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International audience; 1H-benzo[d]imidazole derivatives exhibit antitubercular activity in vitro at a nanomolar range of concentrations and are not toxic to human cells, but their mode of action remains unknown. Here, we showed that these compounds are active against intracellular Mycobacterium tuberculosis. To identify their target, we selected drug-resistant M. tuberculosis mutants and then used whole-genome sequenc-ing to unravel mutations in the essential mmpL3 gene, which encodes the integral membrane protein that catalyzes the export of trehalose monomycolate, a precursor of the mycobacterial outer membrane component trehalose dimycolate (TDM), as well as mycolic acids bound to arabinogalactan. The drug-resistant phenotype was also observed in the parental strain overexpressing the mmpL3 alleles carrying the mutations identified in the resistors. However, no cross-resistance was observed between 1H-benzo[d]imidazole derivatives and SQ109, another MmpL3 inhibitor, or other first-line antitubercular drugs. Metabolic labeling and quantitative thin-layer chromatography (TLC) analysis of radiolabeled lipids from M. tuberculosis cultures treated with the benzoimidazoles indicated an inhibition of trehalose dimycolate (TDM) synthesis, as well as reduced levels of mycolylated arabinogalactan, in agreement with the inhibition of MmpL3 activity. Overall, this study emphasizes the pronounced activity of 1H-benzo[d]imidazole derivatives in interfering with mycolic acid metabolism and their potential for therapeutic application in the fight against tuberculosis.

Brugada syndrome (BrS), early repolarization syndrome (ERS), and idiopathic ventricular fibrillation (iVF) have long been considered primary electrical disorders associated with malignant ventricular arrhythmia and sudden cardiac death. However, recent studies have revealed the presence of subtle microstructural abnormalities of the extracellular matrix in some cases of BrS, ERS, and iVF, particularly within right ventricular subepicardial myocardium. Substrate-based ablation within this region has been shown to ameliorate the electrocardiographic phenotype and to reduce arrhythmia frequency in BrS. Patients with ERS and iVF may also exhibit low-voltage and fractionated electrograms in the ventricular subepicardial myocardium, which can be treated with ablation. A significant proportion of patients with BrS and ERS, as well as some iVF survivors, harbor pathogenic variants in the voltage-gated sodium channel gene, SCN5A , but the majority of genetic susceptibility of these disorders is likely to be polygenic. Here, we postulate that BrS, ERS, and iVF may form part of a spectrum of subtle subepicardial cardiomyopathy. We propose that impaired sodium current, along with genetic and environmental susceptibility, precipitates a reduction in epicardial conduction reserve, facilitating current-to-load mismatch at sites of structural discontinuity, giving rise to electrocardiographic changes and the arrhythmogenic substrate.

Zebrafish hearts can regenerate by replacing damaged tissue with new cardiomyocytes. Although the steps leading up to the proliferation of surviving cardiomyocytes have been extensively studied, little is known about the mechanisms that control proliferation and redifferentiation to a mature state. We found that the cardiac dyad, a structure that regulates calcium handling and excitation-contraction coupling, played a key role in the redifferentiation process. A component of the cardiac dyad called leucine-rich repeat–containing 10 (Lrrc10) acted as a negative regulator of proliferation, prevented cardiomegaly, and induced redifferentiation. We found that its function was conserved in mammalian cardiomyocytes. This study highlights the importance of the underlying mechanisms required for heart regeneration and their application to the generation of fully functional cardiomyocytes.

Sympathovagal balance is important in the pathogenesis of hypertension and independently associated with mortality. We evaluated the value of automated analysis of cross-correlation baroreflex sensitivity (xBRS) and heart rate variability (HRV) and its relationship with clinical covariates in 13,326 participants from the multi-ethnic HELIUS study. Finger blood pressure (BP) was continuously recorded, from which xBRS, standard deviation of normal-to-normal intervals (SDNN), and squared root of mean squared successive difference between normal-to-normal intervals (RMSDD) were determined. A subset of 3356 recordings > 300 s was used to derive the minimally required duration by comparing shortened to complete recordings, defined as intraclass correlation (ICC) > 0.90. For xBRS and SDNN, 120 s and 180 s were required (ICC 0.93); for RMSDD, 60 s (ICC 0.94) was sufficient. We included 10,252 participants (median age 46 years, 54% women) with a recording > 180 s for the regression. xBRS, SDNN, and RMSDD decreased linearly up to 50 years of age. For xBRS, there was a signification interaction with sex, with for every 10 years a decrease of 4.3 ms/mmHg (95%CI 4.0–4.6) for men and 5.9 ms/mmHg (95%CI 5.6–6.1) for women. Using splines, we observed sex-dependent nonlinearities in the relation with BP, waist-to-hip-ratio, and body mass index. Future studies can help unravel the dynamics of these relations and assess their predictive value. Graphical Abstract Panel 1 depicts automatic analysis and filtering of finger BP recordings, panel 2 depicts computation of xBRS from interpolated beat to beat data of systolic BP and interbeat interval, and (IBI) SDNN and RMSDD are computed directly from the filtered IBI dataset. Panel 3 depicts the results of large-scale analysis and relation of xBRS with age, sex, blood pressure and body mass index.

SummaryTranscriptional bursting has been linked to the stochastic positioning of nucleosomes. However, how bursting is regulated by remodeling of promoter nucleosomes is unknown. Here, we use single-molecule live-cell imaging of GAL10 transcription in budding yeast to measure how transcriptional bursting changes upon single and double perturbations of chromatin remodeling factors, the transcription factor Gal4 and preinitiation complex (PIC) components. Using dynamic epistasis analysis, we reveal how remodeling of different nucleosomes regulates individual transcriptional bursting parameters. At the nucleosome covering the Gal4 binding sites, RSC acts synergistically with Gal4 binding to facilitate each burst. Conversely, nucleosome remodeling at the TATA box controls only the first burst upon galactose induction. In the absence of remodelers, nucleosomes at canonical TATA boxes are displaced by TBP binding to allow for transcription activation. Overall, our results reveal how promoter nucleosome remodeling, together with transcription factor and PIC binding regulates the kinetics of transcriptional bursting.

Background The reactivation of genetic programs from early development is a common mechanism for injury-induced organ regeneration. T-box 3 (TBX3) is a member of the T-box family of transcription factors previously shown to regulate pluripotency and subsequent lineage commitment in a number of tissues, including limb and lung. TBX3 is also involved in lung and heart organogenesis. Here, we provide a comprehensive and thorough characterization of TBX3 and its role during pancreatic organogenesis and regeneration. Results We interrogated the level and cell specificity of TBX3 in the developing and adult pancreas at mRNA and protein levels at multiple developmental stages in mouse and human pancreas. We employed conditional mutagenesis to determine its role in murine pancreatic development and in regeneration after the induction of acute pancreatitis. We found that Tbx3 is dynamically expressed in the pancreatic mesenchyme and epithelium. While Tbx3 is expressed in the developing pancreas, its absence is likely compensated by other factors after ablation from either the mesenchymal or epithelial compartments. In an adult model of acute pancreatitis, we found that a lack of Tbx3 resulted in increased proliferation and fibrosis as well as an enhanced inflammatory gene programs, indicating that Tbx3 has a role in tissue homeostasis and regeneration. Conclusions TBX3 demonstrates dynamic expression patterns in the pancreas. Although TBX3 is dispensable for proper pancreatic development, its absence leads to altered organ regeneration after induction of acute pancreatitis.

Background: This study aimed to assess the feasibility of postmortem ultra-high-field magnetic resonance imaging (UHF-MRI) to study fetal musculoskeletal anatomy and explore the contribution of variation in iodine and formaldehyde (paraformaldehyde, PFA) treatment of tissue. Methods: Seven upper extremities from human fetuses with gestational ages of 19 to 24 weeks were included in this experimental study, approved by the Medical Research Ethics Committee. The specimens were treated with various storage (0.2–4% PFA) and staining (Lugol’s solution) protocols and the wrist joint was subsequently imaged with 7.0 T UHF-MRI. Soft-tissue contrast was quantified by determining regions of interest within a chondrified carpal bone (CCB) from the proximal row, the triangular fibrocartilage (TFC), and the pronator quadratus muscle (PQM) and calculating the contrast ratios (CRs) between mean signal intensities of CCB to TFC and CCB to PQM. Results: UHF-MRI showed excellent soft-tissue contrast in different musculoskeletal tissues. Increasing storage time in 4% PFA, CRs decreased, resulting in a shift from relatively hyperintense to hypointense identification of the CCB. Storage in 0.2% PFA barely influenced the CRs over time. Lugol’s solution caused an increase in CRs and might have even contributed to the inversion of the CRs. Conclusions: UHF-MRI is a feasible technique to image musculoskeletal structures in fetal upper extremities and most successful after short storage in 4% PFA or prolonged storage in 0.2% PFA. The use of Lugol’s solution is not detrimental on soft-tissue MRI contrast and therefore enables effectively combining UHF-MRI with contrast-enhanced micro-computed tomography using a single preparation of the specimen. Relevance statement: UHF-MRI can be performed after CE-micro-CT to take advantage of both techniques. Key points: • UHF-MRI is feasible to study human fetal cartilaginous and ligamentous anatomy. • Storage in low PFA concentrations (i.e., 0.2%) improves soft-tissue contrast in UHF-MRI. • Limited preservation time in high concentrations of PFA improves soft-tissue contrast in UHF-MRI. • Prior staining with Lugol’s solution does not reduce soft-tissue contrast in UHF-MRI. Graphical Abstract: [Figure not available: see fulltext.].

Primary amoebic meningoencephalitis and granulomatous amoebic encephalitis are distressing infections of the central nervous system caused by brain-eating amoebae, namely, Naegleria fowleri and Acanthamoeba spp., respectively, and present mortality rates of over 90%. No single drug has been approved for use against these infections, and current therapy is met with an array of obstacles including high toxicity and limited specificity. Thus, the development of alternative effective chemotherapeutic agents for the management of infections due to brain-eating amoebae is a crucial requirement to avert future mortalities. In this paper, we synthesized a conducting polymer-based nanocomposite entailing polyaniline (PANI) and molybdenum disulfide (MoS2) and explored its anti-trophozoite and anti-cyst potentials against Acanthamoeba castellanii and Naegleria fowleri. The intracellular generation of reactive oxygen species (ROS) and ultrastructural appearances of amoeba were also evaluated with treatment. Throughout, treatment with the 1:2 and 1:5 ratios of PANI/MoS2 at 100 μg/mL demonstrated significant anti-amoebic effects toward A. castellanii as well as N. fowleri, appraised to be ROS mediated and effectuate physical alterations to amoeba morphology. Further, cytocompatibility toward human keratinocyte skin cells (HaCaT) and primary human corneal epithelial cells (pHCEC) was noted. For the first time, polymer-based nanocomposites such as PANI/MoS2 are reported in this study as appealing options in the drug discovery for brain-eating amoebae infections.

Background: The cellular mechanisms underlying progression from paroxysmal to persistent atrial fibrillation (AF) are not fully understood, but alterations in (late) sodium current (INa) have been proposed. Human studies investigating electrophysiological changes at the paroxysmal stage of AF are sparse, with the majority employing right atrial appendage cardiomyocytes (CMs). We here investigated action potential (AP) characteristics and (late) INa remodelling in left atrial appendage CMs (LAA-CMs) from patients with paroxysmal and persistent AF and patients in sinus rhythm (SR), as well as the potential contribution of the “neuronal” sodium channel SCN10A/NaV1.8. Methods: Peak INa, late INa and AP properties were investigated through patch-clamp analysis on single LAA-CMs, whereas quantitative polymerase chain reaction was used to assess SCN5A/SCN10A expression levels in LAA tissue. Results: In paroxysmal and persistent AF LAA-CMs, AP duration was shorter than in SR LAA-CMs. Compared with SR, peak INa and SCN5A expression were significantly decreased in paroxysmal AF, whereas they were restored to SR levels in persistent AF. Conversely, although late INa was unchanged in paroxysmal AF compared with SR, it was significantly increased in persistent AF. Peak or late Nav1.8-based INa was not detected in persistent AF LAA-CMs. Similarly, expression of SCN10A was not observed in LAAs at any stage. Conclusions: Our findings demonstrate differences in (late) INa remodeling in LAA-CMs from patients with paroxysmal vs persistent AF, indicating distinct cellular proarrhythmic mechanisms in different AF forms. These observations are of particular relevance when considering potential pharmacologic approaches targeting (late) INa in AF.

Contains fulltext : 291524.pdf (Publisher’s version ) (Open Access) For intradermal (ID) immunisation, novel needle-based delivery systems have been proposed as a better alternative to the Mantoux method. However, the penetration depth of needles in the human skin and its effect on immune cells residing in the different layers of the skin has not been analyzed. A novel and user-friendly silicon microinjection needle (Bella-mu(TM)) has been developed, which allows for a perpendicular injection due to its short needle length (1.4-1.8 mm) and ultrashort bevel. We aimed to characterize the performance of this microinjection needle in the context of the delivery of a particle-based outer membrane vesicle (OMV) vaccine using an ex vivo human skin explant model. We compared the needles of 1.4 and 1.8 mm with the conventional Mantoux method to investigate the depth of vaccine injection and the capacity of the skin antigen-presenting cell (APC) to phagocytose the OMVs. The 1.4 mm needle deposited the antigen closer to the epidermis than the 1.8 mm needle or the Mantoux method. Consequently, activation of epidermal Langerhans cells was significantly higher as determined by dendrite shortening. We found that five different subsets of dermal APCs are able to phagocytose the OMV vaccine, irrespective of the device or injection method. ID delivery using the 1.4 mm needle of a OMV-based vaccine allowed epidermal and dermal APC targeting, with superior activation of Langerhans cells. This study indicates that the use of a microinjection needle improves the delivery of vaccines in the human skin.

Endothelial cells (ECs) form a precisely regulated polarized monolayer in capillary walls. Vascular endothelial growth factor-A (VEGF-A) induces endothelial hyperpermeability, and VEGF-A applied to the basolateral side, but not the apical side, has been shown to be a strong barrier disruptor in blood–retinal barrier ECs. We show here that VEGF-A presented to the basolateral side of human umbilical vein ECs (HUVECs) induces higher permeability than apical stimulation, which is similar to results obtained with bovine retinal ECs. We investigated with immunocytochemistry and confocal imaging the distribution of VEGF receptor-2 (VEGFR2) and neuropilin-2 (NRP2) in perinuclear apical and basolateral membrane domains. Orthogonal z-sections of cultured HUVECs were obtained, and the fluorescence intensity at the apical and basolateral membrane compartments was measured. We found that VEGFR2 and NRP2 are evenly distributed throughout perinuclear apical and basolateral membrane compartments in unstimulated HUVECs grown on Transwell inserts, whereas basolateral VEGF-A stimulation induces a shift toward basolateral VEGFR2 and NRP2 localization. When HUVECs were grown on coverslips, the distribution of VEGFR2 and NRP2 across the perinuclear apical and basolateral membrane domains was different. Our findings demonstrate that HUVECs dynamically regulate VEGFR2 and NRP2 localization on membrane microdomains, depending on growth conditions and the polarity of VEGF-A stimulation.

The specification and growth of organs is controlled simultaneously by networks of transcription factors. While the connection between these transcription factors with fate determinants is increasingly clear, how they establish the link with the cell cycle is far less understood. Here we investigate this link in the developing Drosophila eye, where two transcription factors, the MEIS1 homologue hth and the Zn-finger tsh, synergize to stimulate the proliferation of naïve eye progenitors. Experiments combining transcriptomics, open-chromatin profiling, motif analysis and functional assays indicate that these progenitor transcription factors exert a global regulation of the proliferation program. Rather than directly regulating cell cycle genes, they control proliferation through an intermediary layer of nuclear receptors of the ecdysone/estrogen-signaling pathway. This regulatory subnetwork between hth, tsh and nuclear receptors might be conserved from Drosophila to mammals, as we find a significant co-overexpression of their human homologues in specific cancer types., PSP is a recipient of a Portuguese “Investigator FCT” contract. Grants BFU2012-34324 and BFU2015-66040 (MINECO, Spain) to F.C. Grants from Research Foundation Flanders (FWO, grants G. 0640.13 and G. 0791.14), University of Leuven (OT/13/103) to SA. MNS was funded by a PhD fellowship from FWO.

Aims The adult mammalian heart is a post-mitotic organ. Even in response to necrotic injuries, where regeneration would be essential to reinstate cardiac structure and function, only a minor percentage of cardiomyocytes undergo cytokinesis. The gene programme that promotes cell division within this population of cardiomyocytes is not fully understood. In this study, we aimed to determine the gene expression profile of proliferating adult cardiomyocytes in the mammalian heart after myocardial ischaemia, to identify factors to can promote cardiac regeneration. Methods and results Here, we demonstrate increased 5-ethynyl-2’deoxyuridine incorporation in cardiomyocytes 3 days post-myocardial infarction in mice. By applying multi-colour lineage tracing, we show that this is paralleled by clonal expansion of cardiomyocytes in the borderzone of the infarcted tissue. Bioinformatic analysis of single-cell RNA sequencing data from cardiomyocytes at 3 days post ischaemic injury revealed a distinct transcriptional profile in cardiomyocytes expressing cell cycle markers. Combinatorial overexpression of the enriched genes within this population in neonatal rat cardiomyocytes and mice at postnatal day 12 (P12) unveiled key genes that promoted increased cardiomyocyte proliferation. Therapeutic delivery of these gene cocktails into the myocardial wall after ischaemic injury demonstrated that a combination of thymosin beta 4 (TMSB4) and prothymosin alpha (PTMA) provide a permissive environment for cardiomyocyte proliferation and thereby attenuated cardiac dysfunction. Conclusion This study reveals the transcriptional profile of proliferating cardiomyocytes in the ischaemic heart and shows that overexpression of the two identified factors, TMSB4 and PTMA, can promote cardiac regeneration. This work indicates that in addition to activating cardiomyocyte proliferation, a supportive environment is a key for regeneration to occur.

Background: Epicardial adipose tissue (EAT) accumulation is associated with cardiac arrhythmias. The effect of EAT secretome (EATs) on cardiac electrophysiology remains largely unknown. Objective: The purpose of this study was to investigate the arrhythmogenicity of EATs and its underlying molecular and electrophysiological mechanisms. Methods: We collected atrial EAT and subcutaneous adipose tissue (SAT) from 30 patients with atrial fibrillation (AF), and EAT from 3 donors without AF. The secretome was collected after a 24-hour incubation of the adipose tissue explants. We cultured neonatal rat ventricular myocytes (NRVMs) with EATs, subcutaneous adipose tissue secretome (SATs), and cardiomyocytes conditioned medium (CCM) for 72 hours. We implemented the electrophysiological changes observed after EATs incubation into a model of human left atrium and tested arrhythmia inducibility. Results: Incubation of NRVMs with EATs decreased expression of the potassium channel subunit Kcnj2 by 26% and correspondingly reduced the inward rectifier K+ current IK1 by 35% compared to incubation with CCM, resulting in a depolarized resting membrane of cardiomyocytes. EATs decreased expression of connexin43 (29% mRNA, 46% protein) in comparison to CCM. Cells incubated with SATs showed no significant differences in Kcnj2 or Gja1 expression in comparison to CCM, and their resting potential was not depolarized. Cardiomyocytes incubated with EATs showed reduced conduction velocity and increased conduction heterogeneity compared to SATs and CCM. Computer modeling of human left atrium revealed that the electrophysiological changes induced by EATs promote sustained reentrant arrhythmias if EAT partially covers the myocardium. Conclusion: EAT slows conduction, depolarizes the resting potential, alters electrical cell–cell coupling, and facilitates reentrant arrhythmias.

Contains fulltext : 287837.pdf (Publisher’s version ) (Open Access) The Medical School of Padua (Italy) contributed profoundly to the study of teratology. Many famous physicians and professors of medicine, such as Liceti, Vallisneri, Morgagni, and Malacarne, have studied and investigated these anomalies to better understand the causes and to find a potential explanation, often preserving the specimens for future studies. The present study highlights some historical cases of conjoined twins and a conjoined triplet preserved at the Morgagni Museum of Human Anatomy to show the development of medical theories in the teratological field between the 18th and early 19th century. This approach will provide insights into different study methods and ideas of some of the most famous scholars working in Padua at that time. The current article focuses on rare cases, both human and animal, that were encountered by physicians who worked in the Veneto area in the late 18th and early 19th century. Their detailed descriptions are not only of historical but also of contemporary dysmorphological value.

Abstract To investigate the differences between pre- and post-fire fractures, 30 human forearm bones were subjected to either blunt-force impact, burning, or both. Bones, covered in soft tissue and wrapped in clothing, were burned in a reconstructed house fire. The burning context and dynamics led to differential burning, that was equal amongst the three groups. To evaluate the damage to the bones, a data collection sheet was developed based on the current literature on fracture features. To analyze the relation between exposure temperature and fracture class and occurrence, color was measured to estimate the exposure temperature. Observable and measurable changes on bone, and fracture surfaces, were macro- and microscopically analyzed. Many features overlapped between the three groups, and thus are not usable for differentiation. Fractures caused by blunt force impact (post-mortem, pre-fire) showed a rough fracture surface with smoothness in curved/slanted regions near the margin after burning, while heat-induced bone fractures showed a smooth fracture surface. The margins and surface of bone fractures that originated after the fire (indirect heat-induced) were evenly discolored, whereas heat-induced bone fractures showed uneven discoloration of the fracture margin and surface. Although there were generally more heat-induced fractures in the 450–700 °C range, no other distinctive trend was observed between exposure temperature and class of fractures.

Background: Atherosclerotic plaque ruptures, triggered by blood flow–associated biomechanical forces, cause most myocardial infarctions and strokes. Objectives: This study aims to investigate the exact location and underlying mechanisms of atherosclerotic plaque ruptures, identifying therapeutic targets against cardiovascular events. Methods: Histology, electron microscopy, bulk and spatial RNA sequencing on human carotid plaques were studied in proximal, most stenotic, and distal regions along the longitudinal blood flow direction. Genome-wide association studies were used to examine heritability enrichment and causal relationships of atherosclerosis and stroke. Associations between top differentially expressed genes (DEGs) and preoperative and postoperative cardiovascular events were examined in a validation cohort. Results: In human carotid atherosclerotic plaques, ruptures predominantly occurred in the proximal and most stenotic regions but not in the distal region. Histologic and electron microscopic examination showed that proximal and most stenotic regions exhibited features of plaque vulnerability and thrombosis. RNA sequencing identified DEGs distinguishing the proximal and most stenotic regions from the distal region which were deemed as most relevant to atherosclerosis-associated diseases as shown by heritability enrichment analyses. The identified pathways associated with the proximal rupture-prone regions were validated by spatial transcriptomics, firstly in human atherosclerosis. Of the 3 top DEGs, matrix metallopeptidase 9 emerged particularly because Mendelian randomization suggested that its high circulating levels were causally associated with atherosclerosis risk. Conclusions: Our findings show plaque site–specific transcriptional signatures associated with proximal rupture-prone regions of carotid atherosclerotic plaques. This led to the geographical mapping of novel therapeutic targets, such as matrix metallopeptidase 9, against plaque rupture.

The transmembrane protein 43 (TMEM43/LUMA) p.S358L mutation causes arrhythmogenic cardiomyopathy named as ARVC5, a fully penetrant disease with high risk of ventricular arrhythmias, sudden death, and heart failure. Male gender and vigorous exercise independently predicted deleterious outcome. Our systems genetics analysis revealed the importance of Tmem43 for cardiac and metabolic pathways associated with elevated lipid absorption from small intestine. This study sought to delineate gender-specific cardiac, intestinal, and metabolic phenotypes in vivo and investigate underlying pathophysiological mechanisms of S358L mutation. Serial echocardiography, surface electrocardiography (ECG), treadmill running, and body EchoMRI have been used in knock-in heterozygous (Tmem43WT/S358L), homozygous (Tmem43S358L), and wildtype (Tmem43WT) littermate mice. Electron microscopy, histology, immunohistochemistry, transcriptome, and protein analysis have been performed in cardiac and intestinal tissues. Systolic dysfunction was apparent in 3-mo-old Tmem43S358L and 6-mo-old Tmem43WT/S358L mutants. Both mutant lines displayed intolerance to acute stress at 6 mo of age, arrhythmias, fibro-fatty infiltration, and subcellular abnormalities in the myocardium. Microarray analysis found significantly differentially expressed genes between left ventricular (LV) and right ventricular (RV) myocardium. Mutants displayed diminished PPARG activities and significantly reduced TMEM43 and β-catenin expression in the heart, whereas junctional plakoglobin (JUP) translocated into nuclei of mutant cardiomyocytes. Conversely, elongated villi, fatty infiltration, and overexpression of gut epithelial proliferation markers, β-catenin and Ki-67, were evident in small intestine of mutants. We defined Tmem43 S358L-induced pathological effects on cardiac and intestinal homeostasis via distinctly disturbed WNT-β-catenin and PPARG signaling thereby contributing to ARVC5 pathophysiology. Results suggest that cardiometabolic assessment in mutation carriers may be important for predictive and personalized care.NEW & NOTEWORTHY This manuscript describes the findings of our investigation of cardiac, small intestine, and metabolic features of Tmem43-S358L mouse model. By investigating interorgan pathologies, we uncovered multiple mechanisms of the S358L-induced disease, and these unique mechanisms likely appear to contribute to the disease pathogenesis. We hope our findings are important and novel and open new avenues in the hunting for additional diagnostic and therapeutic targets in subjects carrying TMEM43 mutation.

Aims In the heart, splicing factors orchestrate the functional properties of cardiomyocytes by regulating the alternative splicing of multiple genes. Work in embryonic stem cells has shown that the splicing factor Quaking (QKI) regulates alternative splicing during cardiomyocyte differentiation. However, the relevance and function of QKI in adult cardiomyocytes remains unknown. In this study, we aim to identify the in vivo function of QKI in the adult mouse heart. Methods and results We generated mice with conditional deletion of QKI in cardiomyocytes by the Cre-Lox system. Mice with cardiomyocyte-specific deletion of QKI died during the foetal period (E14.5), without obvious anatomical abnormalities of the heart. Adult mice with tamoxifen-inducible QKI deletion rapidly developed heart failure associated with severe disruption of sarcomeres, already 7 days after knocking out QKI. RNA sequencing revealed that QKI regulates the alternative splicing of more than 1000 genes, including sarcomere and cytoskeletal components, calcium-handling genes, and (post-)transcriptional regulators. Many of these splicing changes corresponded to the loss of muscle-specific isoforms in the heart. Forced overexpression of QKI in cultured neonatal rat ventricular myocytes directed these splicing events in the opposite direction and enhanced contractility of cardiomyocytes. Conclusion Altogether, our findings show that QKI is an important regulator of the muscle-specific alternative splicing program that builds the contractile apparatus of cardiomyocytes.

Trinajstic et al., ( Science , 16 September 2022, p. 1311–1314) describe exceptionally well-preserved organs in fossilized Devonian placoderms to infer the early evolution of the vertebrate heart. We argue that the report has numerous shortcomings and examples of mixed specimen codes. Further, we question whether there indeed is any evidence for a mineralized chambered heart in these placoderms.

Targeting germline (gl-) precursors of broadly neutralizing antibodies (bNAbs) is acknowledged as an important strategy for HIV-1 vaccines. The VRC01-class of bNAbs is attractive because of its distinct genetic signature. However, VRC01-class bNAbs often require extensive somatic hypermutation, including rare insertions and deletions. We describe a BG505 SOSIP trimer, termed GT1.2, to optimize binding to gl-CH31, the unmutated common precursor of the CH30-34 bNAb lineage that acquired a large CDRH1 insertion. The GT1.2 trimer activates gl-CH31 naive B cells in knock-in mice, and B cell responses could be matured by selected boosting immunogens to generate cross-reactive Ab responses. Next-generation B cell sequencing reveals selection for VRC01-class mutations, including insertions in CDRH1 and FWR3 at positions identical to VRC01-class bNAbs, as well as CDRL1 deletions and/or glycine substitutions to accommodate the N276 glycan. These results provide proof of concept for vaccine-induced affinity maturation of B cell lineages that require rare insertions and deletions.

Objectives: Adult spleens show extensive morphological variation, with a reported prevalence of 40–98% clefts (also called notches or fissures) on the splenic surface and 10–30% accessory spleens at autopsy. It is hypothesised that both anatomical variants result from a complete or partial failure of multiple splenic primordia to fuse to the main body. According to this hypothesis, fusion of the spleen primordia is completed after birth and spleen morphological variations are often explained as stagnation of spleen development at the foetal stage. We tested this hypothesis by studying early spleen development in embryos, and compared foetal and adult spleen morphology. Methods and materials: We assessed 22 embryonic, 17 foetal and 90 adult spleens on the presence of clefts using histology, micro-CT and conventional post-mortem CT-scans, respectively. Results: The spleen primordium was observed as a single mesenchymal condensation in all embryonic specimens. The number of clefts varied from 0 to 6 in foetuses, compared to 0–5 in adults. We found no correlation between foetal age and number of clefts (R2 = 0.004). The independent samples Kolmogorov–Smirnov test showed no significant difference in the total number of clefts between adult and foetal spleens (p = 0.068). Conclusion: We found no morphological evidence for a multifocal origin or a lobulated developmental stage of the human spleen. Advances in knowledge: Our findings show that splenic morphology is highly variable, independent of developmental stage and age. We suggest to abandon the term “persistent foetal lobulation” and to regard splenic clefts, regardless of their number or location, as normal variants.

The pacemaker activity of the sinoatrial node (SAN) has been studied extensively in animal species but is virtually unexplored in humans. Here we assess the role of the slowly activating component of the delayed rectifier K+ current (IKs) in human SAN pacemaker activity and its dependence on heart rate and β-adrenergic stimulation. HEK-293 cells were transiently transfected with wild-type KCNQ1 and KCNE1 cDNA, encoding the α- and β-subunits of the IKs channel, respectively. KCNQ1/KCNE1 currents were recorded both during a traditional voltage clamp and during an action potential (AP) clamp with human SAN-like APs. Forskolin (10 µmol/L) was used to increase the intracellular cAMP level, thus mimicking β-adrenergic stimulation. The experimentally observed effects were evaluated in the Fabbri–Severi computer model of an isolated human SAN cell. Transfected HEK-293 cells displayed large IKs-like outward currents in response to depolarizing voltage clamp steps. Forskolin significantly increased the current density and significantly shifted the half-maximal activation voltage towards more negative potentials. Furthermore, forskolin significantly accelerated activation without affecting the rate of deactivation. During an AP clamp, the KCNQ1/KCNE1 current was substantial during the AP phase, but relatively small during diastolic depolarization. In the presence of forskolin, the KCNQ1/KCNE1 current during both the AP phase and diastolic depolarization increased, resulting in a clearly active KCNQ1/KCNE1 current during diastolic depolarization, particularly at shorter cycle lengths. Computer simulations demonstrated that IKs reduces the intrinsic beating rate through its slowing effect on diastolic depolarization at all levels of autonomic tone and that gain-of-function mutations in KCNQ1 may exert a marked bradycardic effect during vagal tone. In conclusion, IKs is active during human SAN pacemaker activity and has a strong dependence on heart rate and cAMP level, with a prominent role at all levels of autonomic tone.

Non-crocodylian reptiles have hearts with a single ventricle, which is partially separated by a muscular ridge that provide some separation of blood flows. An exceptional situation exists in monitor lizards and pythons, where the ventricular left side generates a much higher systolic blood pressure than the right side, thus resembling mammals and birds. This functional division of the ventricle depends on a large muscular ridge and may relate to high metabolic demand. The large leatherback turtle (

Dendritic cells (DCs) control adaptive immunity and are therefore attractive for in vivo targeting to either induce immune activation or tolerance, depending on disease. Liposomes, nanoparticles comprised of a lipid bi-layer, provide a nanoplatform for loading disease-relevant antigen, adjuvant and DC-targeting molecules simultaneously. However, it is yet not fully understood how liposomal formulations affect uptake by DCs and DC function. Here, we examined monocyte-derived DC (moDC) and skin DC uptake of six different liposomal formulations, together with their DC-modulating effect. Contrary to literature, we show using imaging flow cytometry that anionic or neutral liposomes are taken up more efficiently than cationic liposomes by moDCs, or by skin DCs after intradermal injection. None of the formulations yielded significant modulation of DC function as determined by the upregulation of maturation markers and cytokine production. These results suggest that anionic liposomes would be more suitable as vaccine carriers for a dermal application.(c) 2022 The Authors. Published by Elsevier Inc. on behalf of American Pharmacists Association. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Background: Variants in KCNH2, encoding the human ether a-go-go (hERG) channel that is responsible for the rapid component of the cardiac delayed rectifier K+ current (IKr), are causal to long QT syndrome type 2 (LQTS2). We identified eight index patients with a new variant of unknown significance (VUS), KCNH2:c.2717C > T:p.(Ser906Leu). We aimed to elucidate the biophysiological effect of this variant, to enable reclassification and consequent clinical decision-making. Methods: A genotype–phenotype overview of the patients and relatives was created. The biophysiological effects were assessed independently by manual-, and automated calibrated patch clamp. HEK293a cells expressing (i) wild-type (WT) KCNH2, (ii) KCNH2-p.S906L alone (homozygous, Hm) or (iii) KCNH2-p.S906L in combination with WT (1:1) (heterozygous, Hz) were used for manual patching. Automated patch clamp measured the variants function against known benign and pathogenic variants, using Flp-In T-rex HEK293 KCNH2-variant cell lines. Results: Incomplete penetrance of LQTS2 in KCNH2:p.(Ser906Leu) carriers was observed. In addition, some patients were heterozygous for other VUSs in CACNA1C, PKP2, RYR2 or AKAP9. The phenotype of carriers of KCNH2:p.(Ser906Leu) ranged from asymptomatic to life-threatening arrhythmic events. Manual patch clamp showed a reduced current density by 69.8 and 60.4% in KCNH2-p.S906L-Hm and KCNH2-p.S906L-Hz, respectively. The time constant of activation was significantly increased with 80.1% in KCNH2-p.S906L-Hm compared with KCNH2-WT. Assessment of KCNH2-p.S906L-Hz by calibrated automatic patch clamp assay showed a reduction in current density by 35.6%. Conclusion: The reduced current density in the KCNH2-p.S906L-Hz indicates a moderate loss-of-function. Combined with the reduced penetrance and variable phenotype, we conclude that KCNH2:p.(Ser906Leu) is a low penetrant likely pathogenic variant for LQTS2.

Contains fulltext : 291515.pdf (Publisher’s version ) (Open Access) Birt-Hogg-Dubé syndrome (BHD) is an autosomal dominant disorder characterized by fibrofolliculomas, pulmonary cysts, pneumothoraces and renal cell carcinomas. Here, we reveal a novel hereditary disorder in a family with skin and mucosal lesions, extensive lipomatosis and renal cell carcinomas. The proband was initially diagnosed with BHD based on the presence of fibrofolliculomas, but no pathogenic germline variant was detected in FLCN, the gene associated with BHD. By whole exome sequencing we identified a heterozygous missense variant (p.(Cys677Tyr)) in a zinc-finger encoding domain of the PRDM10 gene which co-segregated with the phenotype in the family. We show that PRDM10Cys677Tyr loses affinity for a regulatory binding motif in the FLCN promoter, abrogating cellular FLCN mRNA and protein levels. Overexpressing inducible PRDM10Cys677Tyr in renal epithelial cells altered the transcription of multiple genes, showing overlap but also differences with the effects of knocking out FLCN. We propose that PRDM10 controls an extensive gene program and acts as a critical regulator of FLCN gene transcription in human cells. The germline variant PRDM10Cys677Tyr curtails cellular folliculin expression and underlies a distinguishable syndrome characterized by extensive lipomatosis, fibrofolliculomas and renal cell carcinomas.

Induction of lipid-laden foamy macrophages is a cellular hallmark of tuberculosis (TB) disease, which involves the transformation of infected phagolysosomes from a site of killing into a nutrient-rich replicative niche. Here, we show that a terpenyl nucleoside shed from Mycobacterium tuberculosis, 1-tuberculosinyladenosine (1-TbAd), caused lysosomal maturation arrest and autophagy blockade, leading to lipid storage in M1 macrophages. Pure 1-TbAd, or infection with terpenyl nucleoside-producing M. tuberculosis, caused intralysosomal and peribacillary lipid storage patterns that matched both the molecules and subcellular locations known in foamy macrophages. Lipidomics showed that 1-TbAd induced storage of triacylglycerides and cholesterylesters and that 1-TbAd increased M. tuberculosis growth under conditions of restricted lipid access in macrophages. Furthermore, lipidomics identified 1-TbAd-induced lipid substrates that define Gaucher's disease, Wolman's disease, and other inborn lysosomal storage diseases. These data identify genetic and molecular causes of M. tuberculosis-induced lysosomal failure, leading to successful testing of an agonist of TRPML1 calcium channels that reverses lipid storage in cells. These data establish the host-directed cellular functions of an orphan effector molecule that promotes survival in macrophages, providing both an upstream cause and detailed picture of lysosome failure in foamy macrophages.

Introduction Patients with ventricular tachyarrhythmias (VT) are at high risk of sudden cardiac death. When appropriate, catheter ablation is modestly effective, with relatively high VT recurrence and complication rates. Personalized models that incorporate imaging and computational approaches have advanced VT management. However, 3D patient-specific functional electrical information is typically not considered. We hypothesize that incorporating non-invasive 3D electrical and structural characterization in a patient-specific model improves VT-substrate recognition and ablation targeting.Materials and methods In a 53-year-old male with ischemic cardiomyopathy and recurrent monomorphic VT, we built a structural-functional model based on high-resolution 3D late-gadolinium enhancement (LGE) cardiac magnetic resonance imaging (3D-LGE CMR), multi-detector computed tomography (CT), and electrocardiographic imaging (ECGI). Invasive data from high-density contact and pace mapping obtained during endocardial VT-substrate modification were also incorporated. The integrated 3D electro-anatomic model was analyzed off-line.Results Merging the invasive voltage maps and 3D-LGE CMR endocardial geometry led to a mean Euclidean node-to-node distance of 5 & PLUSMN; 2 mm. Inferolateral and apical areas of low bipolar voltage (< 1.5 mV) were associated with high 3D-LGE CMR signal intensity (> 0.4) and with higher transmurality of fibrosis. Areas of functional conduction delay or block (evoked delayed potentials, EDPs) were in close proximity to 3D-LGE CMR-derived heterogeneous tissue corridors. ECGI pinpointed the epicardial VT exit at & SIM;10 mm from the endocardial site of origin, both juxtaposed to the distal ends of two heterogeneous tissue corridors in the inferobasal left ventricle. Radiofrequency ablation at the entrances of these corridors, eliminating all EDPs, and at the VT site of origin rendered the patient non-inducible and arrhythmia-free until the present day (20 months follow-up). Off-line analysis in our model uncovered dynamic electrical instability of the LV inferolateral heterogeneous scar region which set the stage for an evolving VT circuit.Discussion and conclusion We developed a personalized 3D model that integrates high-resolution structural and electrical information and allows the investigation of their dynamic interaction during arrhythmia formation. This model enhances our mechanistic understanding of scar-related VT and provides an advanced, non-invasive roadmap for catheter ablation. This study was supported by the Special Research Fund (BOF) of Hasselt University (BOF17DOCMA15) and the Maastricht University Medical Center (MUMC+) to JS, the Hein Wellens Foundation, Health Foundation Limburg (Maastricht, The Netherlands), and a Veni grant from the Netherlands Organization for Scientific Research (TTW16772) to MC, the Netherlands CardioVascular Research Initiative (CVON2017-13 VIGILANCE and CVON2018B030 PREDICT2), Den Haag, The Netherlands to PV, and a Veni grant from the Netherlands Organization for Scientific Research (NWO/ZonMw 0915016181013) to RTB.

Context: Applicant perceptions of selection methods can affect motivation, performance and withdrawal and may therefore be of relevance in the context of widening access. However, it is unknown how applicant subgroups perceive different selection methods. Objectives: Using organisational justice theory, the present multi-site study examined applicant perceptions of various selection methods, rationales behind perceptions and subgroup differences. Methods: Applicants to five Dutch undergraduate health professions programmes (N = 704) completed an online survey including demographics and a questionnaire on applicant perceptions applied to 11 commonly used selection methods. Applicants rated general favourability and justice dimensions (7-point Likert scale) and could add comments for each method. Results: Descriptive statistics revealed a preference for selection methods on which applicants feel more ‘in control’: General favourability ratings were highest for curriculum-sampling tests (mean [M] = 5.32) and skills tests (M = 5.13), while weighted lottery (M = 3.05) and unweighted lottery (M = 2.97) were perceived least favourable. Additionally, applicants preferred to distinguish themselves on methods that assess attributes beyond cognitive abilities. Qualitative content analysis of comments revealed several conflicting preferences, including a desire for multiple selection methods versus concerns of experiencing too much stress. Results from a linear mixed model of general favourability indicated some small subgroup differences in perceptions (based on gender, migration background, prior education and parental education), but practical meaning of these differences was negligible. Nevertheless, concerns were expressed that certain selection methods can hinder equitable admission due to inequal access to resources. Conclusions: Our findings illustrate that applicants desire to demonstrate a variety of attributes on a combination of selection tools, but also observe that this can result in multiple drawbacks. The present study can help programmes in deciding which selection methods to include, which more negatively perceived methods should be better justified to applicants, and how to adapt methods to meet applicants' needs.

Heart development and rhythm control are highly Tbx5 dosage-sensitive. TBX5 haploinsufficiency causes congenital conduction disorders, whereas increased expression levels of TBX5 in human heart samples has been associated with atrial fibrillation (AF). We deleted the conserved mouse orthologues of two independent AF-associated genomic regions in the Tbx5 locus, one intronic (RE(int)) and one downstream (RE(down)) of Tbx5. In both lines, we observed a modest (30%) increase of Tbx5 in the postnatal atria. To gain insight into the effects of slight dosage increase in vivo, we investigated the atrial transcriptional, epigenetic and electrophysiological properties of both lines. Increased atrial Tbx5 expression was associated with induction of genes involved in development, ion transport and conduction, with increased susceptibility to atrial arrhythmias, and increased action potential duration of atrial cardiomyocytes. We identified an AF-associated variant in the human RE(int) that increases its transcriptional activity. Expression of the AF-associated transcription factor Prrx1 was induced in Tbx5RE(int)KO cardiomyocytes. We found that some of the transcriptional and functional changes in the atria caused by increased Tbx5 expression were normalized when reducing cardiac Prrx1 expression in Tbx5RE(int)KO mice, indicating an interaction between these two AF genes. We conclude that modest increases in expression of dose-dependent transcription factors, caused by common regulatory variants, significantly impact on the cardiac gene regulatory network and disease susceptibility.

Introduction: High pacing frequency or irregular activity due to arrhythmia produces complex optical mapping signals and challenges for processing. The objective is to establish an automated activation time-based analytical framework applicable to optical mapping images of complex electrical behavior.Methods: Optical mapping signals with varying complexity from sheep (N = 7) ventricular preparations were examined. Windows of activation centered on each action potential upstroke were derived using Hilbert transform phase. Upstroke morphology was evaluated for potential multiple activation components and peaks of upstroke signal derivatives defined activation time. Spatially and temporally clustered activation time points were grouped in to wave fronts for individual processing. Each activation time point was evaluated for corresponding repolarization times. Each wave front was subsequently classified based on repetitive or non-repetitive events. Wave fronts were evaluated for activation time minima defining sites of wave front origin. A visualization tool was further developed to probe dynamically the ensemble activation sequence.Results: Our framework facilitated activation time mapping during complex dynamic events including transitions to rotor-like reentry and ventricular fibrillation. We showed that using fixed AT windows to extract AT maps can impair interpretation of the activation sequence. However, the phase windowing of action potential upstrokes enabled accurate recapitulation of repetitive behavior, providing spatially coherent activation patterns. We further demonstrate that grouping the spatio-temporal distribution of AT points in to coherent wave fronts, facilitated interpretation of isolated conduction events, such as conduction slowing, and to derive dynamic changes in repolarization properties. Focal origins precisely detected sites of stimulation origin and breakthrough for individual wave fronts. Furthermore, a visualization tool to dynamically probe activation time windows during reentry revealed a critical single static line of conduction slowing associated with the rotation core.Conclusion: This comprehensive analytical framework enables detailed quantitative assessment and visualization of complex electrical behavior.

A considerable amount of literature has been published on antidepressants and cardiac ion channel dysfunction. The antidepressant paroxetine has been associated with Brugada syndrome and long QT syndrome, albeit on the basis of conflicting findings. The cardiac voltage-gated sodium channel (NaV1.5) is related to both of these syndromes, suggesting that paroxetine may have an effect on this channel. In the present study, we therefore carried out patch clamp experiments to examine the effect of paroxetine on human NaV1.5 channels stably expressed in human embryonic kidney 293 (HEK-293) cells as well as on action potentials of isolated rabbit left ventricular cardiomyocytes. Additionally, computer simulations were conducted to test the functional effects of the experimentally observed paroxetine-induced changes in the NaV1.5 current. We found that paroxetine led to a decrease in peak NaV1.5 current in a concentration-dependent manner with an IC50 of 6.8 ± 1.1 µM. In addition, paroxetine caused a significant hyperpolarizing shift in the steady-state inactivation of the NaV1.5 current as well as a significant increase in its rate of inactivation. Paroxetine (3 µM) affected the action potential of the left ventricular cardiomyocytes, significantly decreasing its maximum upstroke velocity and amplitude, both of which are mainly regulated by the NaV1.5 current. Our computer simulations demonstrated that paroxetine substantially reduces the fast sodium current of human left ventricular cardiomyocytes, thereby slowing conduction and reducing excitability in strands of cells, in particular if conduction and excitability are already inhibited by a loss-of-function mutation in the NaV1.5 encoding SCN5A gene. In conclusion, paroxetine acts as an inhibitor of NaV1.5 channels, which may enhance the effects of loss-of-function mutations in SCN5A.

Fanconi anaemia (FA) is a rare chromosomal-instability syndrome caused by mutations of any of the 22 known FA DNA-repair genes. FA individuals have an increased risk of head-and-neck squamous-cell-carcinomas (HNSCC), often fatal. Systemic intolerance to standard cisplatin-based protocols due to somatic-cell hypersensitivity underscores the urgent need to develop novel therapies. Here, we performed unbiased siRNA screens to unveil genetic interactions synthetic-lethal with FA-pathway deficiency in FA-patient HNSCC cell lines. We identified based on differential-lethality scores between FA-deficient and FA-proficient cells, next to common-essential genes such as PSMC1, PSMB2, and LAMTOR2, the otherwise non-essential RBBP9 gene. Accordingly, low dose of the FDA-approved RBBP9-targeting drug Emetine kills FA-HNSCC. Importantly both RBBP9-silencing as well as Emetine spared non-tumour FA cells. This study provides a minable genome-wide analyses of vulnerabilities to address treatment challenges in FA-HNSCC. Our investigation divulges a DNA-cross-link-repair independent lead, RBBP9, for targeted treatment of FA-HNSCCs without systemic toxicity.

The mechanical forces that the cells of an organism are subject to are an essential epigenetic factor impacting on their behavior. Heart development is a striking example of a process greatly influenced by mechanical forces. The heart develops from a simple tube in the early embryo into a structurally complex chambered heart with valves and great vessels. Already at the early stages of development, the heart tube starts to function and thus to produce mechanical forces generated by contractions and blood flow. As a consequence, the cells of the developing heart are continuously subject to dynamic shear stress, pressure, and stretch, which change as the developing heart grows and structurally transforms. Through mechanotransduction, the mechanical signals are converted to chemical or electrical signals and transcriptional responses in the cardiac cells that in turn steer the process of heart development. Here we discuss relatively well-studied examples of heart developmental processes that rely on transduction of the dynamic mechanical signals produced by heart function, heart looping and chamber expansion, the formation of the trabeculae, the formation of the valves, and the formation of the outflow vessels.

Background: Symmetric craniorachipagus is an exceedingly rare type of bi-umbilical conjoined twinning, known from only three scantily described cases. Case: We identified a fourth, previously described case that was misdiagnosed as janiceps and as pygopagus. It concerned dorsally conjoined twins that were part of a triplet pregnancy, spontaneously born at 22 weeks of gestation. Radiography confirmed union at the occipital craniums and the thoracolumbar vertebral columns. Both twins had their own separate umbilical cords. To delineate the phenotype of craniorachipagus and to differentiate it from rachipagus without cranial involvement, we compared the present case with the previous three reported cases and with the historical literature concerning comparable conditions. Furthermore, we discuss why exceedingly rare conditions such as these are presently underreported in the literature. Conclusion: Symmetric craniorachipagus is a type of bi-umbilical conjoined twinning, to date represented by four confirmed cases that share a similar phenotype. This includes dorsal conjunction at the sides of the occipital craniums and the vertebral columns, in the absence of any visceral connections. Details on its etiopathogenesis and apparent lethality await additional case investigations. No unequivocally confirmed cases of symmetric rachipagus without cranial involvement have been reported and its existence in humans has yet to be proven.

Niche cells are widely known to regulate stem/progenitor cells in many mammalian tissues. In the hair, dermal papilla niche cells are well accepted to regulate hair stem/progenitor cells. However, how niche cells themselves are maintained is largely unknown. We present evidence implicating hair matrix progenitors and the lipid modifying enzyme, Stearoyl CoA Desaturase 1, in the regulation of the dermal papilla niche during the anagen-catagen transition of the mouse hair cycle. Our data suggest that this takes place via autocrine Wnt signalling and paracrine Hedgehog signalling. To our knowledge, this is the first report demonstrating a potential role for matrix progenitor cells in maintaining the dermal papilla niche. Agency for Science, Technology and Research (A*STAR) Ministry of Health (MOH) National Medical Research Council (NMRC) Published version This work was supported by Biomedical Research Council (BMRC), Agency for Science, Technology, and Research in Singapore (A*STAR); BMRC-A*STAR-EDB IAF-PP for the Skin Research Institute of Singapore (H17/01/a0/004), and Acne and Sebaceous Gland Program (H17/ H17/01/a0/008), and National Medical Research Council Clinician-Scientist Individual Research Grant MOH-CIRG20nov-0009.

Previously, we reported a series of families presenting with trichodiscomas, inherited in an autosomal dominant pattern. The phenotype was named familial multiple discoid fibromas (FMDF). The genetic cause of FMDF remained unknown so far. Trichodiscomas are skin lesions previously reported to be part of the same spectrum as the fibrofolliculoma observed in Birt-Hogg-Dubé syndrome (BHD), an inherited disease caused by pathogenic variants in the FLCN gene. Given the clinical and histological differences with BHD and the exclusion of linkage with the FLCN locus, the phenotype was concluded to be distinct from BHD. We performed extensive clinical evaluations and genetic testing in ten families with FMDF. We identified a FNIP1 frameshift variant in nine families and genealogical studies showed common ancestry for eight families. Using whole exome sequencing, we identified six additional rare variants in the haplotype surrounding FNIP1, including a missense variant in the PDGFRB gene that was found to be present in all tested patients with FMDF. Genome-wide linkage analysis showed that the locus on chromosome 5 including FNIP1 was the only region reaching the maximal possible LOD score. We concluded that FMDF is linked to a haplotype on chromosome 5. Additional evaluations in families with FMDF are required to unravel the exact genetic cause underlying the phenotype. When evaluating patients with multiple trichodisomas without a pathogenic variant in the FLCN gene, further genetic testing is warranted and can include analysis of the haplotype on chromosome 5.

Accuracy and transparency of scientific data are becoming more and more relevant with the increasing concern regarding the evaluation of data reproducibility in many research areas. This concern is also true for quantifying coding and noncoding RNAs, with the remarkable increase in publications reporting RNA profiling and sequencing studies. To address the problem, we propose the following recommendations: (a) accurate documentation of experimental procedures in Materials and methods (and not only in the supplementary information, as many journals have a strict mandate for making Materials and methods as visible as possible in the main text); (b) submission of RT-qPCR raw data for all experiments reported; and (c) adoption of a unified, simple format for submitted RT-qPCR raw data. The Real-time PCR Data Essential Spreadsheet Format (RDES) was created for this purpose.

Student diversity in health professions education (HPE) can be affected by selection procedures. Little is known about how different selection tools impact student diversity across programs using different combinations of traditional and broadened selection criteria. The present multi-site study examined the chances in selection of subgroups of applicants to HPE undergraduate programs with distinctive selection procedures, and their performance on corresponding selection tools. Probability of selection of subgroups (based on gender, migration background, prior education, parental education) of applicants (N = 1935) to five selection procedures of corresponding Dutch HPE undergraduate programs was estimated using multilevel logistic regression. Multilevel linear regression was used to analyze performance on four tools: prior-education grade point average (pe-GPA), biomedical knowledge test, curriculum-sampling test, and curriculum vitae (CV). First-generation Western immigrants and applicants with a foreign education background were significantly less likely to be selected than applicants without a migration background and with pre-university education. These effects did not vary across programs. More variability in effects was found between different selection tools. Compared to women, men performed significantly poorer on CVs, while they had higher scores on biomedical knowledge tests. Applicants with a non-Western migration background scored lower on curriculum-sampling tests. First-generation Western immigrants had lower CV-scores. First-generation university applicants had significantly lower pe-GPAs. There was a variety in effects for applicants with different alternative forms of prior education. For curriculum-sampling tests and CVs, effects varied across programs. Our findings highlight the need for continuous evaluation, identifying best practices within existing tools, and applying alternative tools.

Epigenetics; Tumour-suppressor proteins Epigenética; Proteínas supresoras de tumores Epigenètica; Proteïnes supresores de tumors Colorectal cancers (CRCs) often display histological features indicative of aberrant differentiation but the molecular underpinnings of this trait and whether it directly drives disease progression is unclear. Here, we identify co-ordinate epigenetic inactivation of two epithelial-specific transcription factors, EHF and CDX1, as a mechanism driving differentiation loss in CRCs. Re-expression of EHF and CDX1 in poorly-differentiated CRC cells induced extensive chromatin remodelling, transcriptional re-programming, and differentiation along the enterocytic lineage, leading to reduced growth and metastasis. Strikingly, EHF and CDX1 were also able to reprogramme non-colonic epithelial cells to express colonic differentiation markers. By contrast, inactivation of EHF and CDX1 in well-differentiated CRC cells triggered tumour de-differentiation. Mechanistically, we demonstrate that EHF physically interacts with CDX1 via its PNT domain, and that these transcription factors co-operatively drive transcription of the colonic differentiation marker, VIL1. Compound genetic deletion of Ehf and Cdx1 in the mouse colon disrupted normal colonic differentiation and significantly enhanced colorectal tumour progression. These findings thus reveal a novel mechanism driving epithelial de-differentiation and tumour progression in CRC. This project was supported by NHMRC project grant (1107831), a Cancer Council Victoria Grant (1164674) and the Operational Infrastructure Support Programme, Victorian Government, Australia. JMM was supported by a NHMRC Senior Research Fellowship (1046092). IYL was supported by F J Fletcher Research Scholarship and Randal and Louisa Alcock Scholarship from the University of Melbourne. LJJ was supported by La Trobe University Australian Postgraduate Awards. IN was supported by La Trobe University Postgraduate Research Scholarship. JWTT was supported by the University of Melbourne Australian Postgraduate Awards. OMS is a National Health and Medical Research Council (NHMRC) Senior Research Fellow (APP1136119). Open Access funding enabled and organized by CAUL and its Member Institutions.