Overexpression of connexin 43 using a retroviral vector improves electrical coupling of skeletal myoblasts with cardiac myocytes in vitro.

BACKGROUND: Organ transplantation is presently often the only available option to repair a damaged heart. As heart donors are scarce, engineering of cardiac grafts from autologous skeletal myoblasts is a promising novel therapeutic strategy. The functionality of skeletal muscle cells in the heart milieu is, however, limited because of their inability to integrate electrically and mechanically into the myocardium. Therefore, in pursuit of improved cardiac integration of skeletal muscle grafts we sought to modify primary skeletal myoblasts by overexpression of the main gap-junctional protein connexin 43 and to study electrical coupling of connexin 43 overexpressing myoblasts to cardiac myocytes in vitro. METHODS: To create an efficient means for overexpression of connexin 43 in skeletal myoblasts we constructed a bicistronic retroviral vector MLV-CX43-EGFP expressing the human connexin 43 cDNA and the marker EGFP gene. This vector was employed to transduce primary rat skeletal myoblasts in optimised conditions involving a concomitant use of the retrovirus immobilising protein RetroNectin and the polycation transduction enhancer Transfectam. The EGFP-positive transduced cells were then enriched by flow cytometry. RESULTS: More than four-fold overexpression of connexin 43 in the transduced skeletal myoblasts, compared with non-transduced cells, was shown by Western blotting. Functionality of the overexpressed connexin 43 was demonstrated by microinjection of a fluorescent dye showing enhanced gap-junctional intercellular transfer in connexin 43 transduced myoblasts compared with transfer in non-transduced myoblasts. Rat cardiac myocytes were cultured in multielectrode array culture dishes together with connexin 43/EGFP transduced skeletal myoblasts, control non-transduced skeletal myoblasts or alone. Extracellular field action potential activation rates in the co-cultures of connexin 43 transduced skeletal myoblasts with cardiac myocytes were significantly higher than in the co-cultures of non-transduced skeletal myoblasts with cardiac myocytes and similar to the rates in pure cultures of cardiac myocytes. CONCLUSION: The observed elevated field action potential activation rate in the co-cultures of cardiac myocytes with connexin 43 transduced skeletal myoblasts indicates enhanced cell-to-cell electrical coupling due to overexpression of connexin 43 in skeletal myoblasts. This study suggests that retroviral connexin 43 transduction can be employed to augment engineering of the electrocompetent cardiac grafts from patients own skeletal myoblasts

Climatological and radiative properties of midlatitude cirrus clouds derived by automatic evaluation of lidar measurements

Cirrus, i.e., high, thin clouds that are fully glaciated, play an important role in the Earth's radiation budget as they interact with both long- and shortwave radiation and affect the water vapor budget of the upper troposphere and stratosphere. Here, we present a climatology of midlatitude cirrus clouds measured with the same type of ground-based lidar at three midlatitude research stations: at the Swiss high alpine Jungfraujoch station (3580 m a.s.l.), in Zürich (Switzerland, 510 m a.s.l.), and in Jülich (Germany, 100 m a.s.l.). The analysis is based on 13 000 h of measurements from 2010 to 2014. To automatically evaluate this extensive data set, we have developed the Fast LIdar Cirrus Algorithm (FLICA), which combines a pixel-based cloud-detection scheme with the classic lidar evaluation techniques. We find mean cirrus optical depths of 0.12 on Jungfraujoch and of 0.14 and 0.17 in Zürich and Jülich, respectively. Above Jungfraujoch, subvisible cirrus clouds (τ < 0.03) have been observed during 6 % of the observation time, whereas above Zürich and Jülich fewer clouds of that type were observed. Cirrus have been observed up to altitudes of 14.4 km a.s.l. above Jungfraujoch, whereas they have only been observed to about 1 km lower at the other stations. These features highlight the advantage of the high-altitude station Jungfraujoch, which is often in the free troposphere above the polluted boundary layer, thus enabling lidar measurements of thinner and higher clouds. In addition, the measurements suggest a change in cloud morphology at Jungfraujoch above ∼ 13 km, possibly because high particle number densities form in the observed cirrus clouds, when many ice crystals nucleate in the high supersaturations following rapid uplifts in lee waves above mountainous terrain. The retrieved optical properties are used as input for a radiative transfer model to estimate the net cloud radiative forcing, CRFNET, for the analyzed cirrus clouds. All cirrus detected here have a positive CRFNET. This confirms that these thin, high cirrus have a warming effect on the Earth's climate, whereas cooling clouds typically have cloud edges too low in altitude to satisfy the FLICA criterion of temperatures below −38 °C. We find CRFNET = 0.9 W m−2 for Jungfraujoch and 1.0 W m−2 (1.7 W m−2) for Zürich (Jülich). Further, we calculate that subvisible cirrus (τ < 0.03) contribute about 5 %, thin cirrus (0.03 < τ < 0.3) about 45 %, and opaque cirrus (0.3 < τ) about 50 % of the total cirrus radiative forcing

Molecular basis for the bifunctional Uba4-Urm1 sulfur-relay system in tRNA thiolation and ubiquitin-like conjugation.

The chemical modification of tRNA bases by sulfur is crucial to tune translation and to optimize protein synthesis. In eukaryotes, the ubiquitin-related modifier 1 (Urm1) pathway is responsible for the synthesis of 2-thiolated wobble uridine (U34 ). During the key step of the modification cascade, the E1-like activating enzyme ubiquitin-like protein activator 4 (Uba4) first adenylates and thiocarboxylates the C-terminus of its substrate Urm1. Subsequently, activated thiocarboxylated Urm1 (Urm1-COSH) can serve as a sulfur donor for specific tRNA thiolases or participate in ubiquitin-like conjugation reactions. Structural and mechanistic details of Uba4 and Urm1 have remained elusive but are key to understand the evolutionary branch point between ubiquitin-like proteins (UBL) and sulfur-relay systems. Here, we report the crystal structures of full-length Uba4 and its heterodimeric complex with its substrate Urm1. We show how the two domains of Uba4 orchestrate recognition, binding, and thiocarboxylation of the C-terminus of Urm1. Finally, we uncover how the catalytic domains of Uba4 communicate efficiently during the reaction cycle and identify a mechanism that enables Uba4 to protect itself against self-conjugation with its own product, namely activated Urm1-COSH

Biomechanical analysis of a synthetic femoral spiral fracture model: Do end caps improve retrograde flexible intramedullary nail fixation?

<p>Abstract</p> <p>Background</p> <p>Elastic Stable intramedullary Nailing (ESIN) of dislocated diaphyseal femur fractures has become an accepted method for the treatment in children and adolescents with open physis. Studies focused on complications of this technique showed problems regarding stability, usually in complex fracture types such as spiral fractures and in older children weighing > 40 kg. Biomechanical in vitro testing was performed to evaluate the stability of simulated spiral femoral fractures after retrograde flexible titanium intramedullary nail fixation with and without End caps.</p> <p>Methods</p> <p>Eight synthetic adolescent-size femoral bone models (Sawbones^®with a medullar canal of 10 mm and a spiral fracture of 100 mm length identically sawn by the manufacturer) were used for each group. Both groups underwent retrograde fixation with two 3.5 mm Titanium C-shaped nails inserted from medial and lateral entry portals. In the End Cap group the ends of the nails of the eight specimens were covered with End Caps (Synthes Company, Oberdorf, Switzerland) at the distal entry.</p> <p>Results</p> <p>Beside posterior-anterior stress (4.11 Nm/mm vs. 1.78 Nm/mm, p < 0.001), the use of End Caps demonstrated no higher stability in 4-point bending compared to the group without End Caps (anterior-posterior bending 0.27 Nm/mm vs. 0.77 Nm/mm, p < 0.001; medial-lateral bending 0.8 Nm/mm vs. 1.10 Nm/mm, p < 0.01; lateral-medial bending 0.53 Nm/mm vs. 0.86 Nm/mm, p < 0.001) as well as during internal rotation (0.11 Nm/° vs. 0.14 Nm/°, p < 0.05). During compression in 9°- position and external rotation there was no statistical significant difference (0.37 Nm/° vs. 0.32 Nm/°, p = 0.13 and 1.29 mm vs. 2.18 mm, p = 0.20, respectively) compared to the "classic" 2-C-shaped osteosynthesis without End Caps.</p> <p>Conclusion</p> <p>In this biomechanical study the use of End Caps did not improve the stability of the intramedullary flexible nail osteosynthesis.</p

No longer locally extinct? Tracing the origins of a lion (Panthera leo) living in Gabon

Lions (Panthera leo) are of particular conservation concern due to evidence of recent, widespread population declines in what has hitherto been seen as a common species, robust to anthropogenic disturbance. Here we use non-invasive methods to recover complete mitochondrial genomes from single hair samples collected in the field in order to explore the identity of the Gabonese Plateaux Batéké lion. Comparison of the mitogenomes against a comprehensive dataset of African lion sequences that includes relevant geographically proximate lion populations from both contemporary and ancient sources, enabled us to identify the Plateaux Batéké lion as a close maternal relative to now extirpated populations found in Gabon and nearby Congo during the twentieth century, and to extant populations of Southern Africa. Our study demonstrates the relevance of ancient DNA methods to field conservation work, and the ability of trace field samples to provide copious genetic information about free-ranging animals.Acknowledgements We thank the laboratory technicians of the Centre for GeoGenetics and the staff of the Danish National High-Throughput DNA Sequencing Centre for technical assistance. This project received funding from the European Union�s Seventh Framework Programme for research, technological development and demonstration under grant agreement no. FP7-PEOPLE-2011-IEF-298820, and ERC Consolidator Grant 681396 � Extinction Genomics. Fieldwork in Gabon was funded by Panthera and The Aspinall Foundation. We thank the Staff at Projet Protection Gorilles Gabon and the Gabonese National Park Agency for their assistance in fieldwork. Thanks also to Tony King for helpful comments on the manuscript. Permission to collect and export the samples was granted by the respective Ministries of the Environment (Eaux et Forets) in Gabon and Congo, and by Gabon�s National Park Agency (ANPN).Scopu

Intermittent existence of a southern Californian upwelling cell during submillennial climate change of the last 60 kyr

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/95021/1/palo1108.pd

FHIR-DHP: A standardized clinical data harmonisation pipeline for scalable AI application deployment

Background Increasing digitalisation in the medical domain gives rise to large amounts of healthcare data which has the potential to expand clinical knowledge and transform patient care if leveraged through artificial intelligence (AI). Yet, big data and AI oftentimes cannot unlock their full potential at scale, owing to non-standardised data formats, lack of technical and semantic data interoperability, and limited cooperation between stakeholders in the healthcare system. Despite the existence of standardised data formats for the medical domain, such as Fast Healthcare Interoperability Resources (FHIR), their prevalence and usability for AI remains limited.Objective We developed a data harmonisation pipeline (DHP) for clinical data sets relying on the common FHIR data standard.Methods We validated the performance and usability of our FHIR-DHP with data from the MIMIC IV database including > 40,000 patients admitted to an intensive care unit.Results We present the FHIR-DHP workflow in respect of transformation of “raw” hospital records into a harmonised, AI-friendly data representation. The pipeline consists of five key preprocessing steps: querying of data from hospital database, FHIR mapping, syntactic validation, transfer of harmonised data into the patient-model database and export of data in an AI-friendly format for further medical applications. A detailed example of FHIR-DHP execution was presented for clinical diagnoses records.Conclusions Our approach enables scalable and needs-driven data modelling of large and heterogenous clinical data sets. The FHIR-DHP is a pivotal step towards increasing cooperation, interoperability and quality of patient care in the clinical routine and for medical research

Evidence Based Development of a Novel Lateral Fibula Plate (VariAx Fibula) Using a Real CT Bone Data Based Optimization Process During Device Development

Development of novel implants in orthopaedic trauma surgery is based on limited datasets of cadaver trials or artificial bone models. A method has been developed whereby implants can be constructed in an evidence based method founded on a large anatomic database consisting of more than 2.000 datasets of bones extracted from CT scans. The aim of this study was the development and clinical application of an anatomically pre-contoured plate for the treatment of distal fibular fractures based on the anatomical database

Carbonates from the ancient world's longest aqueduct:A testament of Byzantine water management

The fourth‐ and fifth‐century aqueduct system of Constantinople is, at 426 km, the longest water supply line of the ancient world. Carbonate deposits in the aqueduct system provide an archive of both archaeological developments and palaeo‐environmental conditions during the depositional period. The 246‐km‐long aqueduct line from the fourth century used springs from a small aquifer, whereas a 180‐km‐long fifth‐century extension to the west tapped a larger aquifer. Although historical records testify at least 700 years of aqueduct activity, carbonate deposits in the aqueduct system display less than 27 years of operation. This implies that the entire system must have been cleaned of carbonate, presumably during regular campaigns. A 50‐km‐long double‐aqueduct section in the central part of the system may have been a costly but practical solution to allow repairs and cleaning of the aqueducts of carbonate to ascertain a continuous water supply to the city. The fifth‐century channel was commonly contaminated with clay, caused by the nature of the aqueduct system and possible local damage to the channel. This clay‐rich water could have been one of the reasons for the construction of large reservoirs in Constantinople. imageLeverhulme Trust http://dx.doi.org/10.13039/501100000275Deutsche Forschungsgemeinschaft http://dx.doi.org/10.13039/50110000165

Разработка импульсной системы питания сварочной дуги на базе искусственной формирующей линии

Объектом исследования является импульсная система питания сварочной дуги, горящей в динамическом режиме. Предмет исследования – система импульсного питания сварочной дуги на базе искусственной формирующей линии неплавящимся электродом. Цель работы – анализ и разработка импульсной системы питания сварочной дуги на базе искусственной формирующей линии неплавящимся электродом в аргоне. Создана действующая экспериментальная установка для сварки дугой горящей в динамическом режиме с униполярными импульсами. Достоверность результатов и выводов подтверждена их сходимостью с экспериментальными данными, полученными методом осциллографированияSubject of research – power system for welding in dynamic mode nonconsumable electrode unidirectional pulses of current. Purpose of research – research and development of the power system for non-consumable electrode arc welding in argon in dynamic mode. The acting experimental setup for non-consumable electrode arc welding in argon in dynamic mode was created. The validity of results and inferences was confirmed by their convergence with experimental data obtained by oscillographic testing of developed power system for non-consumable electrode arc welding in argon in dynamic mode