Cell Type-Specific Roles of NF-κB Linking Inflammation and Thrombosis

The transcription factor NF-κB is a central mediator of inflammation with multiple links to thrombotic processes. In this review, we focus on the role of NF-κB signaling in cell types within the vasculature and the circulation that are involved in thrombo-inflammatory processes. All these cells express NF-κB, which mediates important functions in cellular interactions, cell survival and differentiation, as well as expression of cytokines, chemokines, and coagulation factors. Even platelets, as anucleated cells, contain NF-κB family members and their corresponding signaling molecules, which are involved in platelet activation, as well as secondary feedback circuits. The response of endothelial cells to inflammation and NF-κB activation is characterized by the induction of adhesion molecules promoting binding and transmigration of leukocytes, while simultaneously increasing their thrombogenic potential. Paracrine signaling from endothelial cells activates NF-κB in vascular smooth muscle cells and causes a phenotypic switch to a “synthetic” state associated with a decrease in contractile proteins. Monocytes react to inflammatory situations with enforced expression of tissue factor and after differentiation to macrophages with altered polarization. Neutrophils respond with an extension of their life span—and upon full activation they can expel their DNA thereby forming so-called neutrophil extracellular traps (NETs), which exert antibacterial functions, but also induce a strong coagulatory response. This may cause formation of microthrombi that are important for the immobilization of pathogens, a process designated as immunothrombosis. However, deregulation of the complex cellular links between inflammation and thrombosis by unrestrained NET formation or the loss of the endothelial layer due to mechanical rupture or erosion can result in rapid activation and aggregation of platelets and the manifestation of thrombo-inflammatory diseases. Sepsis is an important example of such a disorder caused by a dysregulated host response to infection finally leading to severe coagulopathies. NF-κB is critically involved in these pathophysiological processes as it induces both inflammatory and thrombotic responses

An APRI+ALBI Based Multivariable Model as Preoperative Predictor for Posthepatectomy Liver Failure.

OBJECTIVE AND BACKGROUND Clinically significant posthepatectomy liver failure (PHLF B+C) remains the main cause of mortality after major hepatic resection. This study aimed to establish an APRI+ALBI, aspartate aminotransferase to platelet ratio (APRI) combined with albumin-bilirubin grade (ALBI), based multivariable model (MVM) to predict PHLF and compare its performance to indocyanine green clearance (ICG-R15 or ICG-PDR) and albumin-ICG evaluation (ALICE). METHODS 12,056 patients from the National Surgical Quality Improvement Program (NSQIP) database were used to generate a MVM to predict PHLF B+C. The model was determined using stepwise backwards elimination. Performance of the model was tested using receiver operating characteristic curve analysis and validated in an international cohort of 2,525 patients. In 620 patients, the APRI+ALBI MVM, trained in the NSQIP cohort, was compared with MVM's based on other liver function tests (ICG clearance, ALICE) by comparing the areas under the curve (AUC). RESULTS A MVM including APRI+ALBI, age, sex, tumor type and extent of resection was found to predict PHLF B+C with an AUC of 0.77, with comparable performance in the validation cohort (AUC 0.74). In direct comparison with other MVM's based on more expensive and time-consuming liver function tests (ICG clearance, ALICE), the APRI+ALBI MVM demonstrated equal predictive potential for PHLF B+C. A smartphone application for calculation of the APRI+ALBI MVM was designed. CONCLUSION Risk assessment via the APRI+ALBI MVM for PHLF B+C increases preoperative predictive accuracy and represents an universally available and cost-effective risk assessment prior to hepatectomy, facilitated by a freely available smartphone app

Camostat Mesylate Versus Lopinavir/Ritonavir in Hospitalized Patients With COVID-19—Results From a Randomized, Controlled, Open Label, Platform Trial (ACOVACT)

Background: To date, no oral antiviral drug has proven to be beneficial in hospitalized patients with COVID-19.Methods: In this randomized, controlled, open-label, platform trial, we randomly assigned patients ≥18 years hospitalized with COVID-19 pneumonia to receive either camostat mesylate (CM) (considered standard-of-care) or lopinavir/ritonavir (LPV/RTV). The primary endpoint was time to sustained clinical improvement (≥48 h) of at least one point on the 7-category WHO scale. Secondary endpoints included length of stay (LOS), need for mechanical ventilation (MV) or death, and 29-day mortality.Results: 201 patients were included in the study (101 CM and 100 LPV/RTV) between 20 April 2020 and 14 May 2021. Mean age was 58.7 years, and 67% were male. The median time from symptom onset to randomization was 7 days (IQR 5–9). Patients in the CM group had a significantly shorter time to sustained clinical improvement (HR = 0.67, 95%-CI 0.49–0.90; 9 vs. 11 days, p = 0.008) and demonstrated less progression to MV or death [6/101 (5.9%) vs. 15/100 (15%), p = 0.036] and a shorter LOS (12 vs. 14 days, p = 0.023). A statistically nonsignificant trend toward a lower 29-day mortality in the CM group than the LPV/RTV group [2/101 (2%) vs. 7/100 (7%), p = 0.089] was observed.Conclusion: In patients hospitalized for COVID-19, the use of CM was associated with shorter time to clinical improvement, reduced need for MV or death, and shorter LOS than the use of LPV/RTV. Furthermore, research is needed to confirm the efficacy of CM in larger placebo-controlled trials.Systematic Review Registration: [https://clinicaltrials.gov/ct2/show/NCT04351724, https://www.clinicaltrialsregister.eu/ctr-search/trial/2020-001302-30/AT], identifier [NCT04351724, EUDRACT-NR: 2020–001302-30]

Effect of ultrapure lipopolysaccharides derived from diverse bacterial species on the modulation of platelet activation

Platelets are small circulating blood cells that play essential roles in the maintenance of haemostasis via blood clotting. However, they also play critical roles in the regulation of innate immune responses. Inflammatory receptors, specifically Toll-like receptor (TLR)-4, have been reported to modify platelet reactivity. A plethora of studies have reported controversial functions of TLR4 in the modulation of platelet function using various chemotypes and preparations of its ligand, lipopolysaccharide (LPS). The method of preparation of LPS may explain these discrepancies however this is not fully understood. Hence, to determine the impact of LPS on platelet activation, we used ultrapure preparations of LPS from Escherichia coli (LPSEC), Salmonella minnesota (LPSSM), and Rhodobacter sphaeroides (LPSRS) and examined their actions under diverse experimental conditions in human platelets. LPSEC did not affect platelet activation markers such as inside-out signalling to integrin IIb3 or P-selectin exposure upon agonist-induced activation in platelet-rich plasma or whole blood whereas LPSSM and LPSRS inhibited platelet activation under specific conditions at supraphysiological concentrations. Overall, our data demonstrate that platelet activation is not largely influenced by any of the ultrapure LPS chemotypes used in this study on their own except under certain conditions

Vermittlung wissenschaftlich-experimenteller Methodik und Förderung wissenschaftlichen Denkens mittels Distanzlehre

Objective: The aim of this project was to convert a traditional face-to-face seminar for the teaching of experimental scientific methodology to remote teaching in a timely manner due to the COVID-19 related restrictions to teaching in presence.Methodology: The main focus of the course was on flow cytometry. Basics were developed in a virtual presence phase. Specific teaching contents were taught by an interactive presentation, which came very close to the user experience of a flow cytometer and interactively illustrated the influence of different experimental conditions on the obtained results.Video sequences of authentic sample acquisitions were integrated into Adobe Captivate®. These "virtual acquisitions" were not distinguishable from the original procedure. For interpretation of the resulting diagrams, interactions were inserted, which allowed direct comparison of the obtained results.Implementation: A presentation with interactive elements and video sequences was created and used for the virtual presence phases. After publishing on a web server in HTML 5, contents were made available to the students for post-processing of learning contents by self-paced learning with full (interactive) functionality.Conclusion: Contributions elaborated by the students during the course demonstrate a learning outcome comparable to that archieved in the last years in presence mode. While implementation of this solution represented a highly time-consuming process, narrative feedback was consistently positive. Due to the short time available for implementation, no systematic evaluation could be conducted, which represents a clear limitation of this work.Zielsetzung: Die Zielsetzung des dargestellten Projekts bestand darin, ein Seminar zur Vermittlung experimentell- wissenschaftlicher Methodik zeitnah auf Distanzlehre umstellen. Anlass dafür war der COVID-19-bedingte Entfall der Präsenzlehre.Methodik: Der inhaltliche Schwerpunkt der Lehrveranstaltung lag in der Durchflusszytometrie. Die Grundlagen dazu wurden in einer virtuellen Präsenzphase erarbeitet. Für die weiteren spezifischen Unterrichtsinhalte wurde eine interaktive Präsentation verwendet, welche dem Bedienerlebnis eines Durchflusszytometers sehr nahe kam und den Einfluss unterschiedlicher experimenteller Bedingungen auf die erhaltenen Ergebnisse interaktiv veranschaulichte.Dazu wurden unter anderem Videosequenzen authentischer Probenaquisitionen in Adobe Captivate eingebunden. Diese "virtuellen Aquisitionen" waren in ihrer optischen Darstellung vom Originalvorgang nicht unterscheidbar. Für die Interpretation der damit erhaltenen Diagramme wurden Interaktionen eingefügt, welche ein vergleichendes Gegenüberstellen der erhaltenen Ergebnisse ermöglichten.Umsetzung: Eine Präsentation mit interaktiven Elementen und Videosequenzen wurde erstellt und in den virtuellen Präsenzphasen verwendet. Nach Veröffentlichung im HTML 5-Format stand sie den Studierenden für eine Nachbearbeitung der Lerninhalte in selbstbestimmter Ablaufsteuerung und voller (interaktiver) Funktionalität über das Internet zur Verfügung.Schlussfolgerung: Die durch die Studierenden erarbeiteten Inhalte lassen ein inhaltliches Verständnis erkennen, welches mit dem der letzten Jahre (in Präsenzmodus) vergleichbar ist. Das erhaltene narrative Feedback war durchwegs positiv, der für die Implementierung dieser Lösung notwendige Arbeitsaufwand war unverhältnismäßig hoch. Aufgrund der Kurzfristigkeit der Umsetzung konnte keine systematische Evaluation durchgeführt werden, was eine klare Limitation dieser Arbeit darstellt

Human Cytomegalovirus (HCMV) induces Human Endogenous Retrovirus (HERV) transcription.

BACKGROUND: Emerging evidence suggests that human cytomegalovirus (HCMV) is highly prevalent in tumours of different origin. This virus is implied to have oncogenic and oncomodulatory functions, through its ability to control host gene expression. Human endogenous retroviruses (HERV) are also frequently active in tumours of different origin, and are supposed to contribute as cofactors to cancer development. Due to the high prevalence of HCMV in several different tumours, and its ability to control host cell gene expression, we sought to define whether HCMV may affect HERV transcription. FINDINGS: Infection of 3 established cancer cell lines, 2 primary glioblastoma cells, endothelial cells from 3 donors and monocytes from 4 donors with HCMV (strains VR 1814 or TB40/F) induced reverse transcriptase (RT) activity in all cells tested, but the response varied between donors. Both, gammaretrovirus-related class I elements HERV-T, HERV-W, HERV-F and ERV-9, and betaretrovirus-related class II elements HML-2 - 4 and HML-7 - 8, as well as spuma-virus related class III elements of the HERV-L group were up-regulated in response to HCMV infection in GliNS1 cells. Up-regulation of HERV activity was more pronounced in cells harbouring active HCMV infection, but was also induced by UV-inactivated virus. The effect was only slightly affected by ganciclovir treatment and was not controlled by the IE72 or IE86 HCMV genes. CONCLUSIONS: Within this brief report we show that HCMV infection induces HERV transcriptional activity in different cell types. &nbsp