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Abstract Genetic variants in relevant genes coexisting with MRI lesions in children with drug‐resistant epilepsy (DRE) can negatively influence epilepsy surgery outcomes. Still, presurgical evaluation does not include genetic diagnostics routinely. Here, we report our presurgical evaluation algorithm that includes routine genetic testing. We analyzed retrospectively the data of 68 children with DRE operated at a mean age of 7.8 years (IQR: 8.1 years) at our center. In 49 children, genetic test results were available. We identified 21 gene variants (ACMG III: n = 7, ACMG IV: n = 2, ACMG V: n = 12) in 19 patients (45.2%) in the genes TSC1, TSC2, MECP2, DEPDC5, HUWE1, GRIN1, ASH1I, TRIO, KIF5C, CDON, ANKD11, TGFBR2, ATN1, COL4A1, JAK2, KCNQ2, ATP1A2, and GLI3 by whole‐exome sequencing as well as deletions and duplications by array CGH in six patients. While the results did not change the surgery indication, they supported counseling with respect to postoperative chance of seizure freedom and weaning of antiseizure medication (ASM). The presence of genetic findings leads to the postoperative retention of at least one ASM. In our cohort, the International League against Epilepsy (ILAE) seizure outcome did not differ between patients with and without abnormal genetic findings. However, in the 7/68 patients with an unsatisfactory ILAE seizure outcome IV or V 12 months postsurgery, 2 had an abnormal or suspicious genetic finding as a putative explanation for persisting seizures postsurgery, and 3 had received palliative surgery including one TSC patient. This study highlights the importance of genetic testing in children with DRE to address putative underlying germline variants as genetic epilepsy causes or predisposing factors that guide patient and/or parent counseling on a case‐by‐case with respect to their individual chance of postoperative seizure freedom and ASM weaning. Plain Language Summary Genetic variants in children with drug‐resistant epilepsy (DRE) can negatively influence epilepsy surgery outcomes. However, presurgical evaluation does not include genetic diagnostics routinely. This retrospective study analyzed the genetic testing results of the 68 pediatric patients who received epilepsy surgery in our center. We identified 21 gene variants by whole‐exome sequencing as well as deletions and duplications by array CGH in 6 patients. These results highlight the importance of genetic testing in children with DRE to guide patient and/or parent counseling on a case‐by‐case with respect to their individual chance of postoperative seizure freedom and ASM weaning.

Abstract The importance of halogen bonds (XBs) in the regulation of material properties through a variation in the electrostatic potential of the halogen atom is not attracted much attention. Herein, this study utilizes in situ single crystal X‐ray diffraction and synchrotron‐based X‐ray techniques to investigate the cooling‐triggered irreversible single‐crystal‐to‐single‐crystal transformation of the DMF solvated iodo‐substituted squaraine dye (SQD‐I). Transformation is observed to be mediated by solvent‐involved XB formation and strengthening of electrostatic interaction between adjacent SQD‐I molecules. By immersing a DMF solvate in acetonitrile a solvent exchange without loss of long‐range ordering is observed. This is attributed to conservation of the molecular charge distribution of SQD‐I molecules during the process. The different solvates can be used in combination for temperature‐dependent image encryption. This work emphasizes the changes caused by XB formation to the electrostatic potentials of halogen containing molecules and their influence on material properties and presents the potential utility of XBs in the design of soft‐porous crystals and luminescent materials.

Abstract Primary human trophoblast stem cells (TSCs) and TSCs derived from human pluripotent stem cells (hPSCs) can potentially model placental processes in vitro. Yet, the pluripotent states and factors involved in the differentiation of hPSCs to TSCs remain poorly understood. In this study, we demonstrate that the primed pluripotent state can generate TSCs by activating pathways such as Epidermal Growth Factor (EGF) and Wingless-related integration site (WNT), and by suppressing tumor growth factor beta (TGFβ), histone deacetylases (HDAC), and Rho-associated protein kinase (ROCK) signaling pathways, all without the addition of exogenous Bone morphogenetic protein 4 (BMP4)—a condition we refer to as the TS condition. We characterized this process using temporal single-cell RNA sequencing to compare TS conditions with differentiation protocols involving BMP4 activation alone or BMP4 activation in conjunction with WNT inhibition. The TS condition consistently produced a stable, proliferative cell type that closely mimics first-trimester placental cytotrophoblasts, marked by the activation of endogenous retroviral genes and the absence of amnion expression. This was observed across multiple cell lines, including various primed induced pluripotent stem cell (iPSC) and embryonic stem cell (ESC) lines. Primed-derived TSCs can proliferate for over 30 passages and further specify into multinucleated syncytiotrophoblasts and extravillous trophoblast cells. Our research establishes that the differentiation of primed hPSCs to TSC under TS conditions triggers the induction of TMSB4X, BMP5/7, GATA3, and TFAP2A without progressing through a naive state. These findings propose that the primed hPSC state is part of a continuum of potency with the capacity to differentiate into TSCs through multiple routes.

IntroductionHead-mounted displays (HMDs) that superimpose holograms onto patients are of particular surgical interest as they are believed to dramatically change surgical procedures by including safety warning and allowing real-time offsite consultations. Although there are promising benefits of mixed and augmented reality (MR/AR) technologies in surgery, they also raise new ethical concerns. The aim of this systematic review is to determine the full spectrum of ethical issues that is raised for surgeons in the intraoperative application of MR/AR technology.MethodsFive bibliographic databases were searched for publications on the use of MR/AR, HMDs and other devices, their intraoperative application in surgery, and ethical issues. We applied qualitative content analysis to the n = 50 articles included. Firstly, we coded the material with deductive categories derived from ethical frameworks for surgical innovations, complications and research. Secondly, clinical aspects with ethical relevance were inductively coded as ethical issues within the main categories. Thirdly, we pooled the ethical issues into themes and sub-themes. We report our findings according to the reporting guideline RESERVE.ResultsWe found n = 143 ethical issues across ten main themes, namely patient-physician relationship, informed consent, professionalism, research and innovation, legal and regulatory issues, functioning equipment and optimal operating conditions, allocation of resources, minimizing harm, good communication skills and the ability to exercise sound judgement. The five most prevalent ethical issues are “Need for continuous research and innovation”, “Ensuring improvement of the learning curve”, “MR/AR enables new maneuvers for surgeons”, “Ensuring improvement of comfort, ergonomics, and usability of devices,” and “Not withholding MR/AR if it performs better”.ConclusionsRecognizing the evidence-based limitations of the intraoperative MR/AR application is of paramount importance to avoid ethical issues, but clinical trials in surgery pose particular ethical risks for patients. Regarding the digital surgeon, long-term impact on human workforce, potentially harmful “negative training,” i.e., acquiring inappropriate behaviors, and the fear of surveillance need further attention. MR/AR technologies offer not only challenges but significant advantages, promoting a more equitable distribution of surgical expertise and optimizing healthcare. Aligned with the core principle of social justice, these technologies enable surgeons to collaborate globally, improving training conditions and addressing enduring global healthcare inequalities.

Introduction Atrial fibrillation is highly prevalent in patients on chronic dialysis. It is unclear whether anticoagulant therapy for stroke prevention is beneficial in these patients. Vitamin K-antagonists (VKA) remain the predominant anticoagulant choice. Importantly, anticoagulation remains inconsistently used and a possible benefit remains untested in randomised clinical trials comparing oral anticoagulation with no treatment in patients on chronic dialysis. The Danish Warfarin-Dialysis (DANWARD) trial aims to investigate the safety and efficacy of VKAs in patients with atrial fibrillation on chronic dialysis. The hypothesis is that VKA treatment compared with no treatment is associated with stroke risk reduction and overall benefit.Methods and analysis The DANWARD trial is an investigator-initiated trial at 13 Danish dialysis centres. In an open-label randomised clinical trial study design, a total of 718 patients with atrial fibrillation on chronic dialysis will be randomised in a 1:1 ratio to receive either standard dose VKA targeting an international normalised ratio of 2.0–3.0 or no oral anticoagulation. Principal analyses will compare the risk of a primary efficacy endpoint, stroke or transient ischaemic attack and a primary safety endpoint, major bleeding, in patients allocated to VKA treatment and no treatment, respectively. The first patient was randomised in October 2019. Patients will be followed until 1 year after the inclusion of the last patient.Ethics and dissemination The study protocol was approved by the Regional Research Ethics Committee (journal number H-18050839) and the Danish Medicines Agency (case number 2018101877). The trial is conducted in accordance with the Helsinki declaration and standards of Good Clinical Practice. Study results will be disseminated to participating sites, at research conferences and in peer-reviewed journals.Trial registration numbers NCT03862859, EUDRA-CT 2018-000484-86 and CTIS ID 2022-502500-75-00.

BackgroundSARS-CoV-2 vaccination in rheumatoid arthritis (RA) patients treated with B cell-depleting drugs induced limited seroconversion but robust cellular response. We aimed to document specific T and B cell immunity in response to vaccine booster doses and breakthrough infection (BTI).MethodsWe included 76 RA patients treated with rituximab who received up to four SARS-CoV-2 vaccine doses or three doses plus BTI, in addition to vaccinated healthy donors (HD) and control patients treated with tumor necrosis factor inhibitor (TNFi). We quantified anti-SARS-CoV-2 receptor-binding domain (RBD) Spike IgG, anti-nucleocapsid (NC) IgG, 92 circulating inflammatory proteins, Spike-binding B cells, and Spike-specific T cells along with comprehensive high-dimensional phenotyping and functional assays.FindingsThe time since the last rituximab infusion, persistent inflammation, and age were associated with the anti-SARS-CoV-2 RBD IgG seroconversion. The vaccine-elicited serological response was accompanied by an incomplete induction of peripheral Spike-specific memory B cells but occurred independently of T cell responses. Vaccine- and BTI-elicited cellular immunity was similar between RA and HD ex vivo in terms of frequency or phenotype of Spike-specific cytotoxic T cells and in vitro in terms of the functionality and differentiation profile of Spike-specific T cells.InterpretationSARS-CoV-2 vaccination in RA can induce persistent effector T-cell responses that are reactivated by BTI. Paused rituximab medication allowed serological responses after a booster dose (D4), especially in RA with lower inflammation, enabling efficient humoral and cellular immunity after BTI, and contributed overall to the development of potential durable immunity.

Abstract Although epilepsy surgery is the only curative therapeutic approach for lesional drug‐resistant epilepsy (DRE), there is reluctance to operate on infants due to a fear of complications. A recent meta‐analysis showed that epilepsy surgery in the first 6 months of life can achieve seizure control in about two thirds of children. However, robust data on surgical complications and postoperative cognitive development are lacking. We performed a retrospective multicenter study of infants who underwent epilepsy surgery in the first 6 months of life. 15 infants underwent epilepsy surgery at a median age of 134 days (IQR: 58) at four centers. The most common cause was malformation of cortical development, and 13 patients underwent a hemispherotomy. Two thirds required intraoperative red blood transfusions. Severe intraoperative complications occurred in two patients including death in one infant due to cardiovascular insufficiency. At a median follow‐up of 1.5 years (IQR: 1.8), 57% of patients were seizure‐free. Three patients where reoperated at a later age, resulting in 79% seizure freedom. Anti‐seizure medication could be reduced in two thirds, and all patients improved in their development. Our findings suggest that early epilepsy surgery can result in good seizure control and developmental improvement. However, given the perioperative risks, it should be performed only in specialized centers.

Passerines are the most successfully diversified bird order (around 60% of all avian species). They have developed complicated songs to defend their territories and to attract females for mating that can evolve quickly due to cultural transmission. Complex singing as well as plumage coloration of male birds are honest signals for potential partners and provide information about the males’ quality. To function as honest signals, both traits must be costly for the males. Of course, not all passerine species are equally clever or beautiful. Even within a single family of 50 to 70 species, relevant traits may vary considerably. Tits and chickadees (Paridae) comprise species of similar size, varying a lot in coloration and plumage pattern. The territorial songs are relatively short and simple. We investigated the relationship between song complexity and plumage coloration, taking phylogenetic relationships into account. We studied 55 out of the 64 species with 1084 song recordings retrieved from an online database. In the best model, besides colorfulness, body size had a negative impact on song complexity. Large colorful species were found to sing less complex songs. This result supports the hypothesis of a trade-off between costly traits and their likely intense signal function. This study contributes to a better understanding of how sexual selection influences the diversification of traits. In addition, we found that despite the relatively uniform size, the general negative correlation between body size and song frequency can be recovered. Some song traits are further influenced by distribution and thus by interspecific differences in climate niche.

Located at the crossroads of two continents and at the southeastern edge of the Mediterranean Basin, Anatolia was one of the most important Pleistocene glacial refugia in the Western Palaearctic. As part of the Irano-Anatolian, Caucasus and Mediterranean Basin biodiversity hotspots, this region is also home to a rich avian community including nearly 400 breeding species. Nevertheless, research addressing the genetic structure and diversity of local bird populations is limited, and information on glacial refugia in this region is still scant, especially when compared to other large Mediterranean peninsulas, namely the Balkan, Italian and Iberian ones. In this study, we contribute to filling this gap by addressing the biogeographic pattern of four common resident songbirds—the Eurasian blue tit (Cyanistes caeruleus), the great tit (Parus major), the Eurasian chaffinch (Fringilla coelebs) and the Eurasian blackbird (Turdus merula)—and one endemic species—the Krüper’s nuthatch (Sitta krueperi)—by amplifying two mitochondrial DNA genes in individuals from Anatolia (n = 329) and comparing their sequences to those of conspecifics from the rest of their distribution range across the western Palaearctic (n = 357) deposited in public databases. The overall genetic structure of these species is consistent with a scenario of isolation for multiple populations in different refugia across Anatolia and subsequent secondary contact in the wake of ice retreat, which makes this region a hotspot of genetic diversity for both widespread and endemic avian species.

Background: Immune checkpoint blockade (ICB) has presented a breakthrough in the treatment of malignant tumors and increased the overall survival of patients with various tumor entities. ICB may also cause immune-related adverse events, such as pneumonitis or interstitial lung disease. The lung clearance index (LCI) is a multiple-breath washout technique offering information on lung pathology in addition to conventional spirometry. It measures the degree of pulmonary ventilation inhomogeneity and allows early detection of pulmonary damage, especially that to peripheral airways. Methods: This cross-sectional study compared the lung function of patients with melanoma or metastatic cutaneous squamous cell carcinoma who received programmed cell death 1 (PD-1) and cytotoxic T-Lymphocyte-associated Protein 4 (CTLA-4) antibodies, alone or in combination, to age- and sex-matched controls. Lung function was assessed using spirometry, according to American Thoracic Society and European Respiratory Society standards, the LCI, and a diffusion capacity of carbon monoxide (DLCO) measurement. Results: Sixty-one screened patients and thirty-eight screened controls led to nineteen successfully included pairs. The LCI in the ICB-treated patients was 8.41 ± 1.15 (mean ± SD), which was 0.32 higher compared to 8.07 ± 1.17 in the control group, but the difference was not significant (p = 0.452). The patients receiving their ICB therapy for under five months showed a significantly lower LCI (7.98 ± 0.77) compared to the ICB patients undergoing therapy for over five months (9.63 ± 1.22) at the point of testing (p = 0.014). Spirometric analysis revealed that the forced expiratory volume between 25 and 75% of the forced vital capacity (FEF25–75%) in the ICB-treated patients was significantly reduced (p = 0.047) compared to the control group. DLCO (%predicted and adjusted for hemoglobin) was 94.4 ± 19.7 in the ICB patients and 93.4 ± 21.7 in the control group (p = 0.734). Conclusions: The patients undergoing ICB therapy showed slightly impaired lung function compared to the controls. Longer periods of ICB treatment led to deterioration of the LCI, which may be a sign of a subclinical inflammatory process. The LCI is feasible and may be easily integrated into the clinical daily routine and could contribute to early detection of pulmonary (auto-)inflammation.

Background: Non-melanoma skin cancers (NMSCs) are responsible for up to one-third of all human malignancies. Surgery is usually the treatment of choice, but patients often experience pain during the procedure. Topical rhenium-188 resin skin cancer treatment (RSCT) may be a valid therapeutic alternative. Methods: In this monocentric pilot study, 19 patients suffering from NMSC were treated with RSCT. Most of these patients had also experienced surgery, either because they developed a new NMSC in aftercare, or they had suffered previously from NMSC. Three RSCT-treated patients, who had no exposure to surgery so far, were paired with three matched patients, who had received surgery. We sought to evaluate and compare the patients’ experience with both treatments. A questionnaire assessed patients’ perceptions regarding side effects, aesthetic outcomes, wound care, fear of complications, and personal treatment preferences. Patients evaluated the different parameters of their either RSCT- or surgery-treated lesions on a scale from 0–10. Results: Patients were more afraid of complications before surgery than before RSCT (p = 0.04). Treatment with RSCT caused significantly less pain on treatment day (mean 0.56) than surgery (mean 2.32) (0 no pain, 10 maximum pain) (p = 0.02) and 14 days after the procedure (mean 0.89 versus mean 2.47) (p = 0.02). On day 14, RSCT-treated lesions were also significantly less itchy (mean 0.34) than after surgery (mean 1.50). Most patients were very satisfied with the aesthetic outcome after both RSCT (mean 8.42) and surgery (mean 8.31) (p = 0.89). In the case of a new NMSC, the majority of patients who experienced both treatments would rather be treated primarily with RSCT (44%) or would consider both options (31%); only 19% preferred surgery. Conclusion: Patients evaluated RSCT as less painful than surgery. The aesthetic outcomes of both treatments were comparable. For pain-sensitive patients, RSCT might be a preferable treatment option.

This work investigates the proposed enhanced efficacy of photodynamic therapy (PDT) by activating photosensitizers (PSs) with Cherenkov light (CL). The approaches of Yoon et al. to test the effect of CL with external radiation were taken up and refined. The results were used to transfer the applied scheme from external radiation therapy to radionuclide therapy in nuclear medicine. Here, the CL for the activation of the PSs (psoralen and trioxsalen) is generated by the ionizing radiation from rhenium-188 (a high-energy beta-emitter, Re-188). In vitro cell survival studies were performed on FaDu, B16 and 4T1 cells. A characterization of the PSs (absorbance measurement and gel electrophoresis) and the CL produced by Re-188 (luminescence measurement) was performed as well as a comparison of clonogenic assays with and without PSs. The methods of Yoon et al. were reproduced with a beam line at our facility to validate their results. In our studies with different concentrations of PS and considering the negative controls without PS, the statements of Yoon et al. regarding the positive effect of CL could not be confirmed. There are slight differences in survival fractions, but they are not significant when considering the differences in the controls. Gel electrophoresis showed a dominance of trioxsalen over psoralen in conclusion of single and double strand breaks in plasmid DNA, suggesting a superiority of trioxsalen as a PS (when irradiated with UVA). In addition, absorption measurements showed that these PSs do not need to be shielded from ambient light during the experiment. An observational test setup for a PDT nuclear medicine approach was found. The CL spectrum of Re-188 was measured. Fluctuating inconclusive results from clonogenic assays were found.