Multiple roles of filopodial dynamics in particle capture and phagocytosis and phenotypes of Cdc42 and Myo10 deletion

Macrophage filopodia, finger-like membrane protrusions, were first implicated in phagocytosis more than 100 years ago, but little is still known about the involvement of these actin-dependent structures in particle clearance. Using spinning disk confocal microscopy to image filopodial dynamics in mouse resident Lifeact-EGFP macrophages, we show that filopodia, or filopodia-like structures, support pathogen clearance by multiple means. Filopodia supported the phagocytic uptake of bacterial (Escherichia coli) particles by (i) capturing along the filopodial shaft and surfing toward the cell body, the most common mode of capture; (ii) capturing via the tip followed by retraction; (iii) combinations of surfing and retraction; or (iv) sweeping actions. In addition, filopodia supported the uptake of zymosan (Saccharomyces cerevisiae) particles by (i) providing fixation, (ii) capturing at the tip and filopodia-guided actin anterograde flow with phagocytic cup formation, and (iii) the rapid growth of new protrusions. To explore the role of filopodia-inducing Cdc42, we generated myeloid-restricted Cdc42 knock-out mice. Cdc42-deficient macrophages exhibited rapid phagocytic cup kinetics, but reduced particle clearance, which could be explained by the marked rounded-up morphology of these cells. Macrophages lacking Myo10, thought to act downstream of Cdc42, had normal morphology, motility, and phagocytic cup formation, but displayed markedly reduced filopodia formation. In conclusion, live-cell imaging revealed multiple mechanisms involving macrophage filopodia in particle capture and engulfment. Cdc42 is not critical for filopodia or phagocytic cup formation, but plays a key role in driving macrophage lamellipodial spreading

Didattica e inclusione scolastica - Inklusion im Bildungsbereich

The inequalities in education, which have also been exposed by the pandemic, highlight the need for alternatives and new horizons. The seventh edition of the conference series "Didattica e Inclusione Scolastica – Inclusion in Education", which was organized for the first time under the direction of the Competence Center for Inclusion in Education of the Free University of Bozen-Bolzano, offered a space to deal with the priorities of a just and inclusive education during this time. This volume opens up a dialogue on inclusive didactics, which builds a bridge between german- and Italian-speaking traditions through its multilingual and intercultural orientation. The twelve contributions open up from different perspectives theoretical references, methods and instruments for the development of an inclusive school with a special focus on didactics.Die Ungleichheiten im Bildungsbereich, die auch durch die Pandemie zutage getreten sind, verdeutlichen die Notwendigkeit von Alternativen und neuen Horizonten. Die siebte Ausgabe der Tagungsreihe „Didattica e Inclusione Scolastica – Inklusion im Bildungsbereich“, welche erstmals unter der Leitung des Kompetenzzentrums für Inklusion im Bildungsbereich der Freien Universität Bozen veranstaltet wurde, bot einen Raum zur Auseinandersetzung mit den Prioritäten einer gerechten und inklusiven Bildung in dieser Zeit. Dieser Band eröffnet einen Dialog über inklusive Didaktik, der durch seine mehrsprachige und interkulturelle Ausrichtung eine Brücke zwischen deutsch- und italienischsprachigen Traditionen schlägt. Die zwölf Beiträge erschließen aus unterschiedlichen Blickwinkeln theoretische Bezüge, Methoden und Instrumente für die Entwicklung einer inklusiven Schule mit besonderem Augenmerk auf die Didaktik.Le disuguaglianze in ambito educativo, emerse anche a causa della pandemia, indicano la necessità di alternative e nuovi orizzonti. La settima edizione del Convegno “Didattica e Inclusione Scolastica – Inklusion im Bildungsbereich”, organizzato per la prima volta dal Centro di Competenza per l’Inclusione Scolastica della Libera Università di Bolzano, ha voluto creare uno spazio per mettere a fuoco le priorità per un’educazione equa ed inclusiva in questo tempo. Con questo volume si dà avvio ad un dialogo sulla didattica inclusiva che si fa plurilingue ed interculturale, creando un ponte fra le tradizioni di lingua italiana e tedesca. I dodici contributi presenti delineano, da prospettive diverse, riferimenti teorici, metodologie e strumenti per lo sviluppo della scuola inclusiva, con un’attenzione particolare alla dimensione didattica

Lack of Effective Anti-Apoptotic Activities Restricts Growth of Parachlamydiaceae in Insect Cells

The fundamental role of programmed cell death in host defense is highlighted by the multitude of anti-apoptotic strategies evolved by various microbes, including the well-known obligate intracellular bacterial pathogens Chlamydia trachomatis and Chlamydia (Chlamydophila) pneumoniae. As inhibition of apoptosis is assumed to be essential for a successful infection of humans by these chlamydiae, we analyzed the anti-apoptotic capacity of close relatives that occur as symbionts of amoebae and might represent emerging pathogens. While Simkania negevensis was able to efficiently replicate within insect cells, which served as model for metazoan-derived host cells, the Parachlamydiaceae (Parachlamydia acanthamoebae and Protochlamydia amoebophila) displayed limited intracellular growth, yet these bacteria induced typical features of apoptotic cell death, including formation of apoptotic bodies, nuclear condensation, internucleosomal DNA fragmentation, and effector caspase activity. Induction of apoptosis was dependent on bacterial activity, but not bacterial de novo protein synthesis, and was detectable already at very early stages of infection. Experimental inhibition of host cell death greatly enhanced parachlamydial replication, suggesting that lack of potent anti-apoptotic activities in Parachlamydiaceae may represent an important factor compromising their ability to successfully infect non-protozoan hosts. These findings highlight the importance of the evolution of anti-apoptotic traits for the success of chlamydiae as pathogens of humans and animals

Host cell death during infection with Chlamydia : a double-edged sword

The phylum Chlamydiae constitutes a group of obligate intracellular bacteria that infect a remarkably diverse range of host species. Some representatives are significant pathogens of clinical or veterinary importance. For instance, Chlamydia trachomatis is the leading infectious cause of blindness and the most common bacterial agent of sexually transmitted diseases. Chlamydiae are exceptionally dependent on their eukaryotic host cells as a consequence of their developmental biology. At the same time, host cell death is an integral part of the chlamydial infection cycle. It is therefore not surprising that the bacteria have evolved exquisite and versatile strategies to modulate host cell survival and death programs to their advantage. The recent introduction of tools for genetic modification of Chlamydia spp., in combination with our increasing awareness of the complexity of regulated cell death in eukaryotic cells, and in particular of its connections to cell-intrinsic immunity, has revived the interest in this virulence trait. However, recent advances also challenged long-standing assumptions and highlighted major knowledge gaps. This review summarizes current knowledge in the field and discusses possible directions for future research, which could lead us to a deeper understanding of Chlamydia's virulence strategies and may even inspire novel therapeutic approaches

[Commentary] Chlamydia Anti-apoptosis - A By-product of Metabolic Reprogramming?

Refers to: Munir A. Al-Zeer, Audrey Xavier, Mohammad Abu Lubad, Janine Sigulla, Mirjana Kessler, Robert Hurwitz, Thomas F. Meyer; Chlamydia trachomatis Prevents Apoptosis Via Activation of PDPK1-MYC and Enhanced Mitochondrial Binding of Hexokinase II, EBioMedicine, Volume 23, September 2017, Pages 100-110. DOI: 10.1016/j.ebiom.2017.08.005</p

Chlamydia trachomatis' struggle to keep its host alive

Bacteria of the phylum Chlamydiae infect a diverse range of eukaryotic host species, including vertebrate animals, invertebrates, and even protozoa. Characteristics shared by all Chlamydiae include their obligate intracellular lifestyle and a biphasic developmental cycle. The infectious form, the elementary body (EB), invades a host cell and differentiates into the replicative form, the reticulate body (RB), which proliferates within a membrane-bound compartment, the inclusion. After several rounds of division, RBs retro-differentiate into EBs that are then released to infect neighboring cells. The consequence of this obligatory transition between replicative and infectious forms inside cells is that Chlamydiae absolutely depend on the viability and functionality of their host cell throughout the entire infection cycle. We recently conducted a forward genetic screen in Chlamydia trachomatis, a common sexually transmitted human pathogen, and identified a mutant that caused premature death in the majority of infected host cells. We employed emerging genetic tools in Chlamydia to link this cytotoxicity to the loss of the protein CpoS (Chlamydia promoter of survival) that normally localizes to the membrane of the pathogen-containing vacuole. CpoS-deficient bacteria also induced an exaggerated type-1 interferon response in infected cells, produced reduced numbers of infectious EBs in cell culture, and were cleared faster from the mouse genital tract in a transcervical infection model in vivo. The analysis of this CpoS-deficient mutant yielded unique insights into the nature of cell-autonomous defense responses against Chlamydia and highlighted the importance of Chlamydia-mediated control of host cell fate for the success of the pathogen

Dialectical Behavioral Therapy for Adolescents (DBT-A): a clinical Trial for Patients with suicidal and self-injurious Behavior and Borderline Symptoms with a one-year Follow-up

Abstract Background To date, there are no empirically validated treatments of good quality for adolescents showing suicidality and non-suicidal self-injurious behavior. Risk factors for suicide are impulsive and non-suicidal self-injurious behavior, depression, conduct disorders and child abuse. Behind this background, we tested the main hypothesis of our study; that Dialectical Behavioral Therapy for Adolescents is an effective treatment for these patients. Methods Dialectical Behavioral Therapy (DBT) has been developed by Marsha Linehan - especially for the outpatient treatment of chronically non-suicidal patients diagnosed with borderline personality disorder. The modified version of DBT for Adolescents (DBT-A) from Rathus & Miller has been adapted for a 16-24 week outpatient treatment in the German-speaking area by our group. The efficacy of treatment was measured by a pre-/post- comparison and a one-year follow-up with the aid of standardized instruments (SCL-90-R, CBCL, YSR, ILC, CGI). Results In the pilot study, 12 adolescents were treated. At the beginning of therapy, 83% of patients fulfilled five or more DSM-IV criteria for borderline personality disorder. From the beginning of therapy to one year after its end, the mean value of these diagnostic criteria decreased significantly from 5.8 to 2.75. 75% of patients were kept in therapy. For the behavioral domains according to the SCL-90-R and YSR, we have found effect sizes between 0.54 and 2.14. During treatment, non-suicidal self-injurious behavior reduced significantly. Before the start of therapy, 8 of 12 patients had attempted suicide at least once. There were neither suicidal attempts during treatment with DBT-A nor at the one-year follow-up. Conclusions The promising results suggest that the interventions were well accepted by the patients and their families, and were associated with improvement in multiple domains including suicidality, non-suicidal self-injurious behavior, emotion dysregulation and depression from the beginning of therapy to the one-year follow-up.</p

Chlamydia trachomatis fails to protect its growth niche against pro-apoptotic insults

Chlamydia trachomatis is an obligate intracellular bacterial agent responsible for ocular infections and sexually transmitted diseases. It has been postulated that Chlamydia inhibits apoptosis in host cells to maintain an intact replicative niche until sufficient infectious progeny can be generated. Here we report that, while cells infected with C. trachomatis are protected from apoptosis at early and mid-stages of infection, they remain susceptible to the induction of other cell death modalities. By monitoring the fate of infected cells by time-lapse video microscopy and by analyzing host plasma membrane integrity and the activity of caspases, we determined that C. trachomatis-infected cells exposed to pro-apoptotic stimuli predominately died by a mechanism resembling necrosis. This necrotic death of infected cells occurred with kinetics similar to the induction of apoptosis in uninfected cells, indicating that C. trachomatis fails to considerably prolong the lifespan of its host cell when exposed to pro-apoptotic insults. Inhibitors of bacterial protein synthesis partially blocked necrotic death of infected cells, suggesting that the switch from apoptosis to necrosis relies on an active contribution of the bacteria. Tumor necrosis factor alpha (TNF-α)-mediated induction of necrosis in cells infected with C. trachomatis was not dependent on canonical regulators of necroptosis, such as RIPK1, RIPK3, or MLKL, yet was blocked by inhibition or depletion of CASP8. These results suggest that alternative signaling pathways regulate necrotic death in the context of C. trachomatis infections. Finally, consistent with the inability of C. trachomatis to preserve host cell viability, necrosis resulting from pro-apoptotic conditions significantly impaired production of infectious progeny. Taken together, our findings suggest that Chlamydia's anti-apoptotic activities are not sufficient to protect the pathogen's replicative niche