RESUMO
The Water Framework Directive (WFD) requires member states to routinely assess the river ecological status using community-based indices. However, there is still a lack of published WFD-compliant methods for the French West Indies, especially using diatom-based indices. Martinique and Guadeloupe exhibit diverse landscapes shaped by their complex geological history and tropical climatic conditions. These strong particularities make the existing indices developed for the European mainland unusable. Based on diatom sampling from to 2013 (607 samples) and through multivariate analyses, we developed the Indice Diatomique des Antilles (IDA). We first identified the key abiotic factors influencing diatom communities on both islands, and then characterized taxon sensitivity by considering their presence probability along a pressure gradient.. The index was based on the presence and relative abundance of these taxa in each sample. The last step consisted of using new data from the 2014-2022 sampling surveys (457 samples) as a validation dataset to verify IDA accuracy. Our results suggest that the IDA methodology is well designed to assess the ecological status of rivers in the West Indies.
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Diatomáceas , Monitoramento Ambiental , Rios , Diatomáceas/classificação , Rios/química , Monitoramento Ambiental/métodos , Guadalupe , Ecossistema , Biodiversidade , Martinica , Índias OcidentaisRESUMO
Molecular imaging holds great promise in the noninvasive monitoring of several diseases with nanoparticles (NPs) being considered an efficient imaging tool for cancer, central nervous system, and heart- or bone-related diseases and for disorders of the mononuclear phagocytic system (MPS). In the present study, we used an iron-based nanoformulation, already established as an MRI/SPECT probe, as well as to load different biomolecules, to investigate its potential for nuclear planar and tomographic imaging of several target tissues following its distribution via different administration routes. Iron-doped hydroxyapatite NPs (FeHA) were radiolabeled with the single photon γ-emitting imaging agent [99mTc]TcMDP. Administration of the radioactive NPs was performed via the following four delivery methods: (1) standard intravenous (iv) tail vein, (2) iv retro-orbital injection, (3) intratracheal (it) instillation, and (4) intrarectal installation (pr). Real-time, live, fast dynamic screening studies were performed on a dedicated bench top, mouse-sized, planar SPECT system from t = 0 to 1 hour postinjection (p.i.), and consequently, tomographic SPECT/CT imaging was performed, for up to 24 hours p.i. The administration routes that have been studied provide a wide range of possible target tissues, for various diseases. Studies can be optimized following this workflow, as it is possible to quickly assess more parameters in a small number of animals (injection route, dosage, and fasting conditions). Thus, such an imaging protocol combines the strengths of both dynamic planar and tomographic imaging, and by using iron-based NPs of high biocompatibility along with the appropriate administration route, a potential diagnostic or therapeutic effect could be attained.
Assuntos
Nanopartículas , Animais , Nanopartículas Magnéticas de Óxido de Ferro , Camundongos , Tomografia Computadorizada de Emissão de Fóton Único , Tomografia Computadorizada por Raios X , Fluxo de TrabalhoRESUMO
Magnetic resonance imaging (MRI) has a leading place in medicine as an imaging tool of high resolution for anatomical studies and diagnosis of diseases, in particular for soft tissues that cannot be accessible by other modalities. Many research works are thus focused on improving the images obtained with MRI. This technique has indeed poor sensitivity, which can be compensated by using a contrast agent (CA). Today, the clinically approved CAs on market are solely based on gadolinium complexes that may induce nephrogenic systemic fibrosis for patients with kidney failure, whereas more recent studies on healthy rats also showed Gd retention in the brain. Consequently, researchers try to elaborate other types of safer MRI CAs like manganese-based complexes. In this context, the synthesis of Mn2+ complexes of four 12-membered pyridine-containing macrocyclic ligands based on the pyclen core was accomplished and described herein. Then, the properties of these Mn(II) complexes were studied by two relaxometric methods, 17O NMR spectroscopy and 1H NMR dispersion profiles. The time of residence (τM) and the number of water molecules (q) present in the inner sphere of coordination were determined by these two experiments. The efficacy of the pyclen-based Mn(II) complexes as MRI CAs was evaluated by proton relaxometry at a magnetic field intensity of 1.41 T near those of most medical MRI scanners (1.5 T). Both the 17O NMR and the nuclear magnetic relaxation dispersion profiles indicated that the four hexadentate ligands prepared herein left one vacant coordination site to accommodate one water molecule, rapidly exchanging, in around 6 ns. Furthermore, it has been shown that the presence of an additional amide bond formed when the paramagnetic complex is conjugated to a molecule of interest does not alter the inner sphere of coordination of Mn, which remains monohydrated. These complexes exhibit r1 relaxivities, large enough to be used as clinical MRI CAs (1.7-3.4 mM-1·s-1, at 1.41 T and 37 °C).
Assuntos
Compostos Azabicíclicos/química , Meios de Contraste/química , Complexos de Coordenação/química , Compostos Azabicíclicos/síntese química , Meios de Contraste/síntese química , Complexos de Coordenação/síntese química , Ligantes , Imageamento por Ressonância Magnética , Manganês/química , Isótopos de Oxigênio/química , Estudo de Prova de Conceito , Espectroscopia de Prótons por Ressonância Magnética , Água/químicaRESUMO
The inherent lack of sensitivity of MRI needs the development of new Gd contrast agents in order to extend the application of this technique to cellular imaging. For this purpose, two multimeric MR contrast agents obtained by peptidic coupling between an amido amine dendron and GdDOTAGA chelates (premetalation strategy, G1-4GdDOTAGA) or DO3A derivatives which then were postmetalated (G1-4GdDO3A) have been prepared. By comparison to the monomers, an increase of longitudinal relaxivity has been observed for both structures. Especially for G1-4GdDO3A, a marked increase is observed between 20 and 60 MHz. This structure differs from G1-4GdDOTAGA by an increased rigidity due to the aromatic linker between each chelate and the organic framework. This has the effect of limiting local rotational movements, which has a positive impact on relaxivity.
RESUMO
The synthesis of poly[N,N-bis(3-aminopropyl)glycine] (PAPGly) dendrons Gd-based contrast agents (GdCAs) via an orthogonal protection of the different functional groups and an activation/coupling strategy wherein a specific number of synthetic steps add a generation to the existing dendron has been described. The aim of this protocol is to build up two different generations of dendrons (G-0 or dendron's core, and G-1) with peripheral NH2 groups to conjugate a 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (DO3A) derivative and afterwards to chelate with Gd3+ paramagnetic ions. These complexes, which have a well-defined molecular weight, are of relevance to MRI as an attempt to gain higher 1 H relaxivity by slowing down the rotation of molecule compared to monomeric Gd(III) complexes used as contrast agents and to increase the number of paramagnetic centers present in one molecular structure. From the study of their water 1 H longitudinal relaxation rate at different magnetic fields (NMRD, Nuclear Magnetic Relaxation Dispersion) and by evaluating the variable temperature 17 O-NMR data we determined the parameters characterizing the water exchange rate and the rotational correlation time of each complex, both affecting 1 H relaxivity. Furthermore, these two novel PAPGly GdCAs were objects of i) an inâ vivo study to determine their biodistributions in healthy C57 mice at several time points, and ii) the Dynamic Contrast-Enhanced MRI (DCE-MRI) approach to assess their contrast efficiency measured in the tumor region of C57BL/6 mice transplanted subcutaneously with B16-F10 melanoma cells. The aim of the comparison of these two dendrons GdCAs, having different molecular weights (MW), is to understand how MW and relaxivity may influence the contrast enhancement capabilities inâ vivo at low magnetic field (1â T). Significant contrast enhancement was observed in several organs (vessel, spleen and liver), already at 5â min post-injection, for the investigated CAs. Moreover, these CAs induced a marked contrast enhancement in the tumor region, thanks to the enhanced permeability retention effect of those macromolecular structures.
Assuntos
Meios de Contraste/química , Gadolínio/química , Melanoma/química , Compostos Organometálicos/química , Animais , Meios de Contraste/síntese química , Meios de Contraste/farmacocinética , Gadolínio/farmacocinética , Humanos , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Masculino , Melanoma/diagnóstico por imagem , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Neoplasias Experimentais/química , Neoplasias Experimentais/diagnóstico por imagem , Compostos Organometálicos/síntese química , Compostos Organometálicos/farmacocinética , Distribuição TecidualRESUMO
The growing concern over the toxicity of Gd-based contrast agents used in magnetic resonance imaging (MRI) motivates the search for less toxic and more effective alternatives. Among these alternatives, iron-iron oxide (Fe@FeOx) core-shell architectures have been long recognized as promising MRI contrast agents while limited information on their engineering is available. Here we report the synthesis of 10 nm large Fe@FeOx nanoparticles, their coating with a 11 nm thick layer of dense silica and functionalization by 5 kDa PEG chains to improve their biocompatibility. The nanomaterials obtained have been characterized by a set of complementary techniques such as infra-red and nuclear magnetic resonance spectroscopies, transmission electron microscopy, dynamic light scattering and zetametry, and magnetometry. They display hydrodynamic diameters in the 100 nm range, zetapotential values around -30 mV, and magnetization values higher than the reference contrast agent RESOVIST®. They display no cytotoxicity against 1BR3G and HCT116 cell lines and no hemolytic activity against human red blood cells. Their nuclear magnetic relaxation dispersion (NMRD) profiles are typical for nanomaterials of this size and magnetization. They display high r2 relaxivity values and low r1 leading to enhanced r2/r1 ratios in comparison with RESOVIST®. All these data make them promising contrast agents to detect early stage tumors.
Assuntos
Dextranos/química , Compostos Férricos/química , Ferro/química , Imageamento por Ressonância Magnética , Nanopartículas de Magnetita/química , Dióxido de Silício , Linhagem Celular Tumoral , Materiais Revestidos Biocompatíveis , Humanos , Nanopartículas de Magnetita/ultraestrutura , Modelos Teóricos , Dióxido de Silício/química , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
INTRODUCTION: Human spontaneous osteonecrosis of the knee (SPONK) is still challenging as the current treatments do not allow the production of hyaline cartilage tissue. The aim of the present study was to explore the therapeutic potential of cartilage regeneration using a new biphasic scaffold (type I collagen/hydroxyapatite) previously loaded or not with concentrated bone marrow cells. MATERIAL AND METHODS: Female rabbits were operated of one knee to create articular lesions of the trochlea (three holes of 4 × 4mm). The holes were left empty in the control group or were filled with the scaffold alone or the scaffold previously loaded with concentrated bone marrow cells. After two months, rabbits were sacrificed and the structure of the newly formed tissues were evaluated by macroscopic, MRI, and immunohistochemistry analyses. RESULTS: Macroscopic and MRI evaluation of the knees did not show differences between the three groups (p > 0.05). However, histological analysis demonstrated that a higher O'Driscoll score was obtained in the two groups treated with the scaffold, as compared to the control group (p < 0.05). The number of cells in treated area was higher in scaffold groups compared to the control group (p < 0.05). There was no difference for intensity of collagen type II between the groups (p > 0.05) but subchondral bone repair was significantly thicker in scaffold-treated groups than in the control group (1 mm for the control group vs 2.1 and 2.6 mm for scaffold groups). Furthermore, we observed that scaffolds previously loaded with concentrated bone marrow were more reabsorbed (p < 0.05). CONCLUSION: The use of a biphasic scaffold previously loaded with concentrated bone marrow significantly improves cartilage lesion healing.
Assuntos
Cartilagem Articular/cirurgia , Articulação do Joelho/fisiopatologia , Transplante de Células-Tronco Mesenquimais/métodos , Alicerces Teciduais , Animais , Regeneração Óssea/fisiologia , Cartilagem Articular/metabolismo , Cartilagem Articular/fisiopatologia , Colágeno Tipo I/farmacologia , Colágeno Tipo II/metabolismo , Durapatita/farmacologia , Feminino , Imuno-Histoquímica , Articulação do Joelho/metabolismo , Articulação do Joelho/cirurgia , Imageamento por Ressonância Magnética , CoelhosRESUMO
A new facile synthetic strategy was developed to prepare bifunctional monophosphinic acid Ln-DOTA derivatives, Gd-DO2AGAPNBn and Gd- DO2AGAPABn. The relaxivities of the Gd-complexes are enhanced compared to Gd-DOTA. Monophosphinic acid arm of these Gd-complexes affords enhancement of inner sphere water exchange rate due to its steric bulkiness. The different functionalities of DO2AGAPNBn were appended in trans positions and are designed to conjugate identical or different vectors according to the potential applications. The conjugation of Gd-DO2AGAPABn with E3 peptide known to target apoptosis was successfully performed and in vivo MRI allowed cell death detection in a mouse model.
Assuntos
Meios de Contraste/química , Compostos Heterocíclicos com 1 Anel/química , Imagem Multimodal , Animais , Meios de Contraste/síntese química , Relação Dose-Resposta a Droga , Feminino , Compostos Heterocíclicos com 1 Anel/síntese química , Imageamento por Ressonância Magnética , Camundongos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
It is now widely recognized that the sampling rate of Polar Organic Chemical Integrative Samplers (POCIS) is significantly affected by flow velocity, which can cause a consequent bias when determining time-weighted average concentrations (TWAC). We already observed the desorption of deisopropylatrazine (DIA) over time when added to the receiving phase of a POCIS. This desorption rate was particularly influenced by flow velocity, in an agitated water environment in situ. In the method presented here, we calibrated 30 pesticides under controlled laboratory conditions, varying the flow velocity over four levels. We simultaneously studied the desorption rate of DIA-d5 (a deuterated form of DIA) over time. An empirical model based on a power law involving flow velocity was used to process the information from the accumulation kinetics of the compounds of interest and elimination of DIA-d5. This type of model makes it possible to consider the effect of this crucial factor on exchange kinetics, and then to obtain more accurate TWACs with reduced bias and more acceptable dispersion of results.
Assuntos
Praguicidas , Poluentes Químicos da Água , Poluentes Químicos da Água/análise , Monitoramento Ambiental/métodos , Praguicidas/análise , Compostos Orgânicos/química , CalibragemRESUMO
This paper presents a multi-step methodology to identify relationships between integrative pesticide quantifications and land uses on a given watershed of the Adour-Garonne Basin (Southwestern France). In fact, a large amount of pesticide concentration data was collected from 51 sites located in the Adour-Garonne Basin for a 1 year monitoring period in 2016. The sampling devices used here were polar organic chemical integrative samplers (POCIS), which provided time-weighted average concentration estimates. For each study site, its associated watershed and land cover distribution were determined using Corine Land Cover 2012 (CLC 2012) and Geographic Information System (GIS). The large-scale data were analyzed using multivariate statistical analyses, such as hierarchical cluster analysis (HCA) and principal component analysis (PCA). HCA grouped the 51 sites into five clusters with similar primary land uses. Next, the integrated pesticide concentration and land use distribution data sets were analyzed in a PCA. The key variables responsible for discriminating the sample sites showed distribution patterns consistent with specific land uses. To confirm these observations, pesticide fingerprints from sites with contrasting land uses were compared using a waffle method. The overall multivariate approach allowed for the identification of contamination sources related to their likely initial use, at the watershed level, that could be useful for preventing or containing pesticide pollution beyond simply acting on areas at risk.
RESUMO
In this lab-scale study, the POCIS capacity to integrate short contamination peaks of variable intensity and duration was evaluated. POCIS were immersed for 14 days in tanks filled with tap water and spiked at different concentrations with 12 pesticides of various polarities (log Kow = 1.1-4.7) and classes (herbicides, fungicides, and insecticides). Concentrations were kept relatively constant at 1 µg L-1 and 5 µg L-1, respectively, in two "background" exposure tanks. Three contamination peaks of increasing intensity and decreasing duration were simulated (10 µg L-1 for 24 h, 40 µg L-1 for 6 h, and 60 µg L-1 for 1 h). This lab-scale study demonstrated that ten moderately polar compounds (2 < log Kow < 4) showed a linear uptake, as observed in previous studies, while a non-linear model fits the data of the two most polar pesticides (log Kow < 2). Depending on chemical polarity, some compounds exhibited a "burst effect" or "lag effect" during the first 3 days of exposure. After 14 days of exposure, contamination peaks appeared integrated for seven compounds, showing the ability of POCIS to catch very short pollution events and to provide acceptable time-weighted average concentration estimates under laboratory-controlled conditions.
Assuntos
Praguicidas , Poluentes Químicos da Água , Monitoramento Ambiental , Laboratórios , Compostos Orgânicos , Praguicidas/análise , Poluentes Químicos da Água/análiseRESUMO
Alizarin detection in fish fins is extensively employed because it is easy to use. However, in eels, the eelGFP fluorescent protein may impede the detection of the fluorescent markers in the eel tissues. The study tests the effectiveness of three of the most up-to-date alizarin-detecting technologies on the living body and fins of European glass eels (Anguilla anguilla L.). The findings demonstrated that the control group had a high autofluorescence at alizarin and eelGFP maxima bands. With fluorescence reflectance imaging (FRI), the eel living body autofluorescence impeded the detection of the marked eels. In contrast with experimental excitation-emission-matrix (EEM) fluorescence analyses, 99% of the marked eels were correctly assigned to their group from fluorescence analyses of their fin cellular contents. With epifluorometry (EPI), 100% of the marked eels were detected with the caudal fin tips when excited at 450-490 nm wavelengths due to a weaker autofluorescence signal. EEM and FRI assays unveiled an average fluorescence quenching 60% and 44% of the marked group respectively, in the alizarin and eelGFP maxima bands. The fluorescence quenching observed is discussed. Results will benefit experimental design by examining autofluorescence effects on mark detection and the development of non-invasive detection methods in this critically endangered species.
Assuntos
Anguilla , Anguilla/metabolismo , Animais , Antraquinonas/metabolismo , Corantes/metabolismoRESUMO
The blood-brain barrier (BBB) protects the central nervous system (CNS) from harmful blood-borne factors. Although BBB dysfunction is a hallmark of several neurological disorders, therapies to restore BBB function are lacking. An attractive strategy is to repurpose developmental BBB regulators, such as Wnt7a, into BBB-protective agents. However, safe therapeutic use of Wnt ligands is complicated by their pleiotropic Frizzled signaling activities. Taking advantage of the Wnt7a/b-specific Gpr124/Reck co-receptor complex, we genetically engineered Wnt7a ligands into BBB-specific Wnt activators. In a "hit-and-run" adeno-associated virus-assisted CNS gene delivery setting, these new Gpr124/Reck-specific agonists protected BBB function, thereby mitigating glioblastoma expansion and ischemic stroke infarction. This work reveals that the signaling specificity of Wnt ligands is adjustable and defines a modality to treat CNS disorders by normalizing the BBB.
Assuntos
Barreira Hematoencefálica/fisiologia , Proteínas Ligadas por GPI/agonistas , Glioblastoma/terapia , Receptores Acoplados a Proteínas G/agonistas , Acidente Vascular Cerebral/terapia , Proteínas Wnt/genética , Via de Sinalização Wnt , Animais , Encéfalo/metabolismo , Células Endoteliais/metabolismo , Receptores Frizzled/metabolismo , Glioblastoma/metabolismo , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Mutagênese , Sistema Nervoso/embriologia , Engenharia de Proteínas , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Acidente Vascular Cerebral/metabolismo , Proteínas Wnt/química , Proteínas Wnt/metabolismo , Xenopus laevis , Peixe-ZebraRESUMO
The preparation of an efficient bimodal single probe for magnetic resonance (MRI) and optical imaging (OI) is reported. Paramagnetic properties have been obtained by the non-covalent encapsulation of the clinically used Gd3+ chelate (i.e., Gd-HP-DO3A) within silica nanoparticles through a water-in-oil microemulsion process. To ensure colloidal stability, the surface of the particles was modified by means of treatment using PEG-silane, and further functionalized photochemically using a diazirine linker bearing carboxylic functions. Optical properties were obtained by the covalent grafting of a near-infrared emitting probe (NIR) on the resulting surface. The confinement of Gd complexes within the permeable matrix resulted in a significant increase in longitudinal relaxivities (>500% at 20 MHz) in comparison with the relaxivities of free chelate, while the post-functionalization process of PEG with fluorescent compounds appeared promising for the derivatization procedure. Several physico-chemical properties attested to the efficient surface modification and confirmed covalent grafting. Preliminary imaging experiments complete this study and confirm the potential of the presented system for preclinical imaging experiments.
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Meios de Contraste , Dióxido de Silício , Imageamento por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Imagem ÓpticaRESUMO
Bimodal sub-5 nm superparamagnetic iron oxide nanoparticles (SPIO-5) coated with polyethylene glycol of different chain lengths (i.e. PEG-800, -2000 and -5000) have been prepared and characterized. Fluorescence properties have been obtained by mean of the grafting of a near-infrared-emitting dye (NIR-dye) onto the surface of the oxide, thanks to the carboxylic acid functions introduced towards an organosilane coating. Such modification allowed us to follow in vivo their biodistribution and elimination pathways by T1-w and T2-w high-field magnetic resonance imaging (MRI), as well as by optical and optoacoustic imaging. Interestingly, it has been highlighted that for a given composition, the thickness of the coating strongly influences the pharmacokinetic properties of the administrated SPIO-5.
Assuntos
Nanopartículas Magnéticas de Óxido de Ferro/química , Polietilenoglicóis/química , Animais , Feminino , Camundongos , Camundongos Pelados , Estrutura Molecular , Imagem Óptica , Distribuição TecidualRESUMO
This work describes the preparation, characterization and functionalization with magnetic nanoparticles of a bone tissue-mimetic scaffold composed of collagen and hydroxyapatite obtained through a biomineralization process. Bone remodeling takes place over several weeks and the possibility to follow it in vivo in a quick and reliable way is still an outstanding issue. Therefore, this work aims to produce an implantable material that can be followed in vivo during bone regeneration by using the existing non-invasive imaging techniques (MRI). To this aim, suitably designed biocompatible SPIONs were linked to the hybrid scaffold using two different strategies, one involving naked SPIONs (nMNPs) and the other using coated and activated SPIONs (MNPs) exposing carboxylic acid functions allowing a covalent attachment between MNPs and collagen molecules. Physico-chemical characterization was carried out to investigate the morphology, crystallinity and stability of the functionalized materials followed by MRI analyses and evaluation of a radiotracer uptake ([99mTc]Tc-MDP). Cell proliferation assays in vitro were carried out to check the cytotoxicity and demonstrated no side effects due to the SPIONs. The achieved results demonstrated that the naked and coated SPIONs are more homogeneously distributed in the scaffold when incorporated during the synthesis process. This work demonstrated a suitable approach to develop a biomaterial for bone regeneration that allows the monitoring of the healing progress even for long-term follow-up studies.
Assuntos
Regeneração Óssea , Alicerces Teciduais , Osso e Ossos/diagnóstico por imagem , Colágeno , DurapatitaRESUMO
BACKGROUND: The link between early blood- brain barrier (BBB) breakdown and endothelial cell activation in acute stroke remain poorly defined. We hypothesized that P-selectin, a mediator of the early phase of leukocyte recruitment in acute ischemia is also a major contributor to early BBB dysfunction following stroke. This was investigated by examining the relationship between BBB alterations following transient ischemic stroke and expression of cellular adhesion molecule P-selectin using a combination of magnetic resonance molecular imaging (MRMI), intravital microscopy and immunohistochemistry. MRMI was performed using the contrast, gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) conjugated to Sialyl Lewis X (Slex) where the latter is known to bind to activated endothelium via E- or P selectins. Middle cerebral artery occlusion was induced in male C57/BL 6 wild-type (WT) mice and P-selectin-knockout (KO) mice. At 24 hours following middle cerebral artery occlusion, T1 maps were acquired prior to and following contrast injection. In addition to measuring P- and E-selectin expression in brain homogenates, alterations in BBB function were determined immunohistochemically by assessing the extravasation of immunoglobulin G (IgG) or staining for polymorphonuclear (PMN) leukocytes. In vivo assessment of BBB dysfunction was also investigated optically using intravital microscopy of the pial circulation following the injection of Fluorescein Isothiocyanate (FITC)-dextran (MW 2000 kDa). RESULTS: MRI confirmed similar infarct sizes and T1 values at 24 hours following stroke for both WT and KO animals. However, the blood to brain transfer constant for Gd DTPA (Kgd) demonstrated greater tissue extravasation of Gd DTPA in WT animals than KO mice (P < 0.03). In the P selectin KO mice, Delta T1 stroke -Delta T1 contralateral control cortex, decreased significantly in the Gd-DTPA(sLeX) group compared to Gd-DTPA, indicative of sLeX mediated accumulation of the targeted contrast agent. Regarding BBB function, in the P-selectin KO mice compared to WT control mice, there was an attenuation in the extravasation of IgG (P < 0.001), a trend for decreased FITC extravasation and less infiltration of PMN leukocytes (P < 0.001) thereby supporting the observed increase in Kgd permeability in stroke brain of WT compared to KO mice. CONCLUSION: P-selectin expression contributes to enhanced BBB dysfunction at 24 hours after transient focal cerebral ischemia.
Assuntos
Barreira Hematoencefálica/fisiopatologia , Infarto da Artéria Cerebral Média/fisiopatologia , Ataque Isquêmico Transitório/fisiopatologia , Selectina-P/metabolismo , Animais , Encéfalo/patologia , Encéfalo/fisiopatologia , Permeabilidade Capilar/fisiologia , Modelos Animais de Doenças , Selectina E/metabolismo , Imunoglobulina G/metabolismo , Infarto da Artéria Cerebral Média/patologia , Ataque Isquêmico Transitório/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/fisiologia , Selectina-P/genética , Pia-Máter/irrigação sanguínea , Pia-Máter/fisiopatologia , Acidente Vascular Cerebral/patologia , Acidente Vascular Cerebral/fisiopatologiaRESUMO
Asiatic citrus canker is a major disease worldwide, and its causal agent, Xanthomonas citri pv. citri, is listed as a quarantine organism in many countries. Analysis of the molecular epidemiology of this bacterium is hindered by a lack of molecular typing techniques suitable for surveillance and outbreak investigation. We report a comparative evaluation of three typing techniques, amplified fragment length polymorphism (AFLP) analysis, insertion sequence ligation-mediated PCR (IS-LM-PCR) typing, and multilocus variable-number tandem-repeat analysis (MLVA), with 234 strains originating from Asia, the likely center of origin of the pathogen, and reference strains of pathotypes A, A*, and A(w), which differ in host range. The typing techniques were congruent in describing the diversity of this strain collection, suggesting that the evolution pattern of the bacterium may be clonal. Based on a hierarchical analysis of molecular variance, the AFLP method best described the genetic variation found among pathotypes whereas MLVA best described the variation found among individual strains from the same countries or groups of neighboring countries. IS-LM-PCR data suggested that the transposition of insertion sequences in the genome of X. citri pv. citri occurs rarely enough not to disturb the phylogenetic signal. This technique may be useful for the global surveillance of non-epidemiologically related strains. Although pathological characteristics of strains could be most often predicted from genotyping data, we report the occurrence in the Indian peninsula of strains genetically related to pathotype A* strains but with a host range similar to that of pathotype A, which makes the classification of this bacterium even more complicated.