Advances in Corrosion Casting Methods

This paper briefly discusses the concept of corrosion cast preparation (primarily of blood vessels), the use of the scanning electron microscope (SEM) to study these casts and the observations which can be made, together with the merits and the limitations in various applications. A number of reviews and surveys are quoted in which the different injection media, injection methods, animal preparations and corrosion procedures are described. A new procedure of cleaning the corrosion casts with sodium hydroxide and Triton X-100 is described. The observations which can be made are listed and illustrated both on the cellular level as well as in organ systems as a whole. The discussion centers around some common misconceptions, the feasibility in various applications and the limitations of the method. The conclusion is that the method has proven to be useful especially in conjunction with other methods. Moreover, while the concept of the method may be very straight-forward the approach and the interpretation often need careful consideration and might not be as straight-forward as one tends to expect from the simple sounding principle

Distamycin A Inhibits HMGA1-Binding to the P-Selectin Promoter and Attenuates Lung and Liver Inflammation during Murine Endotoxemia

Background: The architectural transcription factor High Mobility Group-A1 (HMGA1) binds to the minor groove of AT-rich DNA and forms transcription factor complexes (“enhanceosomes”) that upregulate expression of select genes within the inflammatory cascade during critical illness syndromes such as acute lung injury (ALI). AT-rich regions of DNA surround transcription factor binding sites in genes critical for the inflammatory response. Minor groove binding drugs (MGBs), such as Distamycin A (Dist A), interfere with AT-rich region DNA binding in a sequence and conformation-specific manner, and HMGA1 is one of the few transcription factors whose binding is inhibited by MGBs. Objectives: To determine whether MGBs exert beneficial effects during endotoxemia through attenuating tissue inflammation via interfering with HMGA1-DNA binding and modulating expression of adhesion molecules. Methodology/Principal Findings: Administration of Dist A significantly decreased lung and liver inflammation during murine endotoxemia. In intravital microscopy studies, Dist A attenuated neutrophil-endothelial interactions in vivo following an inflammatory stimulus. Endotoxin induction of P-selectin expression in lung and liver tissue and promoter activity in endothelial cells was significantly reduced by Dist A, while E-selectin induction was not significantly affected. Moreover, Dist A disrupted formation of an inducible complex containing NF-κB that binds an AT-rich region of the P-selectin promoter. Transfection studies demonstrated a critical role for HMGA1 in facilitating cytokine and NF-κB induction of P-selectin promoter activity, and Dist A inhibited binding of HMGA1 to this AT-rich region of the P-selectin promoter in vivo. Conclusions/Significance: We describe a novel targeted approach in modulating lung and liver inflammation in vivo during murine endotoxemia through decreasing binding of HMGA1 to a distinct AT-rich region of the P-selectin promoter. These studies highlight the ability of MGBs to function as molecular tools for dissecting transcriptional mechanisms in vivo and suggest alternative treatment approaches for critical illness

Gènes persistants dans les génomes bactériens

Les gènes persistants sont présentés dans presque tous les génomes à l'intérieur d'un clade. Ils sont donc des gènes très représentés et sous sélection très forte. Les gènes persistants incluent la majorité des gènes essentiels. Ils aussi incluent des gènes dont l'inactivation n'est pas automatiquement létale mais qui permettent de répondre à une grande variété de situations défavorables. Les gènes persistants évoluent lentement et typiquement en parallèle avec la sous-unité 16S de l'ARNr. Ils sont très sensibles à des collisions frontales entre le polymérase de l ARN et le polymérase de l'ADN, et ont par conséquence une préférence forte pour le brin précoce. Ils tendent à utiliser des codons optimaux pour assurer une expression efficace. Pour minimiser leur perte fréquente, les gènes persistants sont groupés dans le chromosome. Ces groupes de gènes ont souvent des fonctions voisines. Nous avons construit un réseau représentant les groupes de gènes essentiels duquel résulte le coeur du protéome bactérien. Ce réseau souligne les influences de l'environnement dans le façonnement de la composition du protéome bactérien. On développe aussi deux exemples à partir de la comparaison de profils évolutifs ainsi qu'une plate-forme informatique pour la génomique comparative.Persistent genes are genes present in almost every organism within a group. They are the genes endeavoured through natural selections and widely spread. Persistent genes include most of experimentally essential genes. Furthermore, it also includes genes without immediate lethal effects but allowing to respond to a variety of unfavourable situations. Persistent genes evolve slowly and usually in parallel with the 16S rDNA. They are very sensitive to the head-on collisions between RNA polymerase and DNA polymerase, and consequently have a strong preference for the leading strand. They tend to use optimal codons to ensure efficient expression. To avoid the frequent gene loss, persistent genes are clustered, and such local clusters are usually groups of genes executing coupled functions. By assembling the persistent genes clusters together, a network representing the core bacterial proteome is built up. This network highlights the influences from environment in shaping bacterial proteome composition. Two examples applying evolutionary profiles comparison in genome annotation are presented, as well as a platform to carry out comparative genomics.EVRY-BU (912282101) / SudocSudocFranceF

L-selectin shedding does not regulate human neutrophil attachment, rolling, or transmigration across human vascular endothelium in vitro

Current models of the multistep adhesion cascade for leukocyte-endothelial interactions predict loss of L-selectin from the leukocyte surface before transendothelial migration. We have tested this hypothesis using in vitro adhesion and transendothelial migration assays and a zinc-dependent metalloproteinase inhibitor, Ro 31-9790 (N-2-((2s)-[(hydroxycarbamoyl)methyl)-4-methylvaleryl]-N-1,3 -dimethyl-L-valinamide), which prevents chemoattractant-induced (e.g., IL-8, FMLP, C5a, platelet-activating factor) L-selectin endoproteolytic cleavage from isolated human neutrophils. Inhibitor and vehicle-treated neutrophils exhibited identical behavior during both adhesive interactions with 4- and 24-h TNF-alpha-activated HUVEC monolayers under flow, (including rate of initial attachment, rolling velocities, stable adhesion, and transmigration) and in static adhesion assays. Flow cytometric analysis of transmigrated neutrophils with mAb to L-selectin revealed that vehicle treated neutrophils had minimal detectable surface L-selectin, whereas inhibitor-treated neutrophils retained comparable levels of L-selectin on their surface as neutrophils maintained at 37 degrees C. In addition, mAb to L-selectin that induce rapid shape change and homotypic adhesion (LAM1-116) did not enhance the rate or extent of neutrophil transmigration under flow or static conditions. Neutrophils preincubated with LAM 1-116 displayed similar behavior to neutrophils preincubated with the control anti-L-selectin mAb, LAM1-101. In summary, these results demonstrate that there is no requirement for L-selectin to be shed from the surface of neutrophils before, or during, their migration across endothelial monolayers, and that prevention of surface L-selectin proteolytic cleavage does not enhance or inhibit neutrophil-endothelial cell adhesive interactions

Neutrophil margination, sequestration, and emigration in the lungs of L-selectin-deficient mice.

These studies tested the hypothesis that L-selectin plays a role in neutrophil traffic in the lungs, particularly in neutrophil margination, sequestration, and emigration, using L-selectin-deficient mice. No defect in neutrophil margination within either capillaries or arterioles and venules was observed in uninflamed lungs of L-selectin-deficient mice. The initial rapid sequestration of neutrophils within the pulmonary capillaries 1 min after intravascular injection of complement fragments was not prevented. In contrast, L-selectin did contribute to the prolonged neutrophil sequestration (> or = 5 min). Interestingly, neutrophil accumulation within noncapillary microvessels required L-selectin at both 1 and 5 min after complement injection. During bacterial pneumonias, L-selectin played a role in neutrophil accumulation within noncapillary microvessels in response to either Escherichia coli or Streptococcus pneumoniae and within capillaries in response to E. coli but not S. pneumoniae. However, L-selectin was not required for emigration of neutrophils or edema in response to either organism. These studies demonstrate a role for L-selectin in the prolonged sequestration of neutrophils in response to intravascular complement fragments, in the intracapillary accumulation of neutrophils during E. coli-induced pneumonia, and in the accumulation of neutrophils within noncapillary microvessels when induced by either intravascular complement fragments o