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1.
J Biol Chem ; 292(17): 6965-6977, 2017 04 28.
Artículo en Inglés | MEDLINE | ID: mdl-28258215

RESUMEN

ABCD1 and its homolog ABCD2 are peroxisomal ATP-binding cassette (ABC) half-transporters of fatty acyl-CoAs with both distinct and overlapping substrate specificities. Although it is established that ABC half-transporters have at least to dimerize to generate a functional unit, functional equivalents of tetramers (i.e. dimers of full-length transporters) have also been reported. However, oligomerization of peroxisomal ABCD transporters is incompletely understood but is of potential significance because more complex oligomerization might lead to differences in substrate specificity. In this work, we have characterized the quaternary structure of the ABCD1 and ABCD2 proteins in the peroxisomal membrane. Using various biochemical approaches, we clearly demonstrate that both transporters exist as both homo- and heterotetramers, with a predominance of homotetramers. In addition to tetramers, some larger molecular ABCD assemblies were also found but represented only a minor fraction. By using quantitative co-immunoprecipitation assays coupled with tandem mass spectrometry, we identified potential binding partners of ABCD2 involved in polyunsaturated fatty-acid metabolism. Interestingly, we identified calcium ATPases as ABCD2-binding partners, suggesting a role of ABCD2 in calcium signaling. In conclusion, we have shown here that ABCD1 and its homolog ABCD2 exist mainly as homotetramers in the peroxisomal membrane.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Peroxisomas/metabolismo , Subfamilia D de Transportadores de Casetes de Unión al ATP , Miembro 1 de la Subfamilia D de Transportador de Casetes de Unión al ATP , Adenosina Trifosfato/metabolismo , Animales , Células COS , Señalización del Calcio , ATPasas Transportadoras de Calcio/metabolismo , Carcinoma Hepatocelular/metabolismo , Línea Celular , Chlorocebus aethiops , Proteínas Fluorescentes Verdes/metabolismo , Neoplasias Hepáticas/metabolismo , Espectrometría de Masas , Ratones , Unión Proteica , Mapeo de Interacción de Proteínas , Estructura Cuaternaria de Proteína , Transporte de Proteínas , Ratas , Espectrometría de Masas en Tándem
2.
Mol Plant Microbe Interact ; 28(11): 1227-36, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26106900

RESUMEN

Stomata remain abnormally opened and unresponsive to abscisic acid in grapevine leaves infected by downy mildew. This deregulation occurs from 3 days postinoculation and increases concomitantly with leaf colonization by the pathogen. Using epidermal peels, we demonstrated that the active compound involved in this deregulation is located in the apoplast. Biochemical assays showed that the active compound present in the apoplastic fluids isolated from Plasmopara viticola-infected grapevine leaves (IAF) is a CysCys bridge-independent, thermostable and glycosylated protein. Fractionation guided assays based on chromatography coupled to stomatal response and proteomic analysis allowed the identification of both plant and pathogen proteins in the active fraction obtained from IAF. Further in silico analysis and discriminant filtrations based on the comparison between predictions and experimental indications lead to the identification of two Vitis vinifera proteins as candidates for the observed stomatal deregulation.


Asunto(s)
Glicoproteínas/metabolismo , Oomicetos/metabolismo , Hojas de la Planta/metabolismo , Proteínas de Plantas/metabolismo , Estomas de Plantas/metabolismo , Vitis/metabolismo , Secuencia de Aminoácidos , Pared Celular/genética , Pared Celular/metabolismo , Pared Celular/microbiología , Cromatografía por Intercambio Iónico , Simulación por Computador , Proteínas Fúngicas/metabolismo , Glicoproteínas/clasificación , Glicoproteínas/genética , Interacciones Huésped-Patógeno , Datos de Secuencia Molecular , Oomicetos/fisiología , Filogenia , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiología , Epidermis de la Planta/genética , Epidermis de la Planta/metabolismo , Epidermis de la Planta/microbiología , Hojas de la Planta/genética , Hojas de la Planta/microbiología , Proteínas de Plantas/clasificación , Proteínas de Plantas/genética , Estomas de Plantas/genética , Estomas de Plantas/microbiología , Proteómica/métodos , Homología de Secuencia de Aminoácido , Vitis/genética , Vitis/microbiología
3.
J Intensive Med ; 3(2): 147-154, 2023 Apr 30.
Artículo en Inglés | MEDLINE | ID: mdl-37188118

RESUMEN

Background: This study aimed to investigate renal replacement therapy (RRT) practices in a representative nationwide sample of French intensive care units (ICUs). Methods: From July 1 to October 5 2021, 67 French ICUs provided data regarding their ICU and RRT implementation. We used an online questionnaire to record general data about each participating ICU, including the type of hospital, number of beds, staff ratios, and RRT implementation. Each center then prospectively recorded RRT parameters from 5 consecutive acute kidney injury (AKI) patients, namely the indication, type of dialysis catheter used, type of catheter lock used, type of RRT (continuous or intermittent), the RRT parameters initially prescribed (dose, blood flow, and duration), and the anticoagulant agent used for the circuit. Results: A total of 303 patients from 67 ICUs were analyzed. Main indications for RRT were oligo-anuria (57.4%), metabolic acidosis (52.1%), and increased plasma urea levels (47.9%). The commonest insertion site was the right internal jugular (45.2%). In 71.0% of cases, the dialysis catheter was inserted by a resident. Ultrasound guidance was used in 97.0% and isovolumic connection in 90.1%. Citrate, unfractionated heparin, and saline were used as catheter locks in 46.9%, 24.1%, and 21.1% of cases, respectively. Conclusions: Practices in French ICUs are largely compliant with current national guidelines and international literature. The findings should be interpreted in light of the limitations inherent to this type of study.

4.
Blood ; 115(1): 78-88, 2010 Jan 07.
Artículo en Inglés | MEDLINE | ID: mdl-19864642

RESUMEN

Chronic myelomonocytic leukemia (CMML) is a clonal hematopoietic disorder that occurs in elderly patients. One of the main diagnostic criteria is the accumulation of heterogeneous monocytes in the peripheral blood. We further explored this cellular heterogeneity and observed that part of the leukemic clone in the peripheral blood was made of immature dysplastic granulocytes with a CD14(-)/CD24(+) phenotype. The proteome profile of these cells is dramatically distinct from that of CD14(+)/CD24(-) monocytes from CMML patients or healthy donors. More specifically, CD14(-)/CD24(+) CMML cells synthesize and secrete large amounts of alpha-defensin 1-3 (HNP1-3). Recombinant HNPs inhibit macrophage colony-stimulating factor (M-CSF)-driven differentiation of human peripheral blood monocytes into macrophages. Using transwell, antibody-mediated depletion, suramin inhibition of purinergic receptors, and competitive experiments with uridine diphosphate (UDP)/uridine triphosphate (UTP), we demonstrate that HNP1-3 secreted by CD14(-)/CD24(+) cells inhibit M-CSF-induced differentiation of CD14(+)/CD24(-) cells at least in part through P2Y6, a receptor involved in macrophage differentiation. Altogether, these observations suggest that a population of immature dysplastic granulocytes contributes to the CMML phenotype through production of alpha-defensins HNP1-3 that suppress the differentiation capabilities of monocytes.


Asunto(s)
Diferenciación Celular , Granulocitos/metabolismo , Granulocitos/patología , Leucemia Mielomonocítica Crónica/patología , Monocitos/patología , alfa-Defensinas/metabolismo , Antígeno CD24/metabolismo , Diferenciación Celular/efectos de los fármacos , Citocinas/biosíntesis , Granulocitos/efectos de los fármacos , Humanos , Leucemia Mielomonocítica Crónica/metabolismo , Receptores de Lipopolisacáridos/metabolismo , Factor Estimulante de Colonias de Macrófagos/farmacología , Macrófagos/citología , Macrófagos/efectos de los fármacos , Monocitos/efectos de los fármacos , Receptores Purinérgicos P2/metabolismo , Uridina Difosfato/farmacología , Uridina Trifosfato/farmacología , alfa-Defensinas/farmacología
5.
Med Sci (Paris) ; 23 Spec No 1: 19-22, 2007 Mar.
Artículo en Francés | MEDLINE | ID: mdl-17669348

RESUMEN

Proteins identified in biological fluids of cancer patients could be helpful for both diagnosis and prognosis. However, clinical proteomics based on analysis of protein profiles in biological fluids has demonstrated various flaws, most of them related to the difficulties met in reproducibility. These difficulties could be partly overcome by accurate standardisation of pre-analytical and analytical steps of these studies. The size of the patient cohort is one of the parameters that determine the powerfulness of the study. Recruitment of a cohort with a sufficient size often implies multicentric studies in which analysis of the reproducibility between centres and standardisation of pre-analytical and analytical steps are essential. Such a standardisation requires the use of calibrated samples as common references.


Asunto(s)
Líquidos Corporales/química , Proteínas de Neoplasias/análisis , Proteómica/normas , Manejo de Especímenes/normas , Biomarcadores de Tumor/análisis , Análisis Químico de la Sangre/instrumentación , Análisis Químico de la Sangre/normas , Calibración , Humanos , Estudios Multicéntricos como Asunto/métodos , Estudios Multicéntricos como Asunto/estadística & datos numéricos , Neoplasias/sangre , Estándares de Referencia , Reproducibilidad de los Resultados , Tamaño de la Muestra , Manejo de Especímenes/instrumentación , Manejo de Especímenes/métodos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/instrumentación
6.
Chem Phys Lipids ; 207(Pt B): 179-191, 2017 10.
Artículo en Inglés | MEDLINE | ID: mdl-28576383

RESUMEN

Glaucoma is a progressive and irreversible blinding neuropathy that is characterized by the loss of retinal ganglion cells (RGCs). Muller Glial Cell (MGC) activation is induced in retinal gliosis. MGCs are the most numerous glial cells in the retina and one of their roles is to sustain cholesterol homeostasis. 24S-hydroxycholesterol (24S-OHC) is one of the form of cholesterol elimination from the retina and is overexpressed during glaucoma. The objective of this study was to determine whether 24S-OHC triggers MGC membrane dynamics involving lipid rafts and contributes to gliosis at early and late time points. A proteomic analysis was carried out by nanoLC-MS/MS in raft and non-raft fractions from MGCs after treatment with 24S-OHC (10µM). The expression of structural and functional proteins was further analyzed by Western-blotting, as well as the levels of GM3 ganglioside by LC-MS. Cholesterol, sphingomyelin, saturated fatty acids and ganglioside GM3 are enriched in the rafts fractions in MGCs. Caveolin-1, flotillin-1, connexin-30 and -43 are localized in the MGCs rafts. Proteins implicated in adhesion or oxidative stress pathways in raft fractions were up and down-regulated by the treatment. Our data showed that 24S-OHC induced early changes in protein distribution in raft microdomains; however, further studies are needed to better characterize the surrounded mechanisms.


Asunto(s)
Membrana Celular/efectos de los fármacos , Colesterol/metabolismo , Células Ependimogliales/citología , Glaucoma/metabolismo , Hidroxicolesteroles/farmacología , Animales , Membrana Celular/metabolismo , Células Cultivadas , Ratas , Ratas Long-Evans
7.
Materials (Basel) ; 6(11): 4946-4966, 2013 Oct 25.
Artículo en Inglés | MEDLINE | ID: mdl-28788369

RESUMEN

Widely used in microelectronics and optoelectronics; Gallium Arsenide (GaAs) is a III-V crystal with several interesting properties for microsystem and biosensor applications. Among these; its piezoelectric properties and the ability to directly biofunctionalize the bare surface, offer an opportunity to combine a highly sensitive transducer with a specific bio-interface; which are the two essential parts of a biosensor. To optimize the biorecognition part; it is necessary to control protein coverage and the binding affinity of the protein layer on the GaAs surface. In this paper; we investigate the potential of a specific chemical interface composed of thiolate molecules with different chain lengths; possessing hydroxyl (MUDO; for 11-mercapto-1-undecanol (HS(CH2)11OH)) or carboxyl (MHDA; for mercaptohexadecanoic acid (HS(CH2)15CO2H)) end groups; to reconstitute a dense and homogeneous albumin (Rat Serum Albumin; RSA) protein layer on the GaAs (100) surface. The protein monolayer formation and the covalent binding existing between RSA proteins and carboxyl end groups were characterized by atomic force microscopy (AFM) analysis. Characterization in terms of topography; protein layer thickness and stability lead us to propose the 10% MHDA/MUDO interface as the optimal chemical layer to efficiently graft proteins. This analysis was coupled with insitu MALDI-TOF mass spectrometry measurements; which proved the presence of a dense and uniform grafted protein layer on the 10% MHDA/MUDO interface. We show in this study that a critical number of carboxylic docking sites (10%) is required to obtain homogeneous and dense protein coverage on GaAs. Such a protein bio-interface is of fundamental importance to ensure a highly specific and sensitive biosensor.

8.
J Proteomics ; 75(12): 3665-73, 2012 Jun 27.
Artículo en Inglés | MEDLINE | ID: mdl-22575268

RESUMEN

In order to describe developmental changes in human salivary peptidome, whole saliva was obtained from 98 infants followed longitudinally at 3 and 6months of age. Data on teeth eruption and diet at the age of 6months were also recorded. Salivary peptide extracts were characterised by label-free MALDI-MS. Peptides differentially expressed between the two ages, and those significantly affected by teeth eruption or introduction of solid foods were identified by MALDI TOF-TOF and LC ESI MS-MS. Out of 81 peaks retained for statistical analysis, 26 were overexpressed at the age of 6months. Exposure to solid foods had a more pronounced effect on profiles (overexpression of nine peaks) than teeth eruption (overexpression of one peak). Differential peaks corresponded to fragments of acidic and basic PRPs, statherin and histatin. Comparison with existing knowledge on adult saliva peptidome revealed that proteolytic processing of salivary proteins is qualitatively quite comparable in infants and in adults. However, age and diet are modulators of salivary peptidome in human infants.


Asunto(s)
Envejecimiento/metabolismo , Conducta Alimentaria/fisiología , Péptidos/metabolismo , Proteoma/metabolismo , Saliva/metabolismo , Proteínas y Péptidos Salivales/metabolismo , Erupción Dental/fisiología , Adulto , Femenino , Humanos , Lactante , Masculino , Péptidos/análisis , Proteoma/análisis
9.
J Proteomics ; 72(6): 974-81, 2009 Aug 20.
Artículo en Inglés | MEDLINE | ID: mdl-19481188

RESUMEN

This work is a statistical analysis of reproducibility of a MALDI-TOF mass spectrometry experiment. Its aim is to evaluate measurement variability and compare peak intensities from two types of MALDI-TOF platforms. We compared and commented on the abilities of Principal Component Analysis and mixed-model analysis of variance to evaluate the biological variability and the technical variability of peak intensities in different patients. The properties and hypotheses of both methods are summarized and applied to spectra from plasma of patients with Hodgkin lymphoma. Principal Component Analysis checks rapidly the balance between the two variabilities; however, a mixed-model analysis of variance is necessary to quantify the biological and technical components of the experimental variance as well as their interactions and to split the total variance into between-subjects and within-subject components. The latter method helped to assess the reproducibility of measurements from two MALDI-TOF platforms and to decompose the technical variability according to the experimental design.


Asunto(s)
Enfermedad de Hodgkin/metabolismo , Proteómica/métodos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Adsorción , Análisis de Varianza , Proteínas Sanguíneas/química , Computadores , Humanos , Espectrometría de Masas/métodos , Péptidos/química , Análisis de Componente Principal , Proteoma , Control de Calidad , Reproducibilidad de los Resultados , Programas Informáticos
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