Animales , Glucemia/metabolismo , Calcio/metabolismo , Grasas de la Dieta/efectos adversos , Grasas de la Dieta/metabolismo , Ácidos Grasos/metabolismo , Ácidos Grasos Monoinsaturados/metabolismo , Ácidos Grasos Insaturados/metabolismo , Femenino , Peróxido de Hidrógeno/metabolismo , Masculino , Ratones Endogámicos C57BL , Consumo de Oxígeno , Aceites de Plantas , Embarazo , Efectos Tardíos de la Exposición Prenatal , Respiración , Ácidos Grasos trans/metabolismo , Triglicéridos/sangre

RESUMEN

Cardiolipin (CL) is a phospholipid found in the outer mitochondrial membrane (OMM) and inner mitochondrial membrane (IMM) in animal cells. Isocitrate dehydrogenase (ICDH) is an important catalytic enzyme that is localized at the cytosol and mitochondria; the metabolic pathway catalyzed by ICDH differs between the OMM and IMM. To estimate the possible role of lipid membrane in the enzymatic activity of NADP+-dependent ICDH, CL-modified liposomes were prepared using CL/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/cholesterol (Ch), and their characteristics were analyzed based on the fluorescent probe method. The relative enzymatic activity of ICDH decreased in the presence of CL/DPPC/Ch=(30/50/20) liposome, whereas activity increased in the presence of CL/DPPC/Ch=(5/75/20) liposome. NADP+ had the greatest substrate affinity and was dominant in the regulation of ICDH activity. Analysis of membrane properties indicated that membranes in CL-modified liposomes were dehydrated by ICDH binding. Using circular dichroism analysis, CL/DPPC/Ch=(30/50/20) liposome induced a conformational change in ICDH, indicating that CL-rich membrane domains could inhibit ICDH activity. These results suggest that lipid membranes, including CL molecules, could act as a platform to regulate ICDH-related metabolic pathways such as the tricarboxylic acid cycle and lipid synthesis.

RESUMEN

Tafazzin (TAZ) is a phospholipid transacylase that catalyzes the remodeling of cardiolipin, a mitochondrial phospholipid required for oxidative phosphorylation. Mutations of TAZ cause Barth syndrome, which is characterized by mitochondrial dysfunction and dilated cardiomyopathy, leading to premature death. However, the molecular mechanisms underlying the cause of mitochondrial dysfunction in Barth syndrome remain poorly understood. Here we investigated the role of TAZ in regulating mitochondrial function and mitophagy. Using primary mouse embryonic fibroblasts (MEFs) with doxycycline-inducible knockdown of Taz, we showed that TAZ deficiency in MEFs caused defective mitophagosome biogenesis, but not other autophagic processes. Consistent with a key role of mitophagy in mitochondria quality control, TAZ deficiency in MEFs also led to impaired oxidative phosphorylation and severe oxidative stress. Together, these findings provide key insights on mitochondrial dysfunction in Barth syndrome, suggesting that pharmacological restoration of mitophagy may provide a novel treatment for this lethal condition.

Asunto(s)

Autofagia/fisiología , Cardiolipinas/metabolismo , Mitofagia/fisiología , Factores de Transcripción/metabolismo , Aciltransferasas , Animales , Autofagia/genética , Cardiomiopatía Dilatada/genética , Cardiomiopatía Dilatada/metabolismo , Cardiomiopatía Dilatada/patología , Humanos , Ratones , Mitocondrias/genética , Mutación/genética , Factores de Transcripción/deficiencia , Factores de Transcripción/genética

RESUMEN

An exposure of the yeast Saccharomyces cerevisiae to exogenous palmitoleic acid (POA) elicits "liponecrosis," a mode of programmed cell death (PCD) which differs from the currently known PCD subroutines. Here, we report the following mechanism for liponecrotic PCD. Exogenously added POA is incorporated into POA-containing phospholipids that then amass in the endoplasmic reticulum membrane, mitochondrial membranes and the plasma membrane. The buildup of the POA-containing phospholipids in the plasma membrane reduces the level of phosphatidylethanolamine in its extracellular leaflet, thereby increasing plasma membrane permeability for small molecules and committing yeast to liponecrotic PCD. The excessive accumulation of POA-containing phospholipids in mitochondrial membranes impairs mitochondrial functionality and causes the excessive production of reactive oxygen species in mitochondria. The resulting rise in cellular reactive oxygen species above a critical level contributes to the commitment of yeast to liponecrotic PCD by: (1) oxidatively damaging numerous cellular organelles, thereby triggering their massive macroautophagic degradation; and (2) oxidatively damaging various cellular proteins, thus impairing cellular proteostasis. Several cellular processes in yeast exposed to POA can protect cells from liponecrosis. They include: (1) POA oxidation in peroxisomes, which reduces the flow of POA into phospholipid synthesis pathways; (2) POA incorporation into neutral lipids, which prevents the excessive accumulation of POA-containing phospholipids in cellular membranes; (3) mitophagy, a selective macroautophagic degradation of dysfunctional mitochondria, which sustains a population of functional mitochondria needed for POA incorporation into neutral lipids; and (4) a degradation of damaged, dysfunctional and aggregated cytosolic proteins, which enables the maintenance of cellular proteostasis.

Asunto(s)

Ácidos Grasos Monoinsaturados/toxicidad , Lípidos de la Membrana/metabolismo , Necrosis/inducido químicamente , Necrosis/metabolismo , Muerte Celular/efectos de los fármacos , Muerte Celular/fisiología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Relación Dosis-Respuesta a Droga , Necrosis/patología , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/metabolismo

RESUMEN

Emerging evidence indicates that mitochondrial cardiolipins (CL) are prone to free radical oxidation and this process appears to be intimately associated with multiple biological functions of mitochondria. Our previous work demonstrated that a significant amount of potent lipid electrophiles including 4-hydroxy-nonenal (4-HNE) was generated from CL oxidation through a novel chemical mechanism. Here we provide further evidence that a characteristic class of CL oxidation products, epoxyalcohol-aldehyde-CL (EAA-CL), is formed through this novel mechanism in isolated mice liver mitochondria when treated with the pro-apoptotic protein t-Bid to induce cyt c release. Generation of these oxidation products are dose-dependently attenuated by a peroxidase inhibitor acetaminophen (ApAP). Using a mouse model of atherosclerosis, we detected significant amount of these CL oxidation products in liver tissue of low density lipoprotein receptor knockout (LDLR -/-) mice after Western diet feeding. Our studies highlight the importance of lipid electrophiles formation from CL oxidation in the settings of apoptosis and atherosclerosis as inhibition of CL oxidation and lipid electrophiles formation may have potential therapeutic value in diseases linked to oxidant stress and mitochondrial dysfunctions.

Asunto(s)

Apoptosis , Cardiolipinas/química , Mitocondrias/metabolismo , Acetaminofén/farmacología , Animales , Apoptosis/efectos de los fármacos , Arteriosclerosis/metabolismo , Arteriosclerosis/patología , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/genética , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/farmacología , Cardiolipinas/análisis , Cromatografía Líquida de Alta Presión , Dieta Alta en Grasa , Hígado/metabolismo , Espectrometría de Masas , Ratones , Ratones Noqueados , Oxidación-Reducción/efectos de los fármacos , Receptores de LDL/deficiencia , Receptores de LDL/genética , Receptores de LDL/metabolismo

RESUMEN

A recent study revealed a mechanism of delaying aging in yeast by a natural compound which specifically impacts mitochondrial redox processes. In this mechanism, exogenously added lithocholic bile acid enters yeast cells, accumulates mainly in the inner mitochondrial membrane, and elicits an age-related remodeling of phospholipid synthesis and movement within both mitochondrial membranes. Such remodeling of mitochondrial phospholipid dynamics progresses with the chronological age of a yeast cell and ultimately causes significant changes in mitochondrial membrane lipidome. These changes in the composition of membrane phospholipids alter mitochondrial abundance and morphology, thereby triggering changes in the age-related chronology of such longevity-defining redox processes as mitochondrial respiration, the maintenance of mitochondrial membrane potential, the preservation of cellular homeostasis of mitochondrially produced reactive oxygen species, and the coupling of electron transport to ATP synthesis.

Asunto(s)

Ácido Litocólico/farmacología , Mitocondrias/metabolismo , Fosfolípidos/metabolismo , Levaduras/citología , Levaduras/fisiología , Adenosina Trifosfato/metabolismo , Envejecimiento , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Tamaño de los Orgánulos , Oxidación-Reducción/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo

RESUMEN

We report changes of the content of anionic phospholipids in Bacillus subtilis in response to hypoxic conditions and inhibition of terminal respiration. Cardiolipin accumulates rapidly when bacteria are suspended in non-growth medium under reduced aeration or exposed to the inhibitor cyanide; the increase of cardiolipin occurs at the expense of its precursor phosphatidylglycerol and is temperature-dependent. Depending on the extent of hypoxic stress, membranes containing different levels of cardiolipin can be isolated from B. subtilis cells. The NADH oxidase activity in cardiolipin-enriched membranes is cyanide-resistant; furthermore O2 consumption measurements indicated that cardiolipin-enriched cells are resistant to cyanide. Results point out a possible interdependence between the effect of cyanide on cardiolipin metabolism and the effect of cardiolipin on the effectiveness of cyanide inhibition.