Correction to: Two years later: Is the SARS-CoV-2 pandemic still having an impact on emergency surgery? An international cross-sectional survey among WSES members

Background: The SARS-CoV-2 pandemic is still ongoing and a major challenge for health care services worldwide. In the first WSES COVID-19 emergency surgery survey, a strong negative impact on emergency surgery (ES) had been described already early in the pandemic situation. However, the knowledge is limited about current effects of the pandemic on patient flow through emergency rooms, daily routine and decision making in ES as well as their changes over time during the last two pandemic years. This second WSES COVID-19 emergency surgery survey investigates the impact of the SARS-CoV-2 pandemic on ES during the course of the pandemic. Methods: A web survey had been distributed to medical specialists in ES during a four-week period from January 2022, investigating the impact of the pandemic on patients and septic diseases both requiring ES, structural problems due to the pandemic and time-to-intervention in ES routine. Results: 367 collaborators from 59 countries responded to the survey. The majority indicated that the pandemic still significantly impacts on treatment and outcome of surgical emergency patients (83.1% and 78.5%, respectively). As reasons, the collaborators reported decreased case load in ES (44.7%), but patients presenting with more prolonged and severe diseases, especially concerning perforated appendicitis (62.1%) and diverticulitis (57.5%). Otherwise, approximately 50% of the participants still observe a delay in time-to-intervention in ES compared with the situation before the pandemic. Relevant causes leading to enlarged time-to-intervention in ES during the pandemic are persistent problems with in-hospital logistics, lacks in medical staff as well as operating room and intensive care capacities during the pandemic. This leads not only to the need for triage or transferring of ES patients to other hospitals, reported by 64.0% and 48.8% of the collaborators, respectively, but also to paradigm shifts in treatment modalities to non-operative approaches reported by 67.3% of the participants, especially in uncomplicated appendicitis, cholecystitis and multiple-recurrent diverticulitis. Conclusions: The SARS-CoV-2 pandemic still significantly impacts on care and outcome of patients in ES. Well-known problems with in-hospital logistics are not sufficiently resolved by now; however, medical staff shortages and reduced capacities have been dramatically aggravated over last two pandemic years

Human demineralized bone matrix (DBM) in Tissue Engineering of human cartilage

Traumatische, tumoröse oder kongenitale Läsionen im Bereich des Kopfes erfordern eine Rekonstruktion nicht nur aus funktionellen, sondern auch aus ästhetischen Gesichtspunkten. Das Tissue Engineering ist eine wertvolle Methode, um aus wenigen autologen Zellen mit Hilfe eines Biomaterials größere Gewebestrukturen herzustellen. Die vorliegende Arbeit beschäftigt sich deshalb mit dem viel versprechenden Biomaterial der humanen demineralisierten Knochenmatrix (DBM), die bereits erfolgreich seit Jahren zur Reparatur von Knochendefekten in der Unfall- und Kieferchirurgie, aber auch Hals-Nasen- Ohrenheilkunde eingesetzt wird. Der Mangel an ausreichenden in-vitro Arbeiten, die die Kultivierung von Knorpelzellen in der DBM genauer untersuchen, machen vor dem tierexperimentellen und klinischen Einsatz weitere in-vitro Arbeiten notwendig. In dieser Arbeit wurde die Besiedlung der DBM mit humanen Chondrozyten untersucht und die Interaktion der Knorpelzellen mit der DBM in Hinblick auf einen möglichen proliferationshemmenden oder toxischen Effekt der DBM analysiert. Die einfache Besiedlung von humanen Chondrozyten in der großporigen DBM gelang nur vereinzelt, da es zu einer mangelnden Redifferenzierung und zum Zelltod kam. Deshalb erfolgte eine Besiedlung der DBM mit Chondrozyten-Makroaggregaten. In den histologischen und immunhistochemischen Färbungen konnte die Adhäsion und Proliferation der Chondrozyten-Makroaggregate in der DBM gezeigt werden. Doch im Gegensatz zu den Kontrollknorpelzellen, die nicht mit der DBM in Kontakt kamen, imponierten in der DBM morphologisch veränderte Zellen mit überwiegender Expression von Kollagen I. Die Redifferenzierung mit einer Wiederaufnahme der Expression von Kollagen II konnte nur in geringem Maße nachgewiesen werden. Der immunhistochemische Nachweis des Apoptosemarkers Caspase III in den Zell-DBM- Konstrukten und den Makroaggregaten, die mit dem Medium der DBM kultiviert wurden, legt die Induktion von Apoptose in den Chondrozyten durch die DBM nahe. Unter dem Verdacht auf eine Freisetzung toxischer Substanzen aus der DBM, u.a. von in der Aufarbeitung verwendeter Peressigsäure und Chloroform, wurden Untersuchungen des Zellkulturmediums durchgeführt. Rest¬bestände von Peressigsäure und Chloroform ließen sich mit den verwendeten Detektionssystemen nicht nachweisen. In einem weiteren Versuchsteil wurde mittels zweier Proliferationstests die Vitalität der Chondrozyten in Abhängigkeit von der Kulturdauer und Konzentration von DBM im Bezug zu einer Gruppe von Kontrollchondrozyten bestimmt. In den ersten 48 Stunden bestand mit steigender Konzentration der DBM ein Trend zu höheren Proliferationsraten und Vitalität der Chondrozyten im Vergleich zur Kontrolle. Nach 96 Stunden sanken die Werte dosisabhängig unter die der Kontrollzellen. Die parallel durchgeführten Vitalfärbungen, mittels der Lebend- (FDA-) und Tot- (PI-) Darstellung stimmen mit den Ergebnissen der Proliferationstests überein. Der immunhistochemische Nachweis von Caspase III, die sinkende Vitalität nach 96 Stunden im ELISA und die Anzahl an toten Zellen in den Vitalfärbungen sind nicht allein mit der Freisetzung von toxischen Restbeständen aus der DBM erklärbar. In den Grundlagenarbeiten von Urist et al. (1979) konnte die Freisetzung von chondrogen wirkenden Wachstumsfaktoren aus der DBM gezeigt werden. Der Verbrauch der Wachstumsfaktoren durch die Zellen, der steigende Bedarf durch die starke anfängliche Proliferation und der Wechsel des kompletten CGM alle zwei Tage, könnte nach 48 Stunden zum Entzug von Wachstumsfaktoren, welcher die Einleitung der Apoptose bedingen kann und damit zum immunhistochemischen Nachweis von Caspase III und sinkenden Proliferationsraten im ELISA geführt haben. Prinzipiell ist DBM als Trägermaterial im Tissue Engineering für humane Chondrozyten geeignet, doch die mangelnde Redifferenzierung und die Einleitung der Apoptose machen sowohl weitere toxikologische Untersuchungen als auch Arbeiten mit dem Einsatz von Wachstumsfaktoren notwendig.The reconstruction of traumatic, tumorous or congenital lesions in regiones of the head is necessary because of functional and aesthetic reasons. Tissue Engineering ist an important method for cultivating some few autologous cells with help of biomaterials. That is why this work examinates the hopefullness biomaterial of human demineralized bone matrix (DBM), which has been used for many years for reconstruction of bone in orthopaedic surgery, but also in otorhinolaryngology. For lack of in-vitro studies, analysing the cultivation of chondrocytes in DBM, more in vitro analysis are necessary before application in animal or clinical studies. In this work the seeding of chondrocytes in the DBM and its interaction with the DBM was tested in view of a possible inhibitory effect of the proliferation or toxicity of the DBM. The colonisation of human chondrocytes in the large pores of the DBM succeeded only isolated because of missing redifferentiation and cell death. Therefore DBM was populated with macroaggregates of chondrocytes. In the histological and immunhistochemical coloration the proliferation and adhesion of the cell – macroaggregates was shown in the DBM. In contrast to a controll group, cells which were not cultivated with DBM, impressed in the DBM morphological modified cells with expression of collagen I. The redifferentiation with reuptake of expression of collagen II was found in low amounts only. The detection of caspase III, a marker of apoptosis, in the cell-DBM-constructs and in the macroaggegates, cultivating with the medium of DBM, suggested the induction of apoptosis in the chondrocytes by DBM. Because of having the suspicion of the release of toxic agents of the DBM for example peracetic acid or chloroform, explained throw the sterilisation process, the cell culture medium was analysed. But neither peracetic acid nor chloroform were verified. In another experimental setting the vitality of chondrocytes depending on culture time and concentration of DBM was analized by two different proliferation tests with reference to a control group. Higher concentration of DBM tended to result in higher proliferation rates and vitality of the cells in contrast to the control group in the first 48 hours. After 96 hours the vitality sank dose-dependent below the control group. The synchronously performed vitality stain by vital (FDA) and non-vital (PI) coloring corresponded with the results of the proliferation tests. The detection of Caspase III, the decreasing vitality after 96 hours and the death cells are not only explained by the release of toxic agents of the DBM. The fundamental works of Urist et al. (1979) have shown the release of growth factors by the DBM, which induces the chondrogenesis. The consumption of growth factors by cells, the increasing need because oft he high proliferation rate in the beginning and the complete change oft he cell culture medium each two days, could lead to a lack of growth factors after 48 hours, which induced the apoptosis associated with the immunhistochemical detection of Caspase III and decreasing proliferation rates in ELISA analysis. In principle DBM is qualified as scaffold for Tissue Engineering of human chondroncytes. But the insufficient redifferentiation and the initiation of apoptosis necessitate further toxicological examinations as well as the use of growth factors

Redox Modulation and Induction of Ferroptosis as a New Therapeutic Strategy in Hepatocellular Carcinoma.

Ferroptosis, a newly discovered form of cell death mediated by reactive oxygen species (ROS) and lipid peroxidation, has recently been shown to have an impact on various cancer types; however, so far there are only few studies about its role in hepatocellular carcinoma (HCC). The delicate equilibrium of ROS in cancer cells has found to be crucial for cell survival, thus increased levels may trigger ferroptosis in HCC. In our study, we investigated the effect of different ROS modulators and ferroptosis inducers on a human HCC cell line and a human hepatoblastoma cell line. We identified a novel synergistic cell death induction by the combination of Auranofin and buthionine sulfoxime (BSO) or by Erastin and BSO at subtoxic concentrations. We found a caspase-independent, redox-regulated cell death, which could be rescued by different inhibitors of ferroptosis. Both cotreatments stimulated lipid peroxidation. All these findings indicated ferroptotic cell death. Both cotreatments affected the canonical ferroptosis pathway through GPX4 downregulation. We also found an accumulation of Nrf2 and HO-1, indicating an additional effect on the non-canonical pathway. Our results implicate that targeting these two main ferroptotic pathways simultaneously can overcome chemotherapy resistance in HCC

The modulation of redox homeostasis and induction of ferroptotic cell death in hepatocellular carcinoma as an anticancer strategy

Introduction: Ferroptosis has recently been identified as a form of programmed cell death caused by an accumulation of lipid reactive oxygen species (ROS). However, little is yet known about the role in hepatocellular carcinoma (HCC) and its signalling mechanism as well the modulation of ROS. Material and methods: Human HCC cell lines were treated with different concentrations of ROS modulators (Auranofin, Erastin, BSO). Cell death was determined by analysis of PI-stained nuclei using flow cytometry. ROS production and lipid peroxidation were analysed at early time points before cell death starts. For mechanistic studies we performed Western Blot and a Proteome array. Different inhibitors of cell death target proteins, ROS-scavengers as well as lipoxygenase inhibitors were used. To investigate the functional relevance of NAPDH oxidases (NOX) 1 and 4 for ROS modulation and ferroptosis we genetically silenced its genes using three distinct siRNAs and we used the NOX1/4-inhibitor GKT137831. Results and discussions: Compared to the single treatment, Auranofin/BSO-cotreatment as well as Erastin/BSO-cotreatment acted in concert to trigger cell death and to reduce cell viability of HCC cells in a dose- and time-dependent manner. Furthermore, both cotreatments induce ROS production, lipid peroxidation and ferroptotic cell death, which could be inhibited by the use of Ferrostatin-1 (inhibitor of lipid peroxidation) and Liproxstatin-1 (specific inhibitor of ferroptosis). The broad-range caspase inhibitor zVAD.fmk failed to rescue cells from Auranofin/BSO- or Erastin/BSO-cotreatment induced cell death. No activation of caspases-3 could be seen in the proteome profiler apoptosis assay. Importantly, the selective lipoxygenase (LOX) inhibitor Baicalain and the pan-LOX inhibitor NDGA protect HCC cells from Auranofin/BSO- and Erastin/BSO-cotreatment stimulated lipid peroxidation, ROS generation and cell death, indication that the induction of ferroptosis may bypass apoptosis resistance of HCC cells. Mechanistic studies showed that Auranofin/BSO-cotreatment decreased TrxR-activity, led to Nrf2 accumulation and promoted the activation of HO-1. In contrast, NOX 1 and 4 were involved in Erastin/BSO-mediated cell death and the use of the NOX1/4-inhibitor GKT137831 rescued HCC cells from the Erastin/BSO-induced cell death. Conclusion: By providing new insights into the molecular regulation of ROS and ferroptosis, our study contributes to the development of novel treatment strategies to reactivate programmed cell death in HCC cells

Redox modulation and induction of ferroptosis as a new therapeutic strategy in hepatocellular carcinoma

Ferroptosis, a newly discovered form of cell death mediated by reactive oxygen species (ROS) and lipid peroxidation, has recently been shown to have an impact on various cancer types; however, so far there are only few studies about its role in hepatocellular carcinoma (HCC). The delicate equilibrium of ROS in cancer cells has found to be crucial for cell survival, thus increased levels may trigger ferroptosis in HCC.In our study, we investigated the effect of different ROS modulators and ferroptosis inducers on a human HCC cell line and a human hepatoblastoma cell line. We identified a novel synergistic cell death induction by the combination of Auranofin and buthionine sulfoxime (BSO) or by Erastin and BSO at subtoxic concentrations. We found a caspase-independent, redox-regulated cell death, which could be rescued by different inhibitors of ferroptosis. Both cotreatments stimulated lipid peroxidation. All these findings indicated ferroptotic cell death. Both cotreatments affected the canonical ferroptosis pathway through GPX4 downregulation. We also found an accumulation of Nrf2 and HO-1, indicating an additional effect on the non-canonical pathway. Our results implicate that targeting these two main ferroptotic pathways simultaneously can overcome chemotherapy resistance in HCC

DIALAPP: a prospective validation of a new diagnostic algorithm for acute appendicitis

Purpose: The management of patients with suspected appendicitis remains a challenge in daily clinical practice, and the optimal management algorithm is still being debated. Negative appendectomy rates (NAR) continue to range between 10 and 15%. This prospective study evaluated the accuracy of a diagnostic pathway in acute appendicitis using clinical risk stratification (Alvarado score), routine ultrasonography, gynecology consult for females, and selected CT after clinical reassessment. Methods: Patients presenting with suspected appendicitis between November 2015 and September 2017 from age 18 years and above were included. Decision-making followed a clear management pathway. Patients were followed up for 6 months after discharge. The hypothesis was that the algorithm can reduce the NAR to a value of under 10%. Results: A total of 183 patients were included. In 65 of 69 appendectomies, acute appendicitis was confirmed by histopathology, corresponding to a NAR of 5.8%. Notably, all 4 NAR appendectomies had other pathologies of the appendix. The perforation rate was 24.6%. Only 36 patients (19.7%) received a CT scan. The follow-up rate after 30 days achieved 69%, including no patients with missed appendicitis. The sensitivity and specificity of the diagnostic pathway was 100% and 96.6%, respectively. The potential saving in costs can be as much as 19.8 million €/100,000 cases presenting with the suspicion of appendicitis. Conclusion: The risk-stratified diagnostic algorithm yields a high diagnostic accuracy for patients with suspicion of appendicitis. Its implementation can safely reduce the NAR, simultaneously minimizing the use of CT scans and optimizing healthcare-related costs in the treatment of acute appendicitis

MicroRNA Profiling of Laser-Microdissected Hepatocellular Carcinoma Reveals an Oncogenic Phenotype of the Tumor Capsule

Several microRNAs (miRNAs) are associated with the molecular pathogenesis of hepatocellular carcinoma (HCC). However, previous studies analyzing the dysregulation of miRNAs in HCC show heterogeneous results. We hypothesized that part of this heterogeneity might be attributable to variations of miRNA expression deriving from the HCC capsule or the fibrotic septa within the peritumoral tissue used as controls. Tissue from surgically resected hepatitis C–associated HCC from six well-matched patients was microdissected using laser microdissection and pressure catapulting technique. Four distinct histologic compartments were isolated: tumor parenchyma (TP), fibrous capsule of the tumor (TC), tumor-adjacent liver parenchyma (LP), and cirrhotic septa of the tumor-adjacent liver (LC). MiRNA expression profiling analysis of 1105 mature miRNAs and precursors was performed using miRNA microarray. Principal component analysis and consecutive pairwise supervised comparisons demonstrated distinct patterns of expressed miRNAs not only for TP versus LP (e.g., intratumoral down-regulation of miR-214, miR-199a, miR-146a, and miR-125a; P< .05) but also for TC versus LC (including down-regulation within TC of miR-126, miR-99a/100, miR-26a, and miR-125b; P< .05). The tumor capsule therefore demonstrates a tumor-like phenotype with down-regulation of well-known tumor-suppressive miRNAs. Variations of co-analyzed fibrotic tissue within the tumor or in controls may have profound influence on miRNA expression analyses in HCC. Several miRNAs, which are proposed to be HCC specific, may indeed be rather associated to the tumor capsule. As miRNAs evolve to be important biomarkers in liver tumors, the presented data have important translational implications on diagnostics and treatment in patients with HCC

Hepatitis B Virus DNA is a Substrate for the cGAS/STING Pathway but is not Sensed in Infected Hepatocytes

Hepatitis B virus (HBV) chronic infection is a critical risk factor for hepatocellular carcinoma. The innate immune response to HBV infection is a matter of debate. In particular, viral escape mechanisms are poorly understood. Our study reveals that HBV RNAs are not immunostimulatory in immunocompetent myeloid cells. In contrast, HBV DNA from viral particles and DNA replication intermediates are immunostimulatory and sensed by cyclic GMP-AMP Synthase (cGAS) and Stimulator of Interferon Genes (STING). We show that primary human hepatocytes express DNA sensors to reduced levels compared to myeloid cells. Nevertheless, hepatocytes can respond to HBV relaxed-circular DNA (rcDNA), when transfected in sufficient amounts, but not to HBV infection. Finally, our data suggest that HBV infection does not actively inhibit the DNA-sensing pathway. In conclusion, in infected hepatocytes, HBV passively evades recognition by cellular sensors of nucleic acids by (i) producing non-immunostimulatory RNAs, (ii) avoiding sensing of its DNAs by cGAS/STING without active inhibition of the pathway

Impact of the SARS-CoV-2 pandemic on emergency surgery services—a multi-national survey among WSES members

Background!#!The SARS-CoV-2 pandemic is a major challenge for health care services worldwide. It's impact on oncologic therapies and elective surgery has been described recently, and the literature provides guidelines regarding appropriate elective patient treatment during the pandemic. However, the impact of SARS-CoV-2 pandemic on emergency surgery services has been poorly investigated up to now.!##!Methods!#!A 17-item web survey had been distributed to emergency surgeons in June 2020 around the world, investigating the impact of SARS-CoV-2 pandemic on patients and septic diseases both requiring emergency surgery and the time-to-intervention in emergency surgery routine, as well as experiences with surgery in COVID-19 patients.!##!Results!#!Ninety-eight collaborators from 31 countries responded to the survey. The majority (65.3%) estimated the impact of the SARS-CoV-2 pandemic on emergency surgical patient care as being strong or very strong. Due to the pandemic, 87.8% reported a decrease in the total number of patients undergoing emergency surgery and approximately 25% estimated a delay of more than 2 h in the time-to-diagnosis and another 2 h in the time-to-intervention. Fifty percent make structural problems with in-hospital logistics (e.g. transport of patients, closed normal wards etc.) mainly responsible for delayed emergency surgery and the frequent need (56.1%) for a triage of emergency surgical patients. 56.1% of the collaborators observed more severe septic abdominal diseases during the pandemic, especially for perforated appendicitis and severe septic cholecystitis (41.8% and 40.2%, respectively). 62.2% had experiences with surgery in COVID-19-infected patients.!##!Conclusions!#!The results of The WSES COVID-19 emergency surgery survey are alarming. The combination of an estimated decrease in numbers of emergency surgical patients and an observed increase in more severe septic diseases may be a result of the fear of patients from infection with COVID-19 and a consecutive delayed hospital admission and diagnosis. A critical delay in time-to-diagnosis and time-to-intervention may be a result of changes in in-hospital logistics and operating room as well as intensive care capacities. Both reflect the potentially harmful impact of SARS-CoV-2 pandemic on emergency surgery services