A Novel Neurofilament Light Chain ELISA Validated in Patients with Alzheimer’s Disease,Frontotemporal Dementia,and Subjective Cognitive Decline,and the Evaluation of Candidate Proteins for Immunoassay Calibration

Neurofilament light chain (Nf-L) is a well-known biomarker for axonal damage; however, the corresponding circulating Nf-L analyte in cerebrospinal fluid (CSF) is poorly characterized. We therefore isolated new monoclonal antibodies against synthetic peptides, and these monoclonals were characterized for their specificity on brain-specific intermediate filament proteins. Two highly specific antibodies, ADx206 and ADx209, were analytically validated for CSF applications according to well-established criteria. Interestingly, using three different sources of purified Nf-L proteins, a significant impact on interpolated concentrations was observed. With a lower limit of analytical sensitivity of 100 pg/mL using bovine Nf-L as the calibrator, we were able to quantify the Nf-L analyte in each sample, and these Nf-L concentrations were highly correlated to the Uman diagnostics assay (Spearman rho = 0.97, p < 0.001). In the clinical diagnostic groups, the new Nf-L ELISA could discriminate patients with Alzheimer’s disease (AD, n = 20) from those with frontotemporal lobe dementia (FTD, n = 20) and control samples with subjective cognitive decline (SCD, n = 20). Henceforth, this novel Nf-L ELISA with well-defined specificity and epitopes can be used to enhance our understanding of harmonizing the use of Nf-L as a clinically relevant marker for neurodegeneration in CSF.     

Adaptation of Oxidative Phosphorylation Machinery Compensates for Hepatic Lipotoxicity in Early Stages of MAFLD

Alterations in mitochondrial function are an important control variable in the progression of metabolic dysfunction-associated fatty liver disease (MAFLD), while also noted by increased de novo lipogenesis (DNL) and hepatic insulin resistance. We hypothesized that the organization and function of a mitochondrial electron transport chain (ETC) in this pathologic condition is a consequence of shifted substrate availability. We addressed this question using a transgenic mouse model with increased hepatic insulin resistance and DNL due to constitutively active human SREBP-1c. The abundance of ETC complex subunits and components of key metabolic pathways are regulated in the liver of these animals. Further omics approaches combined with functional assays in isolated liver mitochondria and primary hepatocytes revealed that the SREBP-1c-forced fatty liver induced a substrate limitation for oxidative phosphorylation, inducing enhanced complex II activity. The observed increased expression of mitochondrial genes may have indicated a counteraction. In conclusion, a shift of available substrates directed toward activated DNL results in increased electron flows, mainly through complex II, to compensate for the increased energy demand of the cell. The reorganization of key compounds in energy metabolism observed in the SREBP-1c animal model might explain the initial increase in mitochondrial function observed in the early stages of human MAFLD.     

SARS-CoV-2 Infection Dysregulates Cilia and Basal Cell Homeostasis in the Respiratory Epithelium of Hamsters

Similar to many other respiratory viruses, SARS-CoV-2 targets the ciliated cells of the respiratory epithelium and compromises mucociliary clearance, thereby facilitating spread to the lungs and paving the way for secondary infections. A detailed understanding of mechanism involved in ciliary loss and subsequent regeneration is crucial to assess the possible long-term consequences of COVID-19. The aim of this study was to characterize the sequence of histological and ultrastructural changes observed in the ciliated epithelium during and after SARS-CoV-2 infection in the golden Syrian hamster model. We show that acute infection induces a severe, transient loss of cilia, which is, at least in part, caused by cilia internalization. Internalized cilia colocalize with membrane invaginations, facilitating virus entry into the cell. Infection also results in a progressive decline in cells expressing the regulator of ciliogenesis FOXJ1, which persists beyond virus clearance and the termination of inflammatory changes. Ciliary loss triggers the mobilization of p73⁺ and CK14⁺ basal cells, which ceases after regeneration of the cilia. Although ciliation is restored after two weeks despite the lack of FOXJ1, an increased frequency of cilia with ultrastructural alterations indicative of secondary ciliary dyskinesia is observed. In summary, the work provides new insights into SARS-CoV-2 pathogenesis and expands our understanding of virally induced damage to defense mechanisms in the conducting airways.     

Counting Fluorescent Dye Molecules on DNA Origami by Means of Photon Statistics

Obtaining quantitative information about molecular assemblies with high spatial and temporal resolution is a challenging task in fluorescence microscopy. Single‐molecule techniques build on the ability to count molecules one by one. Here, a method is presented that extends recent approaches to analyze the statistics of coincidently emitted photons to enable reliable counting of molecules in the range of 1–20. This method does not require photochemistry such as blinking or bleaching. DNA origami structures are labeled with up to 36 dye molecules as a new evaluation tool to characterize this counting by a photon statistics approach. Labeled DNA origami has a well‐defined labeling stoichiometry and ensures equal brightness for all dyes incorporated. Bias and precision of the estimating algorithm are determined, along with the minimal acquisition time required for robust estimation. Complexes containing up to 18 molecules can be investigated non‐invasively within 150 ms. The method might become a quantifying add‐on for confocal microscopes and could be especially powerful in combination with STED/RESOLFT‐type microscopy.     

Holistic process planning chain for robot machining

Today, machining of large, integral constructed structural parts requires expensive machining centers. In contrast, modern industrial robots are suitable for a wide field of applications and are characterized large working spaces and low capital investment. Therefore, they provide high economical potential for machining applications in aerospace industry, especially for the machining of near to shape pre-products like extruded profiles. However, their constructive characteristics like low stiffness and high sensitivity to vibrations lead to disadvantages compared with conventional machining centers and have to be considered during process planning. Therefore, several methods for offline and online optimization of robot machining processes were developed and integrated in a new process chain for manufacturing of structural fuselage parts. Thereby, the conventional CAD–CAM process planning chain was extended with simulation based analysis and optimization methods and a load-depending trajectory planning. These methods for offline process optimization within this novel process chain are presented in this paper.     

Damage Model and Progressive Failure Analyses for Filament Wound Composite Laminates

Recent improvements in manufacturing processes and materials properties associated with excellent mechanical characteristics and low weight have made composite materials very attractive for application on civil aircraft structures. However, even new designs are still very conservative, because the composite failure phenomenon is very complex. Several failure criteria and theories have been developed to describe the damage process and how it evolves, but the solution of the problem is still open. Moreover, modern filament winding techniques have been used to produce a wide variety of structural shapes not only cylindrical parts, but also “flat” laminates. Therefore, this work presents the development of a damage model and its application to simulate the progressive failure of flat composite laminates made using a filament winding process. The damage model was implemented as a UMAT (User Material Subroutine), in ABAQUS^TM Finite Element (FE) framework. Progressive failure analyses were carried out using FE simulation in order to simulate the failure of flat filament wound composite structures under different loading conditions. In addition, experimental tests were performed in order to identify parameters related to the material model, as well as to evaluate both the potential and the limitations of the model. The difference between numerical and the average experimental results in a four point bending set-up is only 1.6 % at maximum load amplitude. Another important issue is that the model parameters are not so complicated to be identified. This characteristic makes this model very attractive to be applied in an industrial environment.     

Anticipation and flexibility in dynamic scheduling

Many real-world optimization problems change over time and require frequent re-optimization. We suggest that in such environments, an optimization algorithm should reflect the problem's dynamics and explicitly take into account that changes to the current solution are to be expected. We claim that this can be achieved by having the optimization algorithm search for solutions that are not only good, but also flexible, i.e. easily adjustable if necessary in the case of problem changes. For the example of a job-shop with jobs arriving non-deterministically over time, we demonstrate that avoiding early idle times increases flexibility, and thus that the incorporation of an early idle time penalty as secondary objective into the scheduling algorithm can greatly enhance the overall system performance.     

Hydrogenation of p‐Nitrophenol to p‐Aminophenol as a Test Reaction for the Catalytic Activity of Supported Pt Catalysts

Hydrogenation of p‐nitrophenol (PNP) to p‐aminophenol (PAP) using NaBH₄ as a reducing agent was studied as a test reaction for determining the catalytic activity of supported Pt catalysts. The initial reaction rate, which is accessible within less than 10 min via online UV‐vis spectroscopy at room temperature, ambient pressure, and in water as solvent, was applied as measure for catalytic activity. For three Pt catalysts supported on porous SiO₂, porous glass, and Al₂O₃, respectively, significant differences in the catalytic activity by almost one order of magnitude were observed. However, especially in the case of very active catalysts, limitations of the reaction by internal or external mass transfer have to be considered.