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A major challenge for breath research is the lack of standardization in sampling and analysis. To address this, a test that utilizes a standardized intervention and a defined study protocol has been proposed to explore disparities in breath research across different analytical platforms and to provide benchmark values for comparison. Specifically, the Peppermint Experiment involves the targeted analysis in exhaled breath of volatile constituents of peppermint oil after ingestion of the encapsulated oil. Data from the Peppermint Experiment performed by proton transfer reaction mass spectrometry (PTR-MS) and selected ion flow tube mass spectrometry (SIFT-MS) are presented and discussed herein, including the product ions associated with the key peppermint volatiles, namely limonene, α- and β-pinene, 1,8-cineole, menthol, menthone and menthofuran. The breath washout profiles of these compounds from 65 individuals were collected, comprising datasets from five PTR-MS and two SIFT-MS instruments. The washout profiles of these volatiles were evaluated by comparing the log-fold change over time of the product ion intensities associated with each volatile. Benchmark values were calculated from the lower 95% confidence interval of the linear time-to-washout regression analysis for all datasets combined. Benchmark washout values from PTR-MS analysis were 353 min for the sum of monoterpenes and 1,8-cineole (identical product ions), 173 min for menthol, 330 min for menthofuran, and 218 min for menthone; from SIFT-MS analysis values were 228 min for the sum of monoterpenes, 281 min for the sum of monoterpenes and 1,8-cineole, and 370 min for menthone plus 1,8-cineole. Large inter- and intra-dataset variations were observed, whereby the latter suggests that biological variability plays a key role in how the compounds are absorbed, metabolized and excreted from the body via breath. This variability seems large compared to the influence of sampling and analytical procedures, but further investigations are recommended to clarify the effects of these factors.
We compare results of simulations of solar facular-like conditions performed using the numerical codes MURaM and STAGGER. Both simulation sets have a similar setup, including the initial condition of ≈200 G vertical magnetic flux. After interpolating the output physical quantities to constant optical depth, we compare them and test them against inversion results from solar observations. From the snapshots, we compute the monochromatic continuum in the visible and infrared, and the full Stokes vector of the Fe i spectral line pair around 6301–6302 Å. We compare the predicted spectral lines (at the simulation resolution and after smearing to the HINODE SP/SOT resolution) in terms of their main parameters for the Stokes I line profiles, and of their area and amplitude asymmetry for the Stokes V profiles. The codes produce magnetoconvection with similar appearance and distribution in temperature and velocity. The results also closely match the values from recent relevant solar observations. Although the overall distribution of the magnetic field is similar in both radiation-magnetohydrodynamic (RMHD) simulation sets, a detailed analysis reveals substantial disagreement in the field orientation, which we attribute to the differing boundary conditions. The resulting differences in the synthetic spectra disappear after spatial smearing to the resolution of the observations. We conclude that the two sets of simulations provide robust models of solar faculae. Nevertheless, we also find differences that call for caution when using results from RMHD simulations to interpret solar observational data.
Rheumatoid arthritis is characterised by a progressive, intermittent inflammation at the synovial membrane, which ultimately leads to the destruction of the synovial joint. The synovial membrane as the joint capsule's inner layer is lined with fibroblast-like synoviocytes that are the key player supporting persistent arthritis leading to bone erosion and cartilage destruction. While microfluidic models that model molecular aspects of bone erosion between bone-derived cells and synoviocytes have been established, RA's synovial-chondral axis has not yet been realised using a microfluidic 3D model based on human patient in vitro cultures. Consequently, we established a chip-based three-dimensional tissue coculture model that simulates the reciprocal cross talk between individual synovial and chondral organoids. When co-cultivated with synovial organoids, we could demonstrate that chondral organoids induce a higher degree of cartilage physiology and architecture and show differential cytokine response compared to their respective monocultures highlighting the importance of reciprocal tissue-level cross talk in the modelling of arthritic diseases.
Coculture systems employing adipose tissue-derived mesenchymal stromal/stem cells (ASC) and endothelial cells (EC) represent a widely used technique to model vascularization. Within this system, cell-cell communication is crucial for the achievement of functional vascular network formation. Extracellular vesicles (EVs) have recently emerged as key players in cell communication by transferring bioactive molecules between cells. In this study we aimed to address the role of EVs in ASC/EC cocultures by discriminating between cells, which have received functional EV cargo from cells that have not. Therefore, we employed the Cre-loxP system, which is based on donor cells expressing the Cre recombinase, whose mRNA was previously shown to be packaged into EVs and reporter cells containing a construct of floxed dsRed upstream of the eGFP coding sequence. The evaluation of Cre induced color switch in the reporter system via EVs indicated that there is no EV-mediated RNA transmission either between EC themselves or EC and ASC. However, since Cre mRNA was not found present in EVs, it remains unclear if Cre mRNA is generally not packaged into EVs or if EVs are not taken up by the utilized cell types. Our data indicate that this technique may not be applicable to evaluate EV-mediated cell-to-cell communication in an in vitro setting using EC and ASC. Further investigations will require a functional system showing efficient and specific loading of Cre mRNA or protein into EVs.
Recent progress in machine learning and deep learning has enabled the implementation of plant and crop detection using systematic inspection of the leaf shapes and other morphological characters for identification systems for precision farming. However, the models used for this approach tend to become black-box models, in the sense that it is difficult to trace characters that are the base for the classification. The interpretability is therefore limited and the explanatory factors may not be based on reasonable visible characters. We investigate the explanatory factors of recent machine learning and deep learning models for plant classification tasks. Based on a Daucus carota and a Beta vulgaris image data set, we implement plant classification models and compare those models by their predictive performance as well as explainability. For comparison we implemented a feed forward convolutional neuronal network as a default model. To evaluate the performance, we trained an unsupervised Bayesian Gaussian process latent variable model as well as a convolutional autoencoder for feature extraction and rely on a support vector machine for classification. The explanatory factors of all models were extracted and analyzed. The experiments show, that feed forward convolutional neuronal networks (98.24%
and 96.10% mean accuracy) outperforms the Bayesian Gaussian process latent variable pipeline (92.08% and 94.31% mean accuracy) as well as the convolutional autoenceoder pipeline (92.38% and 93.28%
mean accuracy) based approaches in terms of classification accuracy, even though not significant for Beta vulgaris images. Additionally, we found that the neuronal network used biological uninterpretable image regions for the plant classification task. In contrast to that, the unsupervised learning models rely on explainable visual characters. We conclude that supervised convolutional neuronal networks must be used carefully to ensure biological interpretability. We recommend unsupervised machine learning, careful feature investigation, and statistical feature analysis for biological applications. View Full-Text
Keywords: deep learning; machine learning; plant leaf morphometrics; explainable AI
Diese Arbeit beschreibt eine Augmented Reality (AR) Applikation für den Einsatz in der Hochschullehre zum Thema Industrierobotik. Ziel ist es, sowohl das Lehren als auch das Lernen grundlegender Robotik-Inhalte durch die Bereitstellung einer interaktiven Methode zur Vermittlung der Lehrinhalte für Studierende zu verbessern. Die Studierenden sind in der Lage, direkt mit dem virtuellen Modell eines Industrieroboters zu interagieren und so selbstgesteuert die Lerninhalte zu vertiefen. Diese interaktive Methode verbindet die Studierenden direkt mit den Lehrinhalten und fördert das selbsterforschende Lernen. Eine weitere Anwendungsmöglichkeit sieht die Kombination einer Lektorenversion der AR Experience mit der Studierendenversion vor. Der Lektor hat die Möglichkeit, das Modell in AR zu steuern bzw. zu verändern und die Studierenden können auf Ihren Mobilgeräten die Änderungen live in AR mitverfolgen, um so auch im Distance Learning eine Verbindung Lektor – Studierende – Inhalt zusätzlich zu Videokonferenz-Tools herzustellen.
In the present work, using the recently introduced framework of local geometric deformations, special types of vector fields – so-called hidden Killing vector fields – are constructed, which solve the Killing equation not globally, but only locally, i.e. in local subregions of spacetime. Taking advantage of the fact that the vector fields coincide locally with Killing fields and therefore allow the consideration of integral laws that convert into exact physical conservation laws on local scales, balance laws in dynamical systems without global Killing symmetries are derived that mimic as closely as possible the conservation laws for energy and angular momentum of highly symmetric models. The utility of said balance laws is demonstrated by a concrete geometric example, namely a toy model for the binary merger of two extremal Reissner–Nordström black holes.
Wissensarbeiter:innen verbringen den überwiegenden Teil ihrer Arbeitszeit sitzend vor dem Computer. Die negativen Folgen von langem Sitzen für die Gesundheit sind bekannt: Zu langes Sitzen bedingt einen niedrigen Kalorienverbrauch, der Stoffwechsel und das Herz-Kreislaufsystem laufen auf Sparflamme. Entsprechend steigt das Risiko für Übergewicht, Diabetes, Bandscheibenvorfall und Herz-Kreislauf- Erkrankungen. Unter den Folgen von langem Sitzen leiden aber nicht nur die betroffenen Mitarbeiter:innen selbst, sondern auch deren Arbeitgeber:innen, weil Mitarbeiter:innen mit einem auf Bewegungsmangel zurückzuführenden reduzierten physischen und psychischen Wohlbefinden weniger produktiv und kreativ arbeiten bzw. aufgrund von Erkrankungen erst gar nicht arbeiten können. Der vorliegende Beitrag zeigt auf, wie Unternehmen durch das Setzen sanfter Bewegungszwänge, den Einsatz dynamischer Arbeitsstationen sowie die Integration niederschwelliger Fitnessmodule in die Bürolandschaft für mehr körperliche Aktivität im Arbeitsalltag sorgen können.
Hybrid courses with a focus on practice-orientated education and self-guided learning phases are on the rise on the higher education sector. Disciplines in Life Sciences implicate a high degree of practical laboratory expertise. The University of Applied Sciences (UAS) in Vienna, Austria, has thus been endeavoured offering students a high qualitative education integrating hybrid courses based on PBL principles, which consist of on-site (including the transmission of necessary background and practical laboratory training) and off-site (including self-study phases) sessions. As practical laboratory units are central in those courses, the restrictive measures, including the transition to a complete online teaching format due to the first Covid-19-pandemic lock-down, had severe effects on the implementation and the quality of the curriculum. According to surveys made specifically to address this problematic situation, it can be concluded that on-site practical units are fundamental for certain disciplines such as Life Sciences.
The repair of large bone defects remains challenging and often requires graft material due to limited availability of autologous bone. In clinical settings, collagen sponges loaded with excessive amounts of bone morphogenetic protein 2 (rhBMP-2) are occasionally used for the treatment of bone non-unions, increasing the risk of adverse events. Therefore, strategies to reduce rhBMP-2 dosage are desirable. Silk scaffolds show great promise due to their favorable biocompatibility and their utility for various biofabrication methods. For this study, we generated silk scaffolds with axially aligned pores, which were subsequently treated with 10× simulated body fluid (SBF) to generate an apatitic calcium phosphate coating. Using a rat femoral critical sized defect model (CSD) we evaluated if the resulting scaffold allows the reduction of BMP-2 dosage to promote efficient bone repair by providing appropriate guidance cues. Highly porous, anisotropic silk scaffolds were produced, demonstrating good cytocompatibility in vitro and treatment with 10× SBF resulted in efficient surface coating. In vivo, the coated silk scaffolds loaded with a low dose of rhBMP-2 demonstrated significantly improved bone regeneration when compared to the unmineralized scaffold. Overall, our findings show that this simple and cost-efficient technique yields scaffolds that enhance rhBMP-2 mediated bone healing.
The natural extracellular matrix (ECM) provides the optimal environment for cells. Many enzymatic or non-enzymatic based strategies to extract ECM proteins from tissues were published over the past years. However, every single isolation strategy reported so far is associated with specific bottlenecks. In this study, frequently used strategies to isolate ECM from human placenta or adipose tissue using Tris-, serum-, or pepsin-based buffers were compared. The resulting ECM proteins were biochemically characterized by analysis of cellular remnants using Hoechst DNA staining, glycosaminoglycan (GAG) content by dimethylmethylene blue, visualization of protein bands using sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis combined with amino acid quantification, and assessment of the proangiogenic profile using an angiogenesis array. Tris-NaCl-extracted ECM proteins showed a high heterogenic degree of extracted proteins, bioactive growth factors, and GAGs, but no collagen-I. Active serum-extracted ECM showed significant lower DNA remnants when compared with the Tris-NaCl isolation strategy. Pepsin-extracted ECM was rich in collagen-I and low amounts of remaining bioactive growth factors. This strategy was most effective to reduce DNA amounts when compared with the other isolation strategies. Pepsin-extracted ECM from both tissues easily gelled at 37°C, whereas the other extracted ECM strategies did not gel at 37°C (Tris-NaCl: liquid; serum: sponge). All relevant characteristics (DNA residues, ECM diversity and bioactivity, shape) of the extracted ECM proteins highly depend on its isolation strategy and could still be optimized. Impact statement The natural human extracellular matrix (ECM) is the ideal cell niche. Various strategies were reported to isolate human ECM components from various sources. In this article, we compared frequently used methods and compared their characteristics (DNA remnants, glycosaminoglycan content, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, amino acid quantification, angiogenesis array, and gel formation). We conclude that more research is still necessary to optimize current isolation approaches for in vitro or in vivo applications of human ECM.
The additive manufacturing of low elastic modulus alloys that have a certain level of porosity for biomedical needs is a growing area of research. Here, we show the results of manufacturing of porous and dense samples by a laser powder bed fusion (LPBF) of Ti-Nb alloy, using two distinctive fusion strategies. The nanostructured Ti-Nb alloy powders were produced by mechanical alloying and have a nanostructured state with nanosized grains up to 90 nm. The manufactured porous samples have pronounced open porosity and advanced roughness, contrary to dense samples with a relatively smooth surface profile. The structure of both types of samples after LPBF is formed by uniaxial grains having micro- and nanosized features. The inner structure of the porous samples is comprised of an open interconnected system of pores. The volume fraction of isolated porosity is 2 vol. % and the total porosity is 20 vol. %. Cell viability was assessed in vitro for 3 and 7 days using the MG63 cell line. With longer culture periods, cells showed an increased cell density over the entire surface of a porous Ti-Nb sample. Both types of samples are not cytotoxic and could be used for further in vivo studies.
Due to their strong biomimetic potential, silk fibroin (SF) hydrogels are impressive candidates for tissue engineering, due to their tunable mechanical properties, biocompatibility, low immunotoxicity, controllable biodegradability, and a remarkable capacity for biomaterial modification and the realization of a specific molecular structure. The fundamental chemical and physical structure of SF allows its structure to be altered using various crosslinking strategies. The established crosslinking methods enable the formation of three-dimensional (3D) networks under physiological conditions. There are different chemical and physical crosslinking mechanisms available for the generation of SF hydrogels (SFHs). These methods, either chemical or physical, change the structure of SF and improve its mechanical stability, although each method has its advantages and disadvantages. While chemical crosslinking agents guarantee the mechanical strength of SFH through the generation of covalent bonds, they could cause some toxicity, and their usage is not compatible with a cell-friendly technology. On the other hand, physical crosslinking approaches have been implemented in the absence of chemical solvents by the induction of β-sheet conformation in the SF structure. Unfortunately, it is not easy to control the shape and properties of SFHs when using this method. The current review discusses the different crosslinking mechanisms of SFH in detail, in order to support the development of engineered SFHs for biomedical applications.
Background: In spite of advances in the treatment of cartilage defects using cell and scaffold-based therapeutic strategies, the long-term outcome is still not satisfying since clinical scores decline years after treatment. Scaffold materials currently used in clinical settings have shown limitations in providing suitable biomechanical properties and an authentic and protective environment for regenerative cells. To tackle this problem, we developed a scaffold material based on decellularised human articular cartilage.
Methods: Human articular cartilage matrix was engraved using a CO2 laser and treated for decellularisation and glycosaminoglycan removal. Characterisation of the resulting scaffold was performed via mechanical testing, DNA and GAG quantification and in vitro cultivation with adipose-derived stromal cells (ASC). Cell vitality, adhesion and chondrogenic differentiation were assessed. An ectopic, unloaded mouse model was used for the assessment of the in vivo performance of the scaffold in combination with ASC and human as well as bovine chondrocytes. The novel scaffold was compared to a commercial collagen type I/III scaffold.
Findings: Crossed line engravings of the matrix allowed for a most regular and ubiquitous distribution of cells and chemical as well as enzymatic matrix treatment was performed to increase cell adhesion. The biomechanical characteristics of this novel scaffold that we term CartiScaff were found to be superior to those of commercially available materials. Neo-tissue was integrated excellently into the scaffold matrix and new collagen fibres were guided by the laser incisions towards a vertical alignment, a typical feature of native cartilage important for nutrition and biomechanics. In an ectopic, unloaded in vivo model, chondrocytes and mesenchymal stromal cells differentiated within the incisions despite the lack of growth factors and load, indicating a strong chondrogenic microenvironment within the scaffold incisions. Cells, most noticeably bone marrow-derived cells, were able to repopulate the empty chondrocyte lacunae inside the scaffold matrix.
Interpretation: Due to the better load-bearing, its chondrogenic effect and the ability to guide matrix-deposition, CartiScaff is a promising biomaterial to accelerate rehabilitation and to improve long term clinical success of cartilage defect treatment.
Funding: Austrian Research Promotion Agency FFG ("CartiScaff" #842455), Lorenz Böhler Fonds (16/13), City of Vienna Competence Team Project Signaltissue (MA23, #18-08).
Keywords: Cartilage regeneration; Decellularisation; Ectopic animal model; Laser engraving; Mechanical testing; Repopulation.
Erosion of the epigenetic DNA methylation landscape is a widely recognized hallmark of aging. Emerging advances in high throughput sequencing techniques, in particular DNA methylation data analysis, have resulted in the establishment of precise human and murine age prediction tools. In vertebrates, methylation of cytosine at the C5 position of CpG dinucleotides is executed by DNA methyltransferases (DNMTs) whereas the process of enzymatic demethylation is highly dependent on the activity of the ten-eleven translocation methylcytosine dioxygenase (TET) family of enzymes. Here, we report the identification of the key players constituting the DNA methylation machinery in the short-lived teleost aging model Nothobranchius furzeri. We present a comprehensive spatio-temporal expression profile of the methylation-associated enzymes from embryogenesis into late adulthood, thereby covering the complete killifish life cycle. Data mining of the N. furzeri genome produced five dnmt gene family orthologues corresponding to the mammalian DNMTs (DNMT1, 2, 3A, and 3B). Comparable to other teleost species, N. furzeri harbors multiple genomic copies of the de novo DNA methylation subfamily. A related search for the DNMT1 recruitment factor UHRF1 and TET family members resulted in the identification of N. furzeri uhrf1, tet1, tet2, and tet3. Phylogenetic analysis revealed high cross-species similarity on the amino acid level of all individual dnmts, tets, and uhrf1, emphasizing a high degree of functional conservation. During early killifish development all analyzed dnmts and tets showed a similar expression profile characterized by a strong increase in transcript levels after fertilization, peaking either at embryonic day 6 or at the black eye stage of embryonic development. In adult N. furzeri, DNA methylation regulating enzymes showed a ubiquitous tissue distribution. Specifically, we observed an age-dependent downregulation of dnmts, and to some extent uhrf1, which correlated with a significant decrease in global DNA methylation levels in the aging killifish liver and muscle. The age-dependent DNA methylation profile and spatio-temporal expression characteristics of its enzymatic machinery reported here may serve as an essential platform for the identification of an epigenetic aging clock in the new vertebrate model system N. furzeri.
There is critical unmet need for new vascularized tissues to support or replace injured tissues and organs. Various synthetic and natural materials were already established for use of two-dimensional (2D) and three-dimensional (3D) in vitro neovascularization assays, however, they still cannot mimic the complex functions of the sum of the extracellular matrix (ECM) in native intact tissue. Currently, this issue is only addressed by artificial products such as Matrigel™, which comprises a complex mixture of ECM proteins, extracted from animal tumor tissue. Despite its outstanding bioactivity, the isolation from tumor tissue hinders its translation into clinical applications. Since nonhuman ECM proteins may cause immune reactions, as are frequently observed in clinical trials, human ECM proteins represent the best option when aiming for clinical applications. Here, we describe an effective method of isolating a human placenta substrate (hpS) that induces the spontaneous formation of an interconnected network of green fluorescence-labeled human umbilical vein endothelial cells (gfpHUVECs) in vitro. The substrate was biochemically characterized by using a combination of bicinchoninic acid (BCA) assay, DNA, and glycosaminoglycan (GAG) content assays, sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) analysis and Western blot, angiogenesis arrays, chromatographic thrombin detection, high performance liquid chromatography (HPLC)-based amino acid quantification analysis, and assessment of antimicrobial properties. 2D in vitro cell culture experiments have been performed to determine the vasculogenic potential of hpS, which demonstrated that cell networks developed on hpS show a significantly higher degree of complexity (number of tubules/junctions; total/mean tube length) when compared with Matrigel. As 3D cell culture techniques represent a more accurate representation of the in vivo condition, the substrate was 3D solidified using various natural polymers. 3D in vitro vasculogenesis assays have been performed by seeding gfpHUVECs in an hpS-fibrinogen clot. In conclusion, hpS provides a potent human/material-based alternative to xenogenic-material-based biomaterials for vascularization strategies in tissue engineering.
The responsibility of a lecturer is not only to share his or her knowledge with the students in an easy to understand manner, but also to help the students to embed new knowledge and to encourage the development of higher-order cognitive skills via applied exercises.
In order to meet the growing demand for blended learning approaches a new course concept was established in autumn 2018. To enhance comprehension and to provide opportunities for self-assessment, web-based training units were implemented by using the interactive learning software “Articulate Storyline”. Students had to prepare at home for the course units by completing interactive chapters. Their learning outcome was assessed by online quizzes at the end of each chapter. Online Training chapters allowed time to focus on selected topics and to repeat key messages in following presence units.
Additionally, guided group exercises were performed to promote analytic skills and abstract thinking. The students had to apply and combine their knowledge to solve problem-based challenges.
An optional revision course was offered to the students, which allowed for interactive repetition of the acquired knowledge with the focus on student-to-lecturer dialog.
An analysis based on a written evaluation of this course resulted in a positive feedback from the students, in particular regarding the guided exercises and the offered revision course. According to the students the group exercises allowed to process the learned subjects, promoted the group climate and were a convenient diversion from the frontal lecture format. Students who attended the revision course on a regular basis showed a better performance at the final exam and exceeded especially at interdisciplinary questions.
The first implementation of this master´s degree course indicated that the combination of web-based training elements with frontal lecture elements, guided exercises stimulating cognitive skills and an optional revision course can teach students the basics of biology in an understandable way. This course structure is especially applicable to teach basic subjects for groups of students with varying initial knowledge.
Financial support from the City of Vienna project PBL in Molecular Life Science (21-06) is gratefully acknowledged.
Talipot starch, a non-conventional starch source with a high yield (76%) from the stem pith of talipot palm (Corypha umbraculifera L.) was subjected to three different thermal treatments (dry-heat, heat-moisture and autoclave treatments) prior to phosphorylation. Upon dual modification of starch with thermal treatments and phosphorylation, the phosphorous content and degree of crosslinking significantly increased (p ≤ 0.05) and was confirmed by the increased peak intensity of P=O and P–O–C stretching vibrations compared to phosphorylated talipot starch in the FT-IR spectrum. The highest degree of crosslinking (0.00418) was observed in the autoclave pretreated phosphorylated talipot starch sample. Thermal pretreatment remarkably changed the granule morphology by creating fissures and grooves. The amylose content and relative crystallinity of all phosphorylated talipot starches significantly decreased (p ≤ 0.05) due to crosslinking by the formation of phosphodiester bonds, reducing the swelling power of dual-modified starches. Among all modified starches, dry-heat pretreated phosphorylated starch gel showed an improved light transmittance value of 28.4%, indicating reduced retrogradation tendency. Pasting and rheological properties represented that the thermal pretreated phosphorylated starch formed stronger gels that improved thermal and shear resistance. Autoclave treatment before phosphorylation of talipot starch showed the highest resistant starch content of 48.08%.