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1.
Int J Mol Sci ; 22(2)2021 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-33430070

RESUMEN

The nosocomial opportunistic Gram-negative bacterial pathogen Acinetobacter baumannii is resistant to multiple antimicrobial agents and an emerging global health problem. The polymyxin antibiotic colistin, targeting the negatively charged lipid A component of the lipopolysaccharide on the bacterial cell surface, is often considered as the last-resort treatment, but resistance to colistin is unfortunately increasing worldwide. Notably, colistin-susceptible A. baumannii can also develop a colistin dependence after exposure to this drug in vitro. Colistin dependence might represent a stepping stone to resistance also in vivo. However, the mechanisms are far from clear. To address this issue, we combined proteogenomics, high-resolution microscopy, and lipid profiling to characterize and compare A. baumannii colistin-susceptible clinical isolate (Ab-S) of to its colistin-dependent subpopulation (Ab-D) obtained after subsequent passages in moderate colistin concentrations. Incidentally, in the colistin-dependent subpopulation the lpxA gene was disrupted by insertion of ISAjo2, the lipid A biosynthesis terminated, and Ab-D cells displayed a lipooligosaccharide (LOS)-deficient phenotype. Moreover, both mlaD and pldA genes were perturbed by insertions of ISAjo2 and ISAba13, and LOS-deficient bacteria displayed a capsule with decreased thickness as well as other surface imperfections. The major changes in relative protein abundance levels were detected in type 6 secretion system (T6SS) components, the resistance-nodulation-division (RND)-type efflux pumps, and in proteins involved in maintenance of outer membrane asymmetry. These findings suggest that colistin dependence in A. baumannii involves an ensemble of mechanisms seen in resistance development and accompanied by complex cellular events related to insertional sequences (ISs)-triggered LOS-deficiency. To our knowledge, this is the first study demonstrating the involvement of ISAjo2 and ISAba13 IS elements in the modulation of the lipid A biosynthesis and associated development of dependence on colistin.

2.
Toxins (Basel) ; 12(10)2020 Oct 21.
Artículo en Inglés | MEDLINE | ID: mdl-33096888

RESUMEN

Microcystins (MCs) are hepatotoxins produced by some cyanobacteria. They are cyclic peptides that inhibit the serine/threonine protein phosphatases (PPs) PP1 and PP2A, especially PP2A. The inhibition of PP2A triggers a series of molecular events, which are responsible for most MC cytotoxic and genotoxic effects on animal cells. It is also known that MCs induce oxidative stress in cells due to the production of reactive oxygen species (ROS). However, a complete characterization of the toxic effects of MCs is still not accomplished. This study aimed to clarify additional molecular mechanisms involved in MC-LR toxicity, using Saccharomyces cerevisiae as eukaryotic model organism. First, a shotgun proteomic analysis of S. cerevisiae VL3 cells response to 1 nM, 10 nM, 100 nM, and 1 µM MC-LR was undertaken and compared to the control (cells not exposed to MC-LR). This analysis revealed a high number of proteins differentially expressed related with gene translation and DNA replication stress; oxidative stress; cell cycle regulation and carbohydrate metabolism. Inference of genotoxic effects of S. cerevisiae VL3 cells exposed to different concentrations of MC-LR were evaluated by analyzing the expression of genes Apn1, Apn2, Rad27, Ntg1, and Ntg2 (from the Base Excision Repair (BER) DNA repair system) using the Real-Time RT-qPCR technique. These genes displayed alterations after exposure to MC-LR, particularly the Apn1/Apn2/Rad27, pointing out effects of MC-LR in the Base Excision Repair system (BER). Overall, this study supports the role of oxidative stress and DNA replication stress as important molecular mechanisms of MC-LR toxicity. Moreover, this study showed that even at low-concentration, MC-LR can induce significant changes in the yeast proteome and in gene expression.

3.
Cell Death Dis ; 11(10): 825, 2020 Oct 03.
Artículo en Inglés | MEDLINE | ID: mdl-33011746

RESUMEN

Caspase-2 is a unique and conservative cysteine protease which plays an important role in several cellular processes including apoptotic cell death. Although the molecular mechanisms of its activation remain largely unclear, a major role belongs to the architecture of the caspase-2 active center. We demonstrate that the substitution of the putative phosphorylation site of caspase-2, Serine-384 to Alanine, blocks caspase-2 processing and decreases its enzymatic activity. Strikingly, in silico analysis using molecular dynamics simulations has shown that Serine-384 is crucially involved in interactions within the caspase-2 active center. It stabilizes Arginine-378, which forms a crucial hydrogen bond with the aspartate residue of a substrate. Hence, Serine-384 is essential for supporting a proper architecture of the active center of caspase-2. Moreover, molecular modeling strongly proved steric inaccessibility of Ser-384 to be phosphorylated. Importantly, a multiple alignment has demonstrated that both Serine-384 and Arg-378 residues are highly conservative across all members of caspase family, which allows us to suggest that this diade is indispensable for caspase processing and activity. Spontaneous mutations in this diade might influence oncosuppressive function of caspases, in particular of caspase-2. Likewise, the mutation of Ser-384 is associated with the development of lung squamous cell carcinoma and adenocarcinoma. Taken together, we have uncovered a central feature of the caspase-2 activation mechanism which is crucial for the regulation of its signaling network.

4.
Biomolecules ; 10(8)2020 07 30.
Artículo en Inglés | MEDLINE | ID: mdl-32751491

RESUMEN

The development of alternative ecological and effective antifouling technologies is still challenging. Synthesis of nature-inspired compounds has been exploited, given the potential to assure commercial supplies of potential ecofriendly antifouling agents. In this direction, the antifouling activity of a series of nineteen synthetic small molecules, with chemical similarities with natural products, were exploited in this work. Six (4, 5, 7, 10, 15 and 17) of the tested xanthones showed in vivo activity toward the settlement of Mytilus galloprovincialis larvae (EC50: 3.53-28.60 µM) and low toxicity to this macrofouling species (LC50 > 500 µM and LC50/EC50: 17.42-141.64), and two of them (7 and 10) showed no general marine ecotoxicity (<10% of Artemia salina mortality) after 48 h of exposure. Regarding the mechanism of action in mussel larvae, the best performance compounds 4 and 5 might be acting by the inhibition of acetylcholinesterase activity (in vitro and in silico studies), while 7 and 10 showed specific targets (proteomic studies) directly related with the mussel adhesive structure (byssal threads), given by the alterations in the expression of Mytilus collagen proteins (PreCols) and proximal thread proteins (TMPs). A quantitative structure-activity relationship (QSAR) model was built with predictive capacity to enable speeding the design of new potential active compounds.

5.
Front Microbiol ; 11: 1069, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32523583

RESUMEN

Mitochondria play crucial roles in cellular metabolism, signaling, longevity, and immune defense. Recent evidences have revealed that the host microbiota, including bacterial pathogens, impact mitochondrial behaviors and activities in the host. The pathogenicity of Pseudomonas aeruginosa requires quorum sensing (QS) cell-cell communication allowing the bacteria to sense population density and collectively control biofilm development, virulence traits, adaptation and interactions with the host. QS molecules, like N-3-oxo-dodecanoyl-L-homoserine lactone (3O-C12-HSL), can also modulate the behavior of host cells, e.g., epithelial barrier properties and innate immune responses. Here, in two types of cells, fibroblasts and intestinal epithelial cells, we investigated whether and how P. aeruginosa 3O-C12-HSL impacts the morphology of mitochondrial networks and their energetic characteristics, using high-resolution transmission electron microscopy, fluorescence live-cell imaging, assay for mitochondrial bioenergetics, and quantitative mass spectrometry for mitoproteomics and bioinformatics. We found that 3O-C12-HSL induced fragmentation of mitochondria, disruption of cristae and inner membrane ultrastructure, altered major characteristics of respiration and energetics, and decreased mitochondrial membrane potential, and that there are distinct cell-type specific details of these effects. Moreover, this was mechanistically accompanied by differential expression of both common and cell-type specific arrays of components in the mitochondrial proteome involved in their structural organization, electron transport chain complexes and response to stress. We suggest that this effect of 3O-C12-HSL on mitochondria may represent one of the events in the interaction between P. aeruginosa and host mitochondria and may have an impact on the pathogens strategy to hijack host cell activities to support their own survival and spreading.

6.
Toxins (Basel) ; 12(3)2020 03 20.
Artículo en Inglés | MEDLINE | ID: mdl-32245045

RESUMEN

Toxic cyanobacterial blooms are a major contaminant in inland aquatic ecosystems. Furthermore, toxic blooms are carried downstream by rivers and waterways to estuarine and coastal ecosystems. Concerning marine and estuarine animal species, very little is known about how these species are affected by the exposure to freshwater cyanobacteria and cyanotoxins. So far, most of the knowledge has been gathered from freshwater bivalve molluscs. This work aimed to infer the sensitivity of the marine mussel Mytilus galloprovincialis to single as well as mixed toxic cyanobacterial cultures and the underlying molecular responses mediated by toxic cyanobacteria. For this purpose, a mussel exposure experiment was outlined with two toxic cyanobacteria species, Microcystis aeruginosa and Chrysosporum ovalisporum at 1 × 105 cells/mL, resembling a natural cyanobacteria bloom. The estimated amount of toxins produced by M. aeruginosa and C. ovalisporum were respectively 0.023 pg/cell of microcystin-LR (MC-LR) and 7.854 pg/cell of cylindrospermopsin (CYN). After 15 days of exposure to single and mixed cyanobacteria, a depuration phase followed, during which mussels were fed only non-toxic microalga Parachlorella kessleri. The results showed that the marine mussel is able to filter toxic cyanobacteria at a rate equal or higher than the non-toxic microalga P. kessleri. Filtration rates observed after 15 days of feeding toxic microalgae were 1773.04 mL/ind.h (for M. aeruginosa), 2151.83 mL/ind.h (for C. ovalisporum), 1673.29 mL/ind.h (for the mixture of the 2 cyanobacteria) and 2539.25 mL/ind.h (for the non-toxic P. kessleri). Filtering toxic microalgae in combination resulted in the accumulation of 14.17 ng/g dw MC-LR and 92.08 ng/g dw CYN. Other physiological and biochemical endpoints (dry weight, byssus production, total protein and glycogen) measured in this work did not change significantly in the groups exposed to toxic cyanobacteria with regard to control group, suggesting that mussels were not affected with the toxic microalgae. Nevertheless, proteomics revealed changes in metabolism of mussels related to diet, specially evident in those fed on combined cyanobacteria. Changes in metabolic pathways related with protein folding and stabilization, cytoskeleton structure, and gene transcription/translation were observed after exposure and feeding toxic cyanobacteria. These changes occur in vital metabolic processes and may contribute to protect mussels from toxic effects of the toxins MC-LR and CYN.

8.
Sci Rep ; 9(1): 9841, 2019 07 08.
Artículo en Inglés | MEDLINE | ID: mdl-31285509

RESUMEN

A large number of natural products have been advocated as anticancer agents. Many of these compounds contain functional groups characterized by chemical reactivity. It is not clear whether distinct mechanisms of action can be attributed to such compounds. We used a chemical library screening approach to demonstrate that a substantial fraction (~20%) of cytotoxic synthetic compounds containing Michael acceptor groups inhibit proteasome substrate processing and induce a cellular response characteristic of proteasome inhibition. Biochemical and structural analyses showed binding to and inhibition of proteasome-associated cysteine deubiquitinases, in particular ubiquitin specific peptidase 14 (USP14). The results suggested that compounds bind to a crevice close to the USP14 active site with modest affinity, followed by covalent binding. A subset of compounds was identified where cell death induction was closely associated with proteasome inhibition and that showed significant antineoplastic activity in a zebrafish embryo model. These findings suggest that proteasome inhibition is a relatively common mode of action by cytotoxic compounds containing Michael acceptor groups and help to explain previous reports on the antineoplastic effects of natural products containing such functional groups.


Asunto(s)
Antineoplásicos/administración & dosificación , Inhibidores de Proteasoma/administración & dosificación , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Ubiquitina Tiolesterasa/metabolismo , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Dominio Catalítico , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Células HCT116 , Células HeLa , Humanos , Células MCF-7 , Complejo de la Endopetidasa Proteasomal/química , Complejo de la Endopetidasa Proteasomal/metabolismo , Inhibidores de Proteasoma/química , Inhibidores de Proteasoma/farmacología , Unión Proteica , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Ubiquitina Tiolesterasa/química , Ensayos Antitumor por Modelo de Xenoinjerto , Pez Cebra
9.
J Innate Immun ; 11(3): 263-279, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-30428481

RESUMEN

Cell-to-cell signaling via small molecules is an essential process to coordinate behavior in single species within a community, and also across kingdoms. In this review, we discuss the quorum sensing (QS) systems used by the opportunistic pathogen Pseudomonas aeruginosa to sense bacterial population density and fitness, and regulate virulence, biofilm development, metabolite acquisition, and mammalian host defense. We also focus on the role of N-acylhomoserine lactone-dependent QS signaling in the modulation of innate immune responses connected together via calcium signaling, homeostasis, mitochondrial and cytoskeletal dynamics, and governing transcriptional and proteomic responses of host cells. A future perspective emphasizes the need for multidisciplinary efforts to bring current knowledge of QS into a more detailed understanding of the communication between bacteria and host, as well as into strategies to prevent and treat P. aeruginosa infections and reduce the rate of antibiotic resistance.


Asunto(s)
Interacciones Microbiota-Huesped , Infecciones por Pseudomonas/etiología , Pseudomonas aeruginosa/fisiología , Percepción de Quorum/fisiología , 4-Butirolactona/análogos & derivados , 4-Butirolactona/fisiología , Adhesión Bacteriana , Biopelículas , Señalización del Calcio , Movimiento Celular , Homoserina/análogos & derivados , Homoserina/fisiología , Humanos , Hierro/metabolismo , Lipopolisacáridos/fisiología , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/patogenicidad
10.
BMC Res Notes ; 11(1): 664, 2018 Sep 12.
Artículo en Inglés | MEDLINE | ID: mdl-30208953

RESUMEN

OBJECTIVE: Cough and fever are the initial symptoms of lower respiratory infection. Severe cases might be fatal. Therefore, particularly in the non-equipped centers, the lack of diagnostic methods to identify the severe cases has resulted in overconsumption of antibiotics. On the basis of the knowledge about non-specific immune response at the site of injury, we developed a colorimetric dip-test that shows abrupt, sensitive and quite specific color change upon contact with sputum in the cases of lower respiratory infection. We further explored the mechanism of the test. RESULTS: We detected deoxyribonucleic acid (DNA) and hepatocyte growth factor in the sputum of patients that suffered from respiratory infection (n = 18). The results differed significantly (P < 0.0001) from age-matched patients (n = 18) with other respiratory disorders and highly correlated with the index-test results (Spearman Rank test = 0.84). DNA with a concentration more than 0.03 mg/ml induced a visible and stable color change on index-test within 1 min. The test recognized all of the cases with respiratory infection and the specificity was 72%. With a high negative predictive value. The index test detects, inter alia, cell-free DNA in sputum and might safely rule-out respiratory infection in 2/3 of cases that present symptoms of acute respiratory infection.


Asunto(s)
Infecciones del Sistema Respiratorio/diagnóstico , Esputo/microbiología , Antibacterianos , Tos , ADN/análisis , Humanos , Sensibilidad y Especificidad , Factores de Tiempo
11.
Mar Drugs ; 16(2)2018 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-29364843

RESUMEN

Cnidarian toxic products, particularly peptide toxins, constitute a promising target for biomedicine research. Indeed, cnidarians are considered as the largest phylum of generally toxic animals. However, research on peptides and toxins of sea anemones is still limited. Moreover, most of the toxins from sea anemones have been discovered by classical purification approaches. Recently, high-throughput methodologies have been used for this purpose but in other Phyla. Hence, the present work was focused on the proteomic analyses of whole-body extract from the unexplored sea anemone Bunodactis verrucosa. The proteomic analyses applied were based on two methods: two-dimensional gel electrophoresis combined with MALDI-TOF/TOF and shotgun proteomic approach. In total, 413 proteins were identified, but only eight proteins were identified from gel-based analyses. Such proteins are mainly involved in basal metabolism and biosynthesis of antibiotics as the most relevant pathways. In addition, some putative toxins including metalloproteinases and neurotoxins were also identified. These findings reinforce the significance of the production of antimicrobial compounds and toxins by sea anemones, which play a significant role in defense and feeding. In general, the present study provides the first proteome map of the sea anemone B. verrucosa stablishing a reference for future studies in the discovery of new compounds.


Asunto(s)
Proteómica , Anémonas de Mar/genética , Animales , Biología Computacional , Ontología de Genes , Metaloproteasas/biosíntesis , Metaloproteasas/química , Pruebas de Sensibilidad Microbiana , Neurotoxinas/biosíntesis , Neurotoxinas/química , Péptidos/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Extractos de Tejidos/química
12.
Front Cell Dev Biol ; 5: 98, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-29209610

RESUMEN

Normal epithelial and endothelial renewal and healing after bacterial and viral challenges are essential for homeostasis along the intestine and the blood and lymphatic vessels. We thus investigated whether and how virus affects migration of human epithelial cells and specifically how the nucleocapsid protein (N) modulates the cellular proteome and interactome using human Caco-2 cells in a wound-healing assay with Hazara virus as a model. Here, Hazara virus blocked cell migration in a dose- and time-dependent manner, disrupted the actin cytoskeleton and specifically reduced the expression of the IQ-motif-containing GTPase-activating protein 1 (IQGAP1) and water channel aquaporin 6 (AQP6) that regulate cytoskeletal organization, water homeostasis and vesicle communication. Moreover, in the Caco-2 cell proteome, we identified several distinct groups of molecules associating with N upon Hazara virus infection, being involved in the ensemble of important cellular processes, e.g., chaperone activity, metabolism, cellular defense against infections, cell morphology, and migration. These events do not only facilitate the virus life cycle, but they are also crucial for membrane and cytoskeleton dynamics, cellular self-renewal and wound healing, being so essential for body integrity and homeostasis.

13.
Sci Rep ; 7: 43512, 2017 03 07.
Artículo en Inglés | MEDLINE | ID: mdl-28266628

RESUMEN

Microdialysis (MD) has been shown to be a promising technique for sampling of biomarkers. Implantation of MD probe causes an acute tissue trauma and provokes innate response cascades. In order to normalize tissue a two hours equilibration period for analysis of small molecules has been reported previously. However, how the proteome profile changes due to this acute trauma has yet to be fully understood. To characterize the early proteome events induced by this trauma we compared proteome in muscle dialysate collected during the equilibration period with two hours later in "post-trauma". Samples were collected from healthy females using a 100 kDa MW cut off membrane and analyzed by high sensitive liquid chromatography tandem mass spectrometry. Proteins involved in stress response, immune system processes, inflammatory responses and nociception from extracellular and intracellular fluid spaces were identified. Sixteen proteins were found to be differentially abundant in samples collected during first two hours in comparison to "post-trauma". Our data suggests that microdialysis in combination with mass spectrometry may provide potentially new insights into the interstitial proteome of trapezius muscle, yet should be further adjusted for biomarker discovery and diagnostics. Moreover, MD proteome alterations in response to catheter injury may reflect individual innate reactivity.


Asunto(s)
Líquido Extracelular/metabolismo , Microdiálisis , Proteoma , Proteómica , Músculos Superficiales de la Espalda/lesiones , Músculos Superficiales de la Espalda/metabolismo , Adulto , Biomarcadores , Biología Computacional/métodos , Femenino , Humanos , Microdiálisis/métodos , Persona de Mediana Edad , Anotación de Secuencia Molecular , Mapeo de Interacción de Proteínas , Mapas de Interacción de Proteínas , Proteómica/métodos
15.
Biochem J ; 473(19): 3177-88, 2016 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-27458251

RESUMEN

The ubiquitously expressed IQ motif-containing GTPase activating protein-1 (IQGAP1) is a scaffolding protein implicated in an array of cellular functions, in particular by binding to cytoskeletal elements and signaling proteins. A role of IQGAP1 in adipocytes has not been reported. We therefore investigated the cellular IQGAP1 interactome in primary human adipocytes. Immunoprecipitation and quantitative mass spectrometry identified caveolae and caveolae-associated proteins as the major IQGAP1 interactors alongside cytoskeletal proteins. We confirmed co-localization of IQGAP1 with the defining caveolar marker protein caveolin-1 by confocal microscopy and proximity ligation assay. Most interestingly, insulin enhanced the number of IQGAP1 interactions with caveolin-1 by five-fold. Moreover, we found a significantly reduced abundance of IQGAP1 in adipocytes from patients with type 2 diabetes compared with cells from nondiabetic control subjects. Both the abundance of IQGAP1 protein and mRNA were reduced, indicating a transcriptional defect in diabetes. Our findings suggest a novel role of IQGAP1 in insulin-regulated interaction between caveolae and cytoskeletal elements of the adipocyte, and that this is quelled in the diabetic state.


Asunto(s)
Adipocitos/metabolismo , Caveolas/metabolismo , Citoesqueleto/metabolismo , Insulina/metabolismo , Proteínas Activadoras de ras GTPasa/metabolismo , Adipocitos/citología , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Fosforilación
16.
FEMS Microbiol Lett ; 362(11)2015 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-25956174

RESUMEN

Helicobacter pylori produces outer membrane vesicles (OMV), delivering bacterial substances including the oncogenic cytotoxin-associated CagA protein to their surroundings. We investigated the effects of H. pylori OMV carrying CagA (OMV-CagA) on cell junctions and ATP-binding proteome of epithelial monolayers, using proteomics, mass spectrometry and imaging. OMV-CagA localized in close vicinity of ZO-1 tight junction protein and induced histone H1 binding to ATP. We suggest the expression of novel events in the interactions between H. pylori OMV and epithelia, which may have an influence on host gene transcription and lead to different outcomes of an infection and development of cancer.


Asunto(s)
Adenosina Trifosfato/metabolismo , Antígenos Bacterianos/metabolismo , Proteínas Bacterianas/metabolismo , Helicobacter pylori/metabolismo , Histonas/metabolismo , Antígenos Bacterianos/genética , Proteínas Bacterianas/genética , Células CACO-2 , Helicobacter pylori/genética , Interacciones Huésped-Patógeno , Humanos , Microscopía Confocal , Uniones Estrechas/metabolismo , Uniones Estrechas/ultraestructura , Proteína de la Zonula Occludens-1/metabolismo
17.
Methods Mol Biol ; 1306: 121-34, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25930698

RESUMEN

This chapter describes different strategies for the study of phosphorylation dynamics and stoichiometry in photosynthetic membranes. Detailed procedures for the detection, large-scale identification, and quantification of phosphorylated proteins optimized for plant thylakoid proteins are given.


Asunto(s)
Arabidopsis/metabolismo , Fosfoproteínas/análisis , Proteómica/métodos , Tilacoides/metabolismo , Proteínas de Arabidopsis/metabolismo , Fosforilación , Fotosíntesis
18.
Pain Med ; 15(8): 1379-89, 2014 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-24995488

RESUMEN

PURPOSE: Chronic neck/shoulder pain (CNSP) is one of the most common pain conditions. The understanding of mechanisms, including the peripheral balance between nociceptive and antinociceptive processes, is incomplete. N-acylethanolamines (NAEs) are a class of endogenous compounds that regulate inflammation and pain. The aim of this study was to investigate the levels of two NAEs: the peroxisome proliferator-activated receptor type-α ligand palmitoylethanolamide (PEA) and stearoylethanolamide (SEA) in the muscle interstitium of the trapezius muscle in women with CNSP randomized to two different neck specific training programs and in a healthy pain-free control group (CON). MATERIALS AND METHODS: Fifty-seven women with CNSP were randomized to strength + stretch or stretch alone exercise programs. Twenty-nine subjects underwent microdialysis procedure before and after 4-6 months of exercise. Twenty-four CON subjects underwent microdialysis procedure before and after 4-6 months without any intervention in between. Microdialysate samples were collected from the trapezius muscle and analyzed by mass spectrometry for PEA and SEA levels. RESULTS: PEA and SEA levels were significantly higher in CNSP patients compared with CON. PEA was significantly higher in CNSP than in CON after both training programs. SEA was significantly higher in CNSP than in CON after stretch alone but not after strength + stretch training. A significant positive correlation was found between changes in pain intensity and in SEA levels in the strength + stretch group, but not in the stretch alone group. CONCLUSION: Our results indicate that exercise interventions differentially affect the levels of the bioactive lipids PEA and SEA in the interstitium of the trapezius muscle in women with CNSP.


Asunto(s)
Etanolaminas/metabolismo , Terapia por Ejercicio/métodos , Dolor de Cuello/rehabilitación , Ácidos Palmíticos/metabolismo , Dolor de Hombro/rehabilitación , Ácidos Esteáricos/metabolismo , Adulto , Amidas , Dolor Crónico/metabolismo , Dolor Crónico/rehabilitación , Etanolaminas/análisis , Femenino , Humanos , Espectrometría de Masas , Microdiálisis , Persona de Mediana Edad , Dolor de Cuello/metabolismo , Ácidos Palmíticos/análisis , Dolor de Hombro/metabolismo , Ácidos Esteáricos/análisis , Músculos Superficiales de la Espalda/química , Músculos Superficiales de la Espalda/metabolismo
19.
Proteomics Clin Appl ; 8(3-4): 241-50, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24273187

RESUMEN

PURPOSE: Mutations in apolipoprotein A-I (apoA-I) may affect plasma high-density lipoprotein (HDL) cholesterol levels and the risk for cardiovascular disease but little is known about the presence and effects of circulating apoA-I variants. This study investigates whether the apoA-I mutations, apoA-I(L202P) and apoA-I(K131del) , are present on plasma HDL particles derived from heterozygote carriers and whether this is associated to changes in HDL protein composition. EXPERIMENTAL DESIGN: Plasma HDL of heterozygotes for either apoA-I(L202P) or apoA-I(K131del) and family controls was isolated using ultracentrifugation. HDL proteins were separated by 2DE and analyzed by MS. RESULTS: ApoA-I peptides containing apoA-I(L202P) or apoA-I(K131del) were identified in HDL from heterozygotes. The apoA-I(L202P) mutant peptide was less abundant than wild-type peptide while the apoA-I(K131del) mutant peptide was more abundant than wild-type peptide in the heterozygotes. Two-dimensional gel electrophoresis analyses indicated that, compared to controls, HDL in apoA-I(L202P) carriers contained less apoE and more zinc-α-2-glycoprotein while HDL from the apoA-I(K131del) heterozygotes contained more alpha-1-antitrypsin and transthyretin. CONCLUSIONS AND CLINICAL RELEVANCE: Both apoA-I(L202P) and apoA-I(K131del) were identified in HDL. In heterozygotes, these mutations have markedly differential effects on the concentration of wild-type apoA-I in the circulation, as well as the HDL proteome, both of which might affect the clinical phenotype encountered in the heterozygous carriers.


Asunto(s)
Apolipoproteína A-I/genética , Enfermedades Cardiovasculares/genética , HDL-Colesterol/sangre , Mutación , Apolipoproteína A-I/sangre , Enfermedades Cardiovasculares/sangre , Enfermedades Cardiovasculares/patología , Electroforesis en Gel Bidimensional , Heterocigoto , Humanos , Fenotipo , Prealbúmina/metabolismo , Proteínas de Plasma Seminal/sangre , alfa 1-Antitripsina/sangre
20.
PLoS One ; 8(10): e76526, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24130779

RESUMEN

Astrocytes are highly involved in regulation and homeostasis of the extracellular environment in the healthy brain. In pathological conditions, these cells play a major role in the inflammatory response seen in CNS tissues, which is called reactive astrogliosis and includes hypertrophy and proliferation of astrocytes. Here, we performed 3D confocal microscopy to evaluate the morphological response of reactive astrocytes positive for glial fibrillary acidic protein (GFAP) in rats, to the presence of Aß(1-40) in the rat brain before and after treatment with genistein. In 50 astrocytes per animal, we measured the volume and surface area for the nucleus, cell body, the entire cell, the tissue covered by single astrocytes and quantified the number and length of branches, the density of the astrocytes and the intensity of GFAP immunoreactivity. Injecting Aß(1-40) into the brain of rats caused astrogliosis indicated by increased values for all measured parameters. Mass spectrometric analysis of hippocampal tissue in Aß(1-40)-injected brain showed decreased amounts of tubulins, enolases and myelin basic protein, and increased amounts of dihydropyrimidinase-related protein 2. In Aß(1-40)-injected rats pretreated with genistein, GFAP intensity was decreased to the sham-operated group level, and Aß(1-40)-induced astrogliosis was significantly ameliorated.


Asunto(s)
Péptidos beta-Amiloides/farmacología , Genisteína/farmacología , Genisteína/uso terapéutico , Gliosis/tratamiento farmacológico , Gliosis/patología , Imagenología Tridimensional , Fragmentos de Péptidos/farmacología , Proteoma/efectos de los fármacos , Animales , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Astrocitos/patología , Recuento de Células , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Tamaño de la Célula/efectos de los fármacos , Proteína Ácida Fibrilar de la Glía/metabolismo , Gliosis/inducido químicamente , Gliosis/metabolismo , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Masculino , Microscopía Confocal , Proteoma/metabolismo , Proteómica , Ratas , Ratas Wistar
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