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Intravital microscopy has revolutionized live-cell imaging by allowing the study of spatial–temporal cell dynamics in living animals. However, the complexity of the data generated by this technology has limited the development of effective computational tools to identify and quantify cell processes. Amongst them, apoptosis is a crucial form of regulated cell death involved in tissue homeostasis and host defense. Live-cell imaging enabled the study of apoptosis at the cellular level, enhancing our understanding of its spatial–temporal regulation. However, at present, no computational method can deliver robust detection of apoptosis in microscopy timelapses. To overcome this limitation, we developed ADeS, a deep learning-based apoptosis detection system that employs the principle of activity recognition. We trained ADeS on extensive datasets containing more than 10,000 apoptotic instances collected both in vitro and in vivo, achieving a classification accuracy above 98% and outperforming state-of-the-art solutions. ADeS is the first method capable of detecting the location and duration of multiple apoptotic events in full microscopy timelapses, surpassing human performance in the same task. We demonstrated the effectiveness and robustness of ADeS across various imaging modalities, cell types, and staining techniques. Finally, we employed ADeS to quantify cell survival in vitro and tissue damage in mice, demonstrating its potential application in toxicity assays, treatment evaluation, and inflammatory dynamics. Our findings suggest that ADeS is a valuable tool for the accurate detection and quantification of apoptosis in live-cell imaging and, in particular, intravital microscopy data, providing insights into the complex spatial–temporal regulation of this process.

Summary: Background: The incidence of endometrial cancer is increasing worldwide. While delays in diagnosis reduce survival, case molecular misclassification might be associated with under- and over-treatment. The objective of this study was to evaluate genetic alterations to detect and molecularly classify cases of endometrial cancer using non-invasive samples. Methods: Consecutive patients with incident endometrial cancer (N = 139) and controls (N = 107) from a recent Spanish case–control study were included in this analysis. Overall, 339 cervicovaginal samples (out of which 228 were clinician-collected and 111 were self-collected) were analysed using a test based on next-generation sequencing (NGS), which targets 47 genes. Immunohistochemical markers were evaluated in 133 tumour samples. A total of 159 samples were used to train the detection algorithm and 180 samples were used for validation. Findings: Overall, 73% (N = 94 out of 129 clinician-collected samples, and N = 66 out of 90 self-collected samples) of endometrial cancer cases had detectable mutations in clinician-collected and self-collected samples, while the specificity was 80% (79/99) for clinician-collected samples and 90% (19/21) for self-collected samples. The molecular classifications obtained using tumour samples and non-invasive gynaecologic samples in our study showed moderate-to-good agreement. The molecular classification of cases of endometrial cancer into four groups using NGS of both clinician-collected and self-collected cervicovaginal samples yielded significant differences in disease-free survival. The cases with mutations in POLE had an excellent prognosis, whereas the cases with TP53 mutations had the poorest clinical outcome, which is consistent with the data on tumour samples. Interpretation: This study classified endometrial cancer cases into four molecular groups based on the analysis of cervicovaginal samples that showed significant differences in disease-free survival. The molecular classification of endometrial cancer in non-invasive samples may improve patient care and survival by indicating the early need for aggressive surgery, as well as reducing referrals to highly specialized hospitals in cancers with good prognosis. Validation in independent sets will confirm the potential for molecular classification in non-invasive samples. Funding: This study was funded by a competitive grant from Instituto de Salud Carlos III through the projects PI19/01835, PI23/00790, and FI20/00031, CIBERESP CB06/02/0073 and CIBERONC CB16/12/00231, CB16/12/00234 (Co-funded by European Regional Development Fund. ERDF: A way to build Europe). Samples and data were provided by Biobank HUB-ICO-IDIBELL, integrated into the Spanish Biobank Network, and funded by the Instituto de Salud Carlos III (PT20/00171) and by Xarxa de Bancs de Tumors de Catalunya (XBTC) sponsored by Pla Director d’Oncologia de Catalunya. This work was supported in part by the AECC, Grupos estables (GCTRA18014MATI). It also counts with the support of the Secretariat for Universities and Research of the Department of Business and Knowledge of the Generalitat de Catalunya, and grants to support the activities of research groups 2021SGR01354 and 2021SGR1112.

Key Clinical Message This case aims to report an unusual clinical situation with uncommon and severe side effects, which can even be life threatening for the patient. The ENT and Hematology specialist should be aware of diagnosing and treating adequately.

Se presenta el caso de un varón de 33 años que ingresó por otomastoiditis complicada tras manipulación quirúrgica con aislamiento de Corynebacterium amycolatum, un microorganismo que está cobrando interés por su papel como patógeno en los últimos años. El interés del caso radica en lo infrecuente del mismo, ya que no se han descrito en la literatura revisada casos de otomastoiditis por esta bacteria. En el caso se describe cómo se llegó al diagnóstico y el manejo que se realizó. Además se realizó una revisión de la literatura publicada hasta el momento de infecciones por C. amycolatum. Consideramos de interés el caso, así como la revisión realizada, ya que es una entidad poco conocida y revisada previamente, por lo que este documento puede aportar información sobre la misma.

Abstract Anecdotal evidence suggests that community infection control measures during the COVID-19 outbreak have modified the number and natural history of acute surgical inflammatory processes (ASIP—appendicitis, cholecystitis, diverticulitis and perianal abscesses) admissions. This study aims to evaluate the impact of the COVID-19 pandemic on the presentation and treatment ASIP and quantify the effect of COVID-19 infection on the outcomes of ASIP patients. This was a multicentre, comparative study, whereby ASIP cases from 2019, 2020 and 2021 (March 14th to May 2nd) were analyzed. Data regarding patient and disease characteristics as well as outcomes, were collected from sixteen centres in Madrid, and one in Seville (Spain). The number of patients treated for ASIP in 2019 was 822 compared to 521 in 2020 and 835 in 2021. This 1/3rd reduction occurs mainly in patients with mild cases, while the number of severe cases was similar. Surgical standards suffered a step back during the first wave: Lower laparoscopic approach and longer length of stay. We also found a more conservative approach to the patients this year, non-justified by clinical circumstances. Luckily these standards improved again in 2021. The positive COVID-19 status itself did not have a direct impact on mortality. Strikingly, none of the 33 surgically treated COVID positive patients during both years died postoperatively. This is an interesting finding which, if confirmed through future research with a larger sample size of COVID-19 positive patients, can expedite the recovery phase of acute surgical services.

The Drosophila polarity protein Crumbs is essential for the establishment and growth of the apical domain in epithelial cells. The protein Yurt limits the ability of Crumbs to promote apical membrane growth, thereby defining proper apical/lateral membrane ratio that is crucial for forming and maintaining complex epithelial structures such as tubes or acini. Here, we show that Yurt also increases Myosin-dependent cortical tension downstream of Crumbs. Yurt overexpression thus induces apical constriction in epithelial cells. The kinase aPKC phosphorylates Yurt, thereby dislodging the latter from the apical domain and releasing apical tension. In contrast, the kinase Pak1 promotes Yurt dephosphorylation through activation of the phosphatase PP2A. The Pak1–PP2A module thus opposes aPKC function and supports Yurt-induced apical constriction. Hence, the complex interplay between Yurt, aPKC, Pak1, and PP2A contributes to the functional plasticity of Crumbs. Overall, our data increase our understanding of how proteins sustaining epithelial cell polarization and Myosin-dependent cell contractility interact with one another to control epithelial tissue architecture.

In emerging epithelial tissues, cells undergo dramatic rearrangements to promote tissue shape changes. Dividing cells remain interconnected via transient cytokinetic bridges. Bridges are cleaved during abscission and currently, the consequences of disrupting abscission in developing epithelia are not well understood. We show that the Rab GTPase Rab25 localizes near cytokinetic midbodies and likely coordinates abscission through endomembrane trafficking in the epithelium of the zebrafish gastrula during epiboly. In maternal-zygotic Rab25a and Rab25b mutant embryos, morphogenic activity tears open persistent apical cytokinetic bridges that failed to undergo timely abscission. Cytokinesis defects result in anisotropic cell morphologies that are associated with a reduction of contractile actomyosin networks. This slows cell rearrangements and alters the viscoelastic responses of the tissue, all of which likely contribute to delayed epiboly. We present a model in which Rab25 trafficking coordinates cytokinetic bridge abscission and cortical actin density, impacting local cell shape changes and tissue-scale forces.

The Target Of Rapamycin (TOR) signaling pathway is known to regulate growth in response to nutrient availability and stress in eukaryotic cells. In the present study, we have investigated the TOR pathway in the white-rot fungus Phanerochaete chrysosporium. Inhibition of TOR activity by rapamycin affects conidia germination and hyphal growth highlighting the conserved mechanism of susceptibility to rapamycin. Interestingly, the secreted protein content is also affected by the rapamycin treatment. Finally, homologs of the components of TOR pathway can be identified in P. chrysosporium. Altogether, those results indicate that the TOR pathway of P. chrysosporium plays a central role in this fungus.

Restriction enzymes are the bread and butter of Molecular Biology. Nonetheless, how restriction enzymes recognize and cleave their target is not always clear. When developing a method for the enzymatic production of oligonucleotides, we noticed that type II endonucleases BtsCI and BseGI, which recognize the sequence GGATGNN^, perform incomplete digestions of DNA hairpins, with the top strand nick not always occurring correctly. We tested the cutting of synthetic hairpins containing all possible combinations of dinucleotides following the recognition site and our results show that all sequences containing one adenine following GGATG were digested more efficiently. We further show that the same sequence preference is also observable in double stranded DNA at higher Mg2+ concentrations and even in optimal conditions. Kinetic results show that BtsCI has a noteworthy difference in the first-rate constants between different sequences and between the two catalytic domains. An increase in Mg2+ resulted in a drastic decrease in the catalytic activity of the top (sense) strand that wasn't always accompanied by a nick in the bottom strand (antisense).

Detection and monitoring of circulating tumor DNA (ctDNA) is rapidly becoming a diagnostic, prognostic and predictive tool in cancer patient care. A growing number of gene targets have been identified as diagnostic or actionable, requiring the development of reliable technology that provides analysis of multiple genes in parallel. We have developed the InVision™ liquid biopsy platform which utilizes enhanced TAm-Seq™ (eTAm-Seq™) technology, an amplicon-based next generation sequencing method for the identification of clinically-relevant somatic alterations at low frequency in ctDNA across a panel of 35 cancer-related genes.We present analytical validation of the eTAm-Seq technology across two laboratories to determine the reproducibility of mutation identification. We assess the quantitative performance of eTAm-Seq technology for analysis of single nucleotide variants in clinically-relevant genes as compared to digital PCR (dPCR), using both established DNA standards and novel full-process control material.The assay detected mutant alleles down to 0.02% AF, with high per-base specificity of 99.9997%. Across two laboratories, analysis of samples with optimal amount of DNA detected 94% mutations at 0.25%-0.33% allele fraction (AF), with 90% of mutations detected for samples with lower amounts of input DNA.These studies demonstrate that eTAm-Seq technology is a robust and reproducible technology for the identification and quantification of somatic mutations in circulating tumor DNA, and support its use in clinical applications for precision medicine.

El aneurisma micótico de la arteria glútea superior es excepcional, sólo se han publicado seis casos hasta el momento. Presentamos el caso de un varón de 65 años con un aneurisma micótico en la arteria glútea superior como presentación inicial de una endocarditis bacteriana. Mycotic aneurysm of the superior gluteal artery is exceptional, there are only six cases reported. This is the case of a 65-year-old man with a mycotic aneurysm, the first symtom of a bacterial endocarditis.

Axis elongation is a conserved process in which the head-to-tail or anterior-posterior (AP) axis of an embryo extends. In Drosophila, cellular rearrangements drive axis elongation. Cells exchange neighbours by converging into transient multicellular vertices which resolve through the assembly of new cell interfaces parallel to the AP axis. We found that new interfaces elongate in pulses correlated with periodic contractions of the surrounding cells. Inhibiting actomyosin contractility globally, or specifically in the cells around multicellular vertices, disrupted the rate and directionality of new interface assembly. Laser ablation indicated that new interfaces sustained greater tension than non-elongating ones. We developed a method to apply ectopic tension and found that increasing AP tension locally increased the elongation rate of new edges by more than twofold. Increasing dorsal-ventral tension resulted in vertex resolution perpendicular to the AP direction. We propose that local, periodic contractile forces polarize vertex resolution to drive Drosophila axis elongation.

Hypoxia and pressure-overload induce heme oxygenase-1 (HO-1) in cardiomyocytes and vascular smooth muscle cells (VSMCs). HO-1(-/-) mice exposed to chronic hypoxia develop pulmonary arterial hypertension (PAH) with exaggerated right ventricular (RV) injury consisting of dilation, fibrosis, and mural thrombi. Our objective was to identify the HO-1 product(s) mediating RV protection from hypoxic injury in HO-1(-/-) mice.HO-1(-/-) mice were exposed to seven weeks of hypoxia and treated with inhaled CO or biliverdin injections. CO reduced right ventricular systolic pressure (RVSP) and prevented hypoxic pulmonary arteriolar remodeling in both HO-1(-/-) and control mice. Biliverdin had no significant effect on arteriolar remodeling or RVSP in either genotype. Despite this, biliverdin prevented RV failure in the hypoxic HO-1(-/-) mice (0/14 manifested RV wall fibrosis or thrombus), while CO-treated HO-1(-/-) mice developed RV insults similar to untreated controls. In vitro, CO inhibited hypoxic VSMC proliferation and migration but did not prevent cardiomyocyte death from anoxia-reoxygenation (A-R). In contrast, bilirubin limited A-R-induced cardiomyocyte death but did not inhibit VSMC proliferation and migration.CO and bilirubin have distinct protective actions in the heart and pulmonary vasculature during chronic hypoxia. Moreover, reducing pulmonary vascular resistance may not prevent RV injury in hypoxia-induced PAH; supporting RV adaptation to hypoxia and preventing RV failure must be a therapeutic goal.