Integrative data semantics through a model-enabled data stewardship.

MOTIVATION: The importance of clinical data in understanding the pathophysiology of complex disorders has prompted the launch of multiple initiatives designed to generate patient-level data from various modalities. While these studies can reveal important findings relevant to the disease, each study captures different yet complementary aspects and modalities which, when combined, generate a more comprehensive picture of disease etiology. However, achieving this requires a global integration of data across studies, which proves to be challenging given the lack of interoperability of cohort datasets. RESULTS: Here, we present the Data Steward Tool (DST), an application that allows for semi-automatic semantic integration of clinical data into ontologies and global data models and data standards. We demonstrate the applicability of the tool in the field of dementia research by establishing a Clinical Data Model (CDM) in this domain. The CDM currently consists of 277 common variables covering demographics (e.g. age and gender), diagnostics, neuropsychological tests and biomarker measurements. The DST combined with this disease-specific data model shows how interoperability between multiple, heterogeneous dementia datasets can be achieved. AVAILABILITY AND IMPLEMENTATION: The DST source code and Docker images are respectively available at https://github.com/SCAI-BIO/data-steward and https://hub.docker.com/r/phwegner/data-steward. Furthermore, the DST is hosted at https://data-steward.bio.scai.fraunhofer.de/data-steward. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SCAview: an Intuitive Visual Approach to the Integrative Analysis of Clinical Data in Spinocerebellar Ataxias.

With SCAview, we present a prompt and comprehensive tool that enables scientists to browse large datasets of the most common spinocerebellar ataxias intuitively and without technical effort. Basic concept is a visualization of data, with a graphical handling and filtering to select and define subgroups and their comparison. Several plot types to visualize all data points resulting from the selected attributes are provided. The underlying synthetic cohort is based on clinical data from five different European and US longitudinal multicenter cohorts in spinocerebellar ataxia type 1, 2, 3, and 6 (SCA1, 2, 3, and 6) comprising > 1400 patients with overall > 5500 visits. First, we developed a common data model to integrate the clinical, demographic, and characterizing data of each source cohort. Second, the available datasets from each cohort were mapped onto the data model. Third, we created a synthetic cohort based on the cleaned dataset. With SCAview, we demonstrate the feasibility of mapping cohort data from different sources onto a common data model. The resulting browser-based visualization tool with a thoroughly graphical handling of the data offers researchers the unique possibility to visualize relationships and distributions of clinical data, to define subgroups and to further investigate them without any technical effort. Access to SCAview can be requested via the Ataxia Global Initiative and is free of charge.

[Physical Strain after Abdominal Surgery - Results of a Patient Survey]. / Belastbarkeit nach abdominellen Eingriffen ­ Ergebnisse einer Patientenbefragung.

INTRODUCTION: Incision hernias are common complications after abdominal surgery and affect the recommendations on postoperative physical strain, as it is thought that excessively early strain causes incisional hernias. However, there is no evidence to justify this. This study evaluates the effect of postoperative strain on the risk of incisional hernia. MATERIALS AND METHODS: Patients with a laparoscopy (LS) or laparotomy (LT) were asked to complete a questionnaire on postoperative strain, complaints and quality of life. Patients with hernia surgery, or open abdomen therapy for complicated courses (Clavien-Dindo > III) were excluded. RESULTS: 393 patients completed the questionnaire (43.6%). 274 were LS and 128 LT. The incidence of incisional hernias was 5.2% (LS) and 18.0% (LT, p = 0.001). Incisional hernia patients were younger and more commonly males. 30.5% of incisional hernia patients did not return to normal physical strain postoperatively. Abdominal binders did not affect the hernia rate. The incisional hernia patients showed decreased quality of life scores in both mental and physical domains. CONCLUSION: Early postoperative physical strain was not a risk factor for incisional hernia development in this study. However, prospective studies are needed to create necessary evidence to recommend earlier postoperative return to normal physical strain.

Status quo of the use of DCS concepts and outcome with focus on blunt abdominal trauma : A registry-based analysis from the TraumaRegister DGU®.

INTRODUCTION: Damage control surgery (DCS) is a standardized treatment concept in severe abdominal injury. Despite its evident advantages, DCS bears the risk of substantial morbidity and mortality, due to open abdomen therapy (OAT). Thus, identifying the suitable patients for that approach is of utmost importance. Furthermore, little is known about the use of DCS and the related outcome, especially in blunt abdominal trauma. METHODS: Patients recorded in the TraumaRegister DGU® from 2008 to 2017, and with an Injury Severity Score (ISS) ≥ 9 and an abdominal injury with an Abbreviated Injury Scale (AIS) score ≥ 3 were included in that registry-based analysis. Patients with DCS and temporary abdominal closure (TAC) were compared with patients who were treated with a laparotomy and primary closure (non-DCS) and those who did receive non-operative management (NOM). Following descriptive analysis, a matched-pairs study was conducted to evaluate differences and outcomes between DCS and non-DCS group. Matching criteria were age, abdominal trauma severity, and hemodynamical instability at the scene. RESULTS: The injury mechanism was predominantly blunt (87.1%). Of the 8226 patients included, 2351 received NOM, 5011 underwent laparotomy and primary abdominal closure (non-DCS), and 864 were managed with DCS. Thus, 785 patient pairs were analysed. The rate of hepatic injuries AIS > 3 differed between the groups (DCS 50.3% vs. non-DCS 18.1%). DCS patients had a higher ISS (p = 0.023), required more significant volumes of fluids, more catecholamines, and transfusions (p < 0.001). More DCS patients were in shock at the accident scene (p = 0.022). DCS patients had a higher number of severe hepatic (AIS score ≥ 3) and gastrointestinal injuries and more vascular injuries. Most severe abdominal injuries in non-DCS patients were splenic injuries (AIS, 4 and 5) (52.1% versus 37.9%, p = 0.004). CONCLUSION: DCS is a strategy used in unstable trauma patients, severe hepatic, gastrointestinal, multiple abdominal injuries, and mass transfusions. The expected survival rates were achieved in such extreme trauma situations.

Traumatic diaphragmatic rupture: epidemiology, associated injuries, and outcome-an analysis based on the TraumaRegister DGU®.

INTRODUCTION: Traumatic diaphragmatic rupture is a rare injury in the severely injured patient and is most commonly caused by blunt mechanisms. However, penetrating mechanisms can also dominate depending on regional and local factors. Traumatic diaphragmatic rupture is difficult to diagnose and can be missed by primary diagnostic procedures in the resuscitation room. Initially not life-threatening, diaphragmatic ruptures can cause severe sequelae in the patient's long-term course if untreated. The objective of this study was to assess the epidemiology, associated injuries, and outcome of traumatic diaphragmatic ruptures based on a multicenter registry-based analysis. MATERIAL AND METHODS: Data from all patients enrolled in the TraumaRegister DGU® between 2009 and 2018 were retrospectively analyzed. That multicenter database collects data on prehospital, intra-hospital emergency, intensive care therapy, and discharge. Included were all patients with a Maximum Abbreviated Injury Scale (MAIS) score of 3 or above and patients with a MAIS score of 2 who died or were treated in the intensive care unit, for whom standard documentation forms had been completed and who had sustained a diaphragmatic rupture (AIS score of 3 or 4). The data has been analyzed using descriptive statistics and chi-square test or Mann-Whitney U test. RESULTS: Of the 199,933 patients included in the study population, 687 patients (0.3%) had a diaphragmatic rupture. Of these, 71.9% were male. The mean patient age was 46.1 years. Blunt trauma accounted for 73.5% of the injuries. Primary diagnosis was established in the resuscitation room in 93.1% of the patients. Multislice helical computed tomography (MSCT) was performed in 82.7% of the cases. Rib fractures were detected in 60.7% of the patients with a diaphragmatic injury. Patients with diaphragmatic rupture had a higher mean Injury Severity Score (ISS) than patients without a diaphragmatic injury (32.9 vs. 18.6) and a higher mortality rate (13.2% vs. 9.0%). CONCLUSIONS: In contrast to the literature, primary diagnostic procedures in the resuscitation room detected relevant diaphragmatic ruptures (AIS ≥ 3) in more than 90% of the patients in our study population. In addition, complex associated serial rib fractures are an important diagnostic indicator.

[Diagnosis and Treatment of Severe Fournier's Gangrene: Introduction of a Surgical Approach, Evaluation of Risk Factors, Microbiological Characteristics and Review of the Literature]. / Diagnose und zielgerichtete Therapie der Fournier-Gangrän mit septischem Verlauf: Vorstellung eines Behandlungsalgorithmus, Identifikation von Risikofaktoren, Betrachtung des Mikrobioms und Abgleich mit der Literatur.

BACKGROUND: Fournier's gangrene is a necrotising fasciitis type I occurring in the perineal and genital region. The disease expands progressively and still has poor outcome, especially in critical ill patients. This study's focus was placed on the evaluation of risk factors and comorbidities, on the bacteriological spectrum, laboratory analyses, mortality and the course according to the algorithmic approach. METHOD: The medical records were reviewed of 10 patients with severe Fournier's gangrene from 2010 to 2019 who underwent intensive care therapy for at least 48 hours. Ten patients with Fournier's gangrene and sepsis and intensive therapy lasting at least 48 hours were recorded and analysed descriptively. RESULTS: The patient cohort consisted of six men and four women with a median age of 62 years (range 42 - 78 years). The median time between the onset of symptoms and hospitalisation was four days (range 3 - 5 days). The commonest aetiological event was a minor trauma. Predisposing factors included diabetes mellitus (30%) and overweight (80% BMI > 25). 90% of patients exhibited polymicrobial infection. All patients had systemic sepsis with SOFA-Score between 3 and 17 points. The mortality rate was 40%. Significant differences between surviving and deceased patients were shown by PCT, INR and aPTT (p < 0.05). CONCLUSION: Severe Fournier's gangrene continues to be a major challenge with a high mortality rate. Only rapid diagnosis, urgent extensive surgical debridement and intensive care therapy can lead to a favourable outcome in these critically ill patients. Vacuum-assisted closure can be successfully used in the management of soft-tissue defects.

The Significance of Visceral Protection in Preventing Enteroatmospheric Fistulae During Open Abdomen Treatment in Patients With Secondary Peritonitis: A Propensity Score-matched Case-control Analysis.

OBJECTIVE: To evaluate the influence of a visceral protective layer (VPL) on the formation of enteroatmospheric fistulae (EAF) in open abdomen treatment (OAT) for peritonitis. BACKGROUND: EAF formation is a severe complication of OAT. Despite the widespread use of OAT, there are no robust evidence-based recommendations for preventing EAF. METHODS: A total of 120 peritonitis patients with secondary peritonitis as a result of a perforation of a hollow viscus or anastomotic insufficiency who had undergone OAT were included, and 14 clinical parameters were recorded in prospective OAT databases at 2 tertiary referral centers. For this analysis, patients with a VPL were assigned to the treatment group and those without a VPL to the control group. Propensity Score (PS) matching was performed. Known risk factors in OAT such as malignant disease, mortality, emergency operation, OAT duration, and fascial closure were matching variables. The influence of VPL on EAF formation was statistically evaluated using logistic regression analysis. RESULTS: With 34 patients in each group, no notable differences were identified with regard to age, sex, underlying disease, mortality, emergency operation, fascial closure, and OAT duration. Overall, a mortality rate of 22.1% for OAT due to peritonitis was observed. Mean OAT duration was approximately 9 days, and secondary fascial closure was achieved in more than two-thirds of all patients. Fascial traction was used in more than 75% of cases. EAF formation was significantly more frequent in the control group (EAF formation: VPL group 2.9% vs control 26.5%; P = 0.00). In the final regression analysis, the use of VPL resulted in a significant reduction in the risk of EAF formation (odds ratio 0.08; 95% confidence interval 0.01-0.71, P = 0.02), which translates to a relative risk reduction of 89.1%. CONCLUSION: VPL effectively prevents EAF formation during OAT in patients with peritonitis. We recommend the consistent use of VPL as part of a standardized OAT treatment algorithm.

[The Impact of Lockdown during the COVID-19 Pandemic on Care for Elective Patients (C-elective Study) - Results of a Multicenter Survey]. / Die Auswirkung des Lockdowns während der COVID-19-Pandemie auf die Versorgung von elektiven Patienten (C-elective-Studie).

BACKGROUND: Due to the COVID-19-related lockdown regulations, surgical clinics had to cut down elective procedures. The impact of the cancellation and postponing surgery on patients is unclear. METHODS: All patients from six hospitals with canceled surgery during the first lockdown (03/16-04/24/2020) were asked to answer a questionnaire. RESULTS: In total, 225 patients took part. In 88 (39.1%) patients, the disease-related complaints changed, mainly towards an increase in severity (82.6%). That was especially true for hernia patients (44.4%). In 4% of the cases, there was a complication requiring surgery in the time interval between the original date of the operation and the interview. For about a third, the cancelation of scheduled surgery caused major administrative difficulties. Most of the patients (76.3%) understood the measures taken, though 40.4% of them considered that their indication was very urgent. DISCUSSION AND CONCLUSION: The overall complication rate was low, and patients agreed to the measures taken; however, especially hernia patients showed increasing symptoms and some acute incarcerations. It seems reasonable to monitor symptomatic patients closely, in order to prioritise them when surgical capacity is restricted.

Cancer-associated circulating large extracellular vesicles in cholangiocarcinoma and hepatocellular carcinoma.

BACKGROUND & AIMS: Large extracellular vesicles, specifically AnnexinV+ EpCAM+ CD147+ tumour-associated microparticles (taMPs), facilitate the detection of colorectal carcinoma (CRC), non-small cell lung carcinoma (NSCLC) as well as pancreas carcinoma (PaCa). Here we assess the diagnostic value of taMPs for detection and monitoring of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA). Specifically, the aim of this study was to differentiate liver taMPs from other cancer taMPs, such as CRC and NSCLC. METHODS: Fluorescence-activated cell scanning (FACS) was applied to detect various taMP populations in patients' sera that were associated with the presence of a tumour (AnnexinV+ EpCAM+ CD147+ taMPs) or could discriminate between cirrhosis (due to HCV or HBV) and liver cancers (AnnexinV+ EpCAM+ ASGPR1+ taMPs). In total 172 patients with liver cancer (HCC or CCA), 54 with cirrhosis and no liver neoplasia, and 202 control subjects were enrolled. RESULTS: The results indicate that AnnexinV+ EpCAM+ CD147+ taMPs were elevated in HCC and CCA. Furthermore, AnnexinV+ EpCAM+ ASGPR1+ CD133+ taMPs allowed the distinction of liver malignancies (HCC or CCA) and cirrhosis from tumour-free individuals and, more importantly, from patients carrying other non-liver cancers. In addition, AnnexinV+ EpCAM+ ASGPR1+ taMPs were increased in liver cancer-bearing patients compared to patients with cirrhosis that lacked any detectable liver malignancy. The smallest sizes of successfully detected cancers were ranging between 11-15mm. AnnexinV+ EpCAM+ ASGPR1+ taMPs decreased at 7days after curative R0 tumour resection suggesting close correlations with tumour presence. ROC values, sensitivity/specificity scores and positive/negative predictive values (>78%) indicated a potent diagnostic accuracy of AnnexinV+ EpCAM+ ASGPR1+ taMPs. CONCLUSION: These data provide strong evidence that AnnexinV+ EpCAM+ ASGPR1+ taMPs are a novel biomarker of HCC and CCA liquid biopsy that permit a non-invasive assessment of the presence and possible extent of these cancers in patients with advanced liver diseases. LAY SUMMARY: Microparticles (MPs) are small vesicles that bleb from the membrane of every cell, including cancer cells, and are released to circulate in the bloodstream. Since their surface composition is similar to the surface of their underlying parental cell, MPs from the bloodstream can be isolated and by screening their surface components, the presence of their parental cells can be identified. This way, it was possible to detect and discriminate between patients bearing liver cancer and chronic liver cirrhosis.

Functional improvement and maturation of rat and human engineered heart tissue by chronic electrical stimulation.

Spontaneously beating engineered heart tissue (EHT) represents an advanced in vitro model for drug testing and disease modeling, but cardiomyocytes in EHTs are less mature and generate lower forces than in the adult heart. We devised a novel pacing system integrated in a setup for videooptical recording of EHT contractile function over time and investigated whether sustained electrical field stimulation improved EHT properties. EHTs were generated from neonatal rat heart cells (rEHT, n=96) or human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (hEHT, n=19). Pacing with biphasic pulses was initiated on day 4 of culture. REHT continuously paced for 16-18 days at 0.5Hz developed 2.2× higher forces than nonstimulated rEHT. This was reflected by higher cardiomyocyte density in the center of EHTs, increased connexin-43 abundance as investigated by two-photon microscopy and remarkably improved sarcomere ultrastructure including regular M-bands. Further signs of tissue maturation include a rightward shift (to more physiological values) of the Ca(2+)-response curve, increased force response to isoprenaline and decreased spontaneous beating activity. Human EHTs stimulated at 2Hz in the first week and 1.5Hz thereafter developed 1.5× higher forces than nonstimulated hEHT on day 14, an ameliorated muscular network of longitudinally oriented cardiomyocytes and a higher cytoplasm-to-nucleus ratio. Taken together, continuous pacing improved structural and functional properties of rEHTs and hEHTs to an unprecedented level. Electrical stimulation appears to be an important step toward the generation of fully mature EHT.

Automated analysis of contractile force and Ca2+ transients in engineered heart tissue.

Contraction and relaxation are fundamental aspects of cardiomyocyte functional biology. They reflect the response of the contractile machinery to the systolic increase and diastolic decrease of the cytoplasmic Ca(2+) concentration. The analysis of contractile function and Ca(2+) transients is therefore important to discriminate between myofilament responsiveness and changes in Ca(2+) homeostasis. This article describes an automated technology to perform sequential analysis of contractile force and Ca(2+) transients in up to 11 strip-format, fibrin-based rat, mouse, and human fura-2-loaded engineered heart tissues (EHTs) under perfusion and electrical stimulation. Measurements in EHTs under increasing concentrations of extracellular Ca(2+) and responses to isoprenaline and carbachol demonstrate that EHTs recapitulate basic principles of heart tissue functional biology. Ca(2+) concentration-response curves in rat, mouse, and human EHTs indicated different maximal twitch forces (0.22, 0.05, and 0.08 mN in rat, mouse, and human, respectively; P < 0.001) and different sensitivity to external Ca(2+) (EC50: 0.15, 0.39, and 1.05 mM Ca(2+) in rat, mouse, and human, respectively; P < 0.001) in the three groups. In contrast, no difference in myofilament Ca(2+) sensitivity was detected between skinned rat and human EHTs, suggesting that the difference in sensitivity to external Ca(2+) concentration is due to changes in Ca(2+) handling proteins. Finally, this study confirms that fura-2 has Ca(2+) buffering effects and is thereby changing the force response to extracellular Ca(2+).

Effects of proarrhythmic drugs on relaxation time and beating pattern in rat engineered heart tissue.

The assessment of proarrhythmic risks of drugs remains challenging. To evaluate the suitability of rat engineered heart tissue (EHT) for detecting proarrhythmic effects. We monitored drug effects on spontaneous contractile activity and, in selected cases, on action potentials (sharp microelectrode) and Ca2+ transients (Fura-2) and contraction under electrical pacing. The Ito-blocker inhibitor 4-aminopyridine increased action potential duration and T2 and caused aftercontractions, which were abolished by inhibitors of ryanodine receptors (RyR2; JTV-519) or sodium calcium exchanger (NCX; SEA0400). 77 Drugs were then tested at 1-10-100× free therapeutic plasma concentrations (FTPC): Inhibitors of IKr, IKs, Ito, antiarrhythmics (8), drugs withdrawn from market for torsades des pointes arrhythmias (TdP, 5), drugs with measurable (7) or isolated TdP incidence (13), drugs considered safe (14), 28 new chemical entities (NCE). Inhibitors of IKr or IKs had no effect alone, but substantially prolonged relaxation time (T2) when combined at high concentration. 15/33 drugs associated with TdP and 6/14 drugs considered non-torsadogenic (cibenzoline, diltiazem, ebastine, ketoconazole, moxifloxacin, and phenytoin) induced concentration-dependent T2 prolongations (10-100× FTPC). Bepridil, desipramine, imipramine, thioridazine, and erythromycin induced irregular beating. Three NCE prolonged T2, one reduced force. Drugs inhibiting repolarization prolong relaxation in rat EHTs and cause aftercontractions involving RyR2 and NCX. Insensitivity to IKr inhibitors makes rat EHTs unsuitable as general proarrhythmia screen, but favors detection of effects on Ito, IKs + Ito or IKs + IKr. Screening a large panel of drugs suggests that effects on these currents, in addition to IKr, are more common than anticipated.

Analysis of surgical quality indicators after certification as a Hernia Center.

Certifications are an increasingly used tool of quality management in the health care system. The primary goal is to improve the quality of treatment due to implemented measures based on a defined catalog of criteria and standardization of the treatment processes. However, the extent to which this affects medical and health-economic indicators is unknown. Therefore, the study aims to examine the possible effects of the certification as a Reference Center for Hernia Surgery on the treatment quality and reimbursement dimensions. The observation and recording periods were defined as 3 years before (2013-2015) and 3 years after certification as a "Reference Center for Hernia Surgery" (2016-2018). Possible changes due to the certification were examined based on multidimensional data collection and analysis. In addition, the aspects of structure, process and result quality, and the reimbursement situation were reported. One thousand three hundred and nineteen cases before and one thousand four hundred and three cases after certification were included. After the certification, the patients were older (58.1 ± 16.1 vs. 64.0 ± 16.1 years, p < 0.01), had a higher CMI (1.01 vs. 1.06), and a higher ASA score (< III 86.9 vs. 85.5%, p < 0.01). The interventions became more complex (e.g., recurrent incisional hernias 0.5% vs. 1.9%, p < 0.01). The mean length of hospital stay was significantly reduced for incisional hernias (8.8 ± 5.8 vs. 6.7 ± 4.1 days, p < 0.001). The reoperation rate for incisional hernias also decreased significantly from 8.24 to 3.66% (p = 0.04). The postoperative complication rate for inguinal hernias was significantly reduced (3.1 vs. 1.1%, p = 0.002). The reimbursement of the hernia center increased by 27.6%. There were positive changes in process and outcome quality and reimbursement after the certification, which supports the effectivity of certifications in hernia surgery.

[The use of botulinum toxin in hernia surgery: results of a survey in certified hernia centers]. / Die Anwendung von Botulinumtoxin in der Hernienchirurgie: Ergebnisse einer Umfrage an zertifizierten Hernienzentren.

BACKGROUND: Botulinum toxin application into the abdominal wall prior to major hernia repair can reduce the complexity of surgery and has been increasingly used in recent years, even if it is an off-label use. OBJECTIVE: To what extent is botulinum toxin used in hernia surgery in German-speaking countries and what is the current evidence in the literature? MATERIAL AND METHODS: In a voluntary online survey of German Society for General and Visceral Surgery (DGAV)-certified competence centers and reference centers for hernia surgery, aspects of botulinum toxin application were surveyed and the results analyzed. RESULTS: A total of 57 centers took part in the survey, of which 27 (47%) use botulinum toxin. The main reasons for not using it were lack of experience and reimbursement. Of the centers 85% have treated less than 50 patients with botulinum toxin. The main indications were midline hernias (M2-4 according to the EHS classification) with a hernia gap > 10 cm (W3 according to EHS classification) and loss of domain situations. The application was predominantly ultrasound-guided by designated hernia surgeons with 100-200 Allergan or 500 Speywood units 4-6 weeks preoperatively and without complications related to the botulinum toxin application. CONCLUSION: Botulinum toxin injections in hernia surgery appear to be safe and effective. Ultrasound-guided preoperative bilateral administration is supported by the available data. Specific course and information formats should be offered by the hernia surgery institutions.

Contractile abnormalities and altered drug response in engineered heart tissue from Mybpc3-targeted knock-in mice.

Myosin-binding protein C (Mybpc3)-targeted knock-in mice (KI) recapitulate typical aspects of human hypertrophic cardiomyopathy. We evaluated whether these functional alterations can be reproduced in engineered heart tissue (EHT) and yield novel mechanistic information on the function of cMyBP-C. EHTs were generated from cardiac cells of neonatal KI, heterozygous (HET) or wild-type controls (WT) and developed without apparent morphological differences. KI had 70% and HET 20% lower total cMyBP-C levels than WT, accompanied by elevated fetal gene expression. Under standard culture conditions and spontaneous beating, KI EHTs showed more frequent burst beating than WT and occasional tetanic contractions (14/96). Under electrical stimulation (6Hz, 37°C) KI EHTs exhibited shorter contraction and relaxation times and a twofold higher sensitivity to external [Ca(2+)]. Accordingly, the sensitivity to verapamil was 4-fold lower and the response to isoprenaline or the Ca(2+) sensitizer EMD 57033 2- to 4-fold smaller. The loss of EMD effect was verified in 6-week-old KI mice in vivo. HET EHTs were apparently normal under basal conditions, but showed similarly altered contractile responses to [Ca(2+)], verapamil, isoprenaline and EMD. In contrast, drug-induced changes in intracellular Ca(2+) transients (Fura-2) were essentially normal. In conclusion, the present findings in auxotonically contracting EHTs support the idea that cMyBP-C's normal role is to suppress force generation at low intracellular Ca(2+) and stabilize the power-stroke step of the cross bridge cycle. Pharmacological testing in EHT unmasked a disease phenotype in HET. The altered drug response may be clinically relevant.

Next-generation sequencing and syntenic integration of flow-sorted arms of wheat chromosome 4A exposes the chromosome structure and gene content.

Wheat is the third most important crop for human nutrition in the world. The availability of high-resolution genetic and physical maps and ultimately a complete genome sequence holds great promise for breeding improved varieties to cope with increasing food demand under the conditions of changing global climate. However, the large size of the bread wheat (Triticum aestivum) genome (approximately 17 Gb/1C) and the triplication of genic sequence resulting from its hexaploid status have impeded genome sequencing of this important crop species. Here we describe the use of mitotic chromosome flow sorting to separately purify and then shotgun-sequence a pair of telocentric chromosomes that together form chromosome 4A (856 Mb/1C) of wheat. The isolation of this much reduced template and the consequent avoidance of the problem of sequence duplication, in conjunction with synteny-based comparisons with other grass genomes, have facilitated construction of an ordered gene map of chromosome 4A, embracing ≥85% of its total gene content, and have enabled precise localization of the various translocation and inversion breakpoints on chromosome 4A that differentiate it from its progenitor chromosome in the A genome diploid donor. The gene map of chromosome 4A, together with the emerging sequences of homoeologous wheat chromosome groups 4, 5 and 7, represent unique resources that will allow us to obtain new insights into the evolutionary dynamics between homoeologous chromosomes and syntenic chromosomal regions.

Liquid Biopsy in Organ Damage: small extracellular vesicle chip-based assessment of polytrauma.

Background: Despite major advances in medicine, blood-borne biomarkers are urgently needed to support decision-making, including polytrauma. Here, we assessed serum-derived extracellular vesicles (EVs) as potential markers of decision-making in polytrauma. Objective: Our Liquid Biopsy in Organ Damage (LiBOD) study aimed to differentiate polytrauma with organ injury from polytrauma without organ injury. We analysed of blood-borne small EVs at the individual level using a combination of immunocapture and high-resolution imaging. Methods: To this end, we isolated, purified, and characterized small EVs according to the latest Minimal Information for Studies of Extracellular Vesicles (MISEV) guidelines from human blood collected within 24 h post-trauma and validated our results using a porcine polytrauma model. Results: We found that small EVs derived from monocytes CD14+ and CD14+CD61+ were significantly elevated in polytrauma with organ damage. To be precise, our findings revealed that CD9+CD14+ and CD14+CD61+ small EVs exhibited superior performance compared to CD9+CD61+ small EVs in accurately indicating polytrauma with organ damage, reaching a sensitivity and a specificity of 0.81% and 0.97%, respectively. The results in humans were confirmed in an independent porcine model of polytrauma. Conclusion: These findings suggest that these specific types of small EVs may serve as valuable, non-invasive, and objective biomarkers for assessing and monitoring the severity of polytrauma and associated organ damage.

Increased afterload induces pathological cardiac hypertrophy: a new in vitro model.

Increased afterload results in 'pathological' cardiac hypertrophy, the most important risk factor for the development of heart failure. Current in vitro models fall short in deciphering the mechanisms of hypertrophy induced by afterload enhancement. The aim of this study was to develop an experimental model that allows investigating the impact of afterload enhancement (AE) on work-performing heart muscles in vitro. Fibrin-based engineered heart tissue (EHT) was cast between two hollow elastic silicone posts in a 24-well cell culture format. After 2 weeks, the posts were reinforced with metal braces, which markedly increased afterload of the spontaneously beating EHTs. Serum-free, triiodothyronine-, and hydrocortisone-supplemented medium conditions were established to prevent undefined serum effects. Control EHTs were handled identically without reinforcement. Endothelin-1 (ET-1)- or phenylephrine (PE)-stimulated EHTs served as positive control for hypertrophy. Cardiomyocytes in EHTs enlarged by 28.4 % under AE and to a similar extent by ET-1- or PE-stimulation (40.6 or 23.6 %), as determined by dystrophin staining. Cardiomyocyte hypertrophy was accompanied by activation of the fetal gene program, increased glucose consumption, and increased mRNA levels and extracellular deposition of collagen-1. Importantly, afterload-enhanced EHTs exhibited reduced contractile force and impaired diastolic relaxation directly after release of the metal braces. These deleterious effects of afterload enhancement were preventable by endothelin-A, but not endothelin-B receptor blockade. Sustained afterload enhancement of EHTs alone is sufficient to induce pathological cardiac remodeling with reduced contractile function and increased glucose consumption. The model will be useful to investigate novel therapeutic approaches in a simple and fast manner.

Development of a drug screening platform based on engineered heart tissue.

RATIONALE: Tissue engineering may provide advanced in vitro models for drug testing and, in combination with recent induced pluripotent stem cell technology, disease modeling, but available techniques are unsuitable for higher throughput. OBJECTIVE: Here, we present a new miniaturized and automated method based on engineered heart tissue (EHT). METHODS AND RESULTS: Neonatal rat heart cells are mixed with fibrinogen/Matrigel plus thrombin and pipetted into rectangular casting molds in which two flexible silicone posts are positioned from above. Contractile activity is monitored video-optically by a camera and evaluated by a custom-made software program. Fibrin-based mini-EHTs (FBMEs) (150 microL, 600 000 cells) were transferred from molds to a standard 24-well plate two hours after casting. Over time FBMEs condensed from a 12x3x3 mm gel to a muscle strip of 8 mm length and, depending on conditions, 0.2 to 1.3 mm diameter. After 8 to 10 days, FBMEs started to rhythmically deflect the posts. Post properties and the extent of post deflection allowed calculation of rate, force (0.1 to 0.3 mN), and kinetics which was validated in organ baths experiments. FBMEs exhibited a well-developed, longitudinally aligned actinin-positive cardiac muscle network and lectin-positive vascular structures interspersed homogeneously throughout the construct. Analysis of a large series of FBME (n=192) revealed high yield and reproducibility and stability for weeks. Chromanol, quinidine, and erythromycin exerted concentration-dependent increases in relaxation time, doxorubicin decreases in contractile force. CONCLUSIONS: We developed a simple technique to construct large series of EHT and automatically evaluate contractile activity. The method shall be useful for drug screening and disease modeling.