195 research outputs found
Die Bedeutung des AdhĂ€sionsmolekĂŒls JAM-A sowie der Chemokinrezeptoren Ccr1, -2 und -5 fĂŒr die Rekrutierung von Leukozyten bei EntzĂŒndung und IschĂ€mie-Reperfusion
Die Eigenschaft von Leukozyten, das GefĂ€Ăsystem zu verlassen und in das umliegende Gewebe auszuwandern, ist von essentieller Bedeutung fĂŒr die BekĂ€mpfung von Infektionen und darĂŒber hinaus entscheidend fĂŒr die Pathogenese des I/R-Schadens.
Die Extravasation von Leukozyten stellt dabei einen kaskadenartig verlaufenden Prozess dar, welcher sich in die Schritte Rolling, AdhĂ€renz, transendotheliale und interstitielle Migration gliedern lĂ€sst. Ein geeignetes Versuchsmodell, welches am M. cremaster der Maus in vivo alle Schritte des leukozytĂ€ren Rekrutierungsprozesses wĂ€hrend I/R zu analysieren erlaubt, liegt bisher nicht vor. WĂ€hrend die frĂŒhen Schritte des leukozytĂ€ren Extravasationsprozesses weitgehend aufgeklĂ€rt sind, sind die Schritte der transendothelialen und interstitiellen Migration von Leukozyten unzureichend verstanden. In vitro Untersuchungen zeigen, dass das MolekĂŒl JAM-A in die Transmigration von Leukozyten involviert ist, jĂŒngste in vivo Studien zeigen jedoch kontroverse Ergebnisse. Ferner gibt es zunehmend Hinweise darauf, dass die Chemokinrezeptoren Ccr1, Ccr2 und Ccr5 an der Extravasation von Leukozyten beteiligt sind. Welche Bedeutung diese Chemokinrezeptoren fĂŒr die einzelnen Schritte des leukozytĂ€ren Rekrutierungsprozesses bei EntzĂŒndung und I/R besitzen, ist bislang unklar.
Die Ziele der vorliegenden Arbeit waren daher i) ein geeignetes Modell zur Untersuchung aller Schritte des leukozytĂ€ren Rekrutierungsprozesses bei I/R am M. cremaster der Maus zu entwickeln, ii) die Bedeutung des AdhĂ€sionsmolekĂŒls JAM-A fĂŒr die Transmigration von Leukozyten zu untersuchen und iii) die Rolle der Chemokinrezeptoren Ccr1, Ccr2 und Ccr5 fĂŒr die einzelnen Schritte des leukozytĂ€ren Rekrutierungsprozesses bei EntzĂŒndung und I/R zu analysieren.
In unterschiedlichen VersuchsansĂ€tzen wurde mit Hilfe der RLOT-Intravitalmikroskopie am M. cremaster anĂ€sthesierter MĂ€use die leukozytĂ€ren Migrationsparameter untersucht. Zur Bestimmung des PhĂ€notyps transmigrierter Leukozyten wurden immunhistochemische FĂ€rbungen von Paraffinschnitten durchgefĂŒhrt.
In einer ersten Versuchsreihe wurden die einzelnen Schritte des leukozytĂ€ren Extravasations-prozesses systematisch in AbhĂ€ngigkeit von IschĂ€miedauer und Reperfusionszeit untersucht. Die Ergebnisse zeigen, dass es bereits nach 30 min IschĂ€mie und 120 min Reperfusion gegenĂŒber schein-operierten Kontrolltieren zu einem starken Anstieg von Leukozyten-adhĂ€renz und -transmigration kommt. Eine VerlĂ€ngerung der IschĂ€miezeit auf 60 bzw. 90 min konnte keine Steigerung der Effekte erzielen. Diese Befunde waren der Ausgangspunkt fĂŒr weitergehende Untersuchungen, welche die Mechanismen des leukozytĂ€ren Rekrutierungs-prozesses nĂ€her charakterisieren sollen.
In diesem Zusammenhang wurde in einer zweiten Versuchsreihe unter Verwendung von JAM-A-defizienten MĂ€usen die Bedeutung des AdhĂ€sionsmolekĂŒls JAM-A fĂŒr die Leukozytenrekrutierung systematisch unter verschiedenen inflammatorischen Bedingungen analysiert. Unsere Daten belegen, dass die transendotheliale Migration von neutrophilen Granulozyten und Monozyten einer Stimulus-spezifischen Regulation durch JAM-A unterliegt. Ferner lassen die Ergebnisse unserer Untersuchungen in eJAM-A-defizienten Tieren darauf schlieĂen, dass endotheliales JAM-A die Transmigration von neutrophilen Granulozyten und Monozyten zwar in der Initialphase entzĂŒndlicher Prozesse vermittelt, zu spĂ€teren Zeitpunkten jedoch keine Bedeutung mehr zu besitzen scheint. SchlieĂlich deuten unsere Befunde darauf hin, dass leukozytĂ€res JAM-A an den der interstitiellen Leukozytenmigration zugrunde liegenden Mechanismen beteiligt ist.
In einer letzten Versuchsreihe wurde die Rolle der Chemokinrezeptoren Ccr1, Ccr2 und Ccr5 fĂŒr die Rekrutierung von Leukozyten bei Chemokin-induzierter EntzĂŒndung und I/R untersucht. Es konnte gezeigt werden, dass diese Chemokinrezeptoren die Extravasation von neutrophilen Granulozyten und Monozyten bei Chemokin-induzierter EntzĂŒndung durch Effekte auf AdhĂ€renz und (konsekutive) transendotheliale Migration mediieren und keinen Einfluss auf das interstitielle Migrationsverhalten transmigrierter Leukozyten besitzen. Des Weiteren ist es mittels durchflusszytometrischer Analyse gelungen, die Expression von Ccr2 und Ccr5 auf nativen neutrophilen Granulozyten nachzuweisen. DarĂŒber hinaus konnte erstmals gezeigt werden, dass die Chemokinrezeptoren Ccr1, Ccr2 und Ccr5 zur Rekrutierung von neutrophilen Granulozyten und Monozyten in das postischĂ€mische Gewebe durch dynamische bzw. differentielle Regulation von AdhĂ€renz und (konsekutiver) Transmigration beitragen
Ccl2 and Ccl3 Mediate Neutrophil Recruitment via Induction of Protein Synthesis and Generation of Lipid Mediators
Objective: Although the chemokines monocyte chemoattractant protein-1 (Ccl2/JE/MCP-1) and macrophage inflammatory protein-1α (Ccl3/MIP-1α) have recently been implicated in neutrophil migration, the underlying mechanisms remain largely unclear.
Methods and Results: Stimulation of the mouse cremaster muscle with Ccl2/JE/MCP-1 or Ccl3/MIP-1α induced a significant increase in numbers of firmly adherent and transmigrated leukocytes (>70% neutrophils) as observed by in vivo microscopy. This increase was significantly attenuated in mice receiving an inhibitor of RNA transcription (actinomycin D) or antagonists of platelet activating factor (PAF; BN 52021) and leukotrienes (MK-886; AA-861). In contrast, leukocyte responses elicited by PAF and leukotriene-B4 (LTB4) themselves were not affected by actinomycin D, BN 52021, MK-886, or AA-861. Conversely, PAF and LTB4, but not Ccl2/JE/MCP-1 and Ccl3/MIP-1α, directly activated neutrophils as indicated by shedding of CD62L and marked upregulation of CD11b. Moreover, Ccl2/JE/
MCP-1- and Ccl3/MIP-1α-elicited leakage of fluorescein isothiocyanate dextran as well as collagen IV remodeling within the venular basement membrane were completely absent in neutrophil-depleted mice.
Conclusions: Ccl2/JE/MCP-1 and Ccl3/MIP-1α mediate firm adherence and (subsequent) transmigration of neutrophils via protein synthesis and secondary generation of leukotrienes and PAF, which in turn directly activate neutrophils. Thereby, neutrophils facilitate basement membrane remodeling and promote microvascular leakage
Inhibitor of Apoptosis Proteins as Novel Targets in Inflammatory Processes
Objective: Inhibitor of apoptosis proteins (IAPs), such as X-linked or cellular IAP 1/2 (XIAP, cIAP1/2), are important regulators of apoptosis. IAP antagonists are currently under clinical investigation as anticancer agents. Interestingly, IAPs participate in the inflammation-associated TNF receptor signaling complex and regulate NFÎșB signaling. This raises the question about the role of IAPs in inflammation. Here, we investigated the anti-inflammatory potential of IAP inhibitors and the role of IAPs in inflammatory processes of endothelial cells.
Methods and Results: In mice, the small molecule IAP antagonist A-4.10099.1 (ABT) suppressed antigen-induced arthritis, leukocyte infiltration in concanavalin A-evoked liver injury, and leukocyte transmigration in the TNFα-activated cremaster muscle. In vitro, we observed an attenuation of leukocyteâ endothelial cell interaction by downregulation of the intercellular adhesion molecule-1. ABT did not impair NFÎșB signaling but decreased the TNFα-induced activation of the TGF-ÎČâactivated kinase 1, p38, and c-Jun N-terminal kinase. These effects are based on the proteasomal degradation of cIAP1/2 accompanied by an altered ratio of the levels of membrane-localized TNF receptor-associated factors 2 and 5.
Conclusion: Our results reveal IAP antagonism as a profound anti-inflammatory principle in vivo and highlight IAPs as important regulators of inflammatory processes in endothelial cells
KĂŒnstliche Intelligenz in der Hals-Nasen-Ohren-Heilkunde
BACKGROUND The continued advancement of digitalization increasingly allows deployment of artificial intelligence (AI) algorithms, leveraging profound effects on society and medicine. OBJECTIVE This article aims to provide an overview of current developments and futures perspectives of AI in otorhinolaryngology. MATERIALS AND METHODS Scientific studies and expert analyses were evaluated and discussed. RESULTS AI can increase the value of current diagnostic tools in otorhinolaryngology and enhance surgical precision in head and neck surgery. CONCLUSION AI has the potential to further improve diagnostic and therapeutic procedures in otorhinolaryngology. This technology, however, is associated with challenges, for example in the domain of privacy and data security. ZUSAMMENFASSUNG HINTERGRUND: Die fortschreitende Digitalisierung ermöglicht zunehmend den Einsatz von kĂŒnstlicher Intelligenz (KI). Sie wird Gesellschaft und Medizin in den nĂ€chsten Jahren maĂgeblich beeinflussen. ZIEL DER ARBEIT Darstellung des gegenwĂ€rtigen Einsatzspektrums von KI in der Hals-Nasen-Ohren-Heilkunde und Skizzierung zukĂŒnftiger Entwicklungen bei der Anwendung dieser Technologie. MATERIAL UND METHODEN Es erfolgte die Auswertung und Diskussion wissenschaftlicher Studien und Expertenanalysen. ERGEBNISSE Durch die Verwendung von KI kann der Nutzen herkömmlicher diagnostischer Werkzeuge in der Hals-Nasen-Ohren-Heilkunde gesteigert werden. Zudem kann der Einsatz dieser Technologie die chirurgische PrĂ€zision in der Kopf-Hals-Chirurgie weiter erhöhen. SCHLUSSFOLGERUNGEN KI besitzt ein groĂes Potenzial zur weiteren Verbesserung diagnostischer und therapeutischer Verfahren in der Hals-Nasen-Ohren-Heilkunde. Allerdings ist die Anwendung dieser Technologie auch mit Herausforderungen verbunden, beispielsweise im Bereich des Datenschutzes
Arabidopsis thaliana GLYCINE RICH RNAâBINDING PROTEIN 7 interaction with its iCLIP target LHCB1.1 correlates with changes in RNA stability and circadian oscillation
The importance of RNAâbinding proteins (RBPs) for plant responses to environmental stimuli and development is well documented. Insights into the portfolio of RNAs they recognize, however, clearly lack behind the understanding gathered in nonâplant model organisms. Here, we characterize binding of the circadian clockâregulated Arabidopsis thaliana GLYCINEâRICH RNAâBINDING PROTEIN 7 (AtGRP7) to its target transcripts. We identified novel RNA targets from individualânucleotide resolution UV crosslinking and immunoprecipitation (iCLIP) data using an improved bioinformatics pipeline that will be broadly applicable to plant RBP iCLIP data. 2705 transcripts with binding sites were identified in plants expressing AtGRP7âGFP that were not recovered in plants expressing an RNAâbinding dead variant or GFP alone. A conserved RNA motif enriched in uridine residues was identified at the AtGRP7 binding sites. NMR titrations confirmed the preference of AtGRP7 for RNAs with a central Uârich motif. Among the bound RNAs, circadian clockâregulated transcripts were overrepresented. Peak abundance of the LHCB1.1 transcript encoding a chlorophyllâbinding protein was reduced in plants overexpressing AtGRP7 whereas it was elevated in atgrp7 mutants, indicating that LHCB1.1 was regulated by AtGRP7 in a doseâdependent manner. In plants overexpressing AtGRP7, the LHCB1.1 halfâlife was shorter compared to wildâtype plants whereas in atgrp7 mutant plants, the halfâlife was significantly longer. Thus, AtGRP7 modulates circadian oscillations of its in vivo binding target LHCB1.1 by affecting RNA stability.Deutsche Forschungsgemeinschaft
http://dx.doi.org/10.13039/501100001659Peer Reviewe
Optimized dispersion of nanoparticles for biological in vitro and in vivo studies
Background: The aim of this study was to establish and validate a practical method to disperse nanoparticles in physiological solutions for biological in vitro and in vivo studies. Results: TiO(2) (rutile) dispersions were prepared in distilled water, PBS, or RPMI 1640 cell culture medium. Different ultrasound energies, various dispersion stabilizers (human, bovine, and mouse serum albumin, Tween 80, and mouse serum), various concentrations of stabilizers, and different sequences of preparation steps were applied. The size distribution of dispersed nanoparticles was analyzed by dynamic light scattering and zeta potential was measured using phase analysis light scattering. Nanoparticle size was also verified by transmission electron microscopy. A specific ultrasound energy of 4.2 x 10(5) kJ/m(3) was sufficient to disaggregate TiO(2) (rutile) nanoparticles, whereas higher energy input did not further improve size reduction. The optimal sequence was first to sonicate the nanoparticles in water, then to add dispersion stabilizers, and finally to add buffered salt solution to the dispersion. The formation of coarse TiO(2) (rutile) agglomerates in PBS or RPMI was prevented by addition of 1.5 mg/ml of human, bovine or mouse serum albumin, or mouse serum. The required concentration of albumin to stabilize the nanoparticle dispersion depended on the concentration of the nanoparticles in the dispersion. TiO(2) (rutile) particle dispersions at a concentration lower than 0.2 mg/ml could be stabilized by the addition of 1.5 mg/ml albumin. TiO(2) (rutile) particle dispersions prepared by this method were stable for up to at least 1 week. This method was suitable for preparing dispersions without coarse agglomerates (average diameter < 290 nm) from nanosized TiO(2) (rutile), ZnO, Ag, SiO(x), SWNT, MWNT, and diesel SRM2975 particulate matter. Conclusion: The optimized dispersion method presented here appears to be effective and practicable for preparing dispersions of nanoparticles in physiological solutions without creating coarse agglomerates
Components of the Plasminogen Activation System Promote Engraftment of Porous Polyethylene Biomaterial via Common and Distinct Effects
Rapid fibrovascularization is a prerequisite for successful biomaterial engraftment. In addition to their well-known roles in fibrinolysis, urokinase-type plasminogen activator (uPA) and tissue plasminogen activator (tPA) or their inhibitor plasminogen activator inhibitor-1 (PAI-1) have recently been implicated as individual mediators in non-fibrinolytic processes, including cell adhesion, migration, and proliferation. Since these events are critical for fibrovascularization of biomaterial, we hypothesized that the components of the plasminogen activation system contribute to biomaterial engraftment. Employing in vivo and ex vivo microscopy techniques, vessel and collagen network formation within porous polyethylene (PPE) implants engrafted into dorsal skinfold chambers were found to be significantly impaired in uPA-, tPA-, or PAI-1-deficient mice. Consequently, the force required for mechanical disintegration of the implants out of the host tissue was significantly lower in the mutant mice than in wild-type controls. Conversely, surface coating with recombinant uPA, tPA, non-catalytic uPA, or PAI-1, but not with non-catalytic tPA, accelerated implant vascularization in wild-type mice. Thus, uPA, tPA, and PAI-1 contribute to the fibrovascularization of PPE implants through common and distinct effects. As clinical perspective, surface coating with recombinant uPA, tPA, or PAI-1 might provide a novel strategy for accelerating the vascularization of this biomaterial
Symmetric microwave potentials for interferometry with thermal atoms on a chip
International audienceA trapped atom interferometer involving state-selective adiabatic potentials with two microwave frequencies on a chip is proposed. We show that this configuration provides a way to achieve a high degree of symmetry between the two arms of the interferometer, which is necessary for coherent splitting and recombination of thermal (i.e., noncondensed) atoms. The resulting interferometer holds promise to achieve high contrast and long coherence time, while avoiding the mean-field interaction issues of interferometers based on trapped Bose-Einstein condensates
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