Search

Abstract Anaerobic microbial fermentations provide high product yields and are a cornerstone of industrial bio-based processes. However, the need for redox balancing limits the array of fermentable substrate-product combinations. To overcome this limitation, here we design an aerobic fermentative metabolism that allows the introduction of selected respiratory modules. These can use oxygen to re-balance otherwise unbalanced fermentations, hence achieving controlled respiro-fermentative growth. Following this design, we engineer and characterize an obligate fermentative Escherichia coli strain that aerobically ferments glucose to stoichiometric amounts of lactate. We then re-integrate the quinone-dependent glycerol 3-phosphate dehydrogenase and demonstrate glycerol fermentation to lactate while selectively transferring the surplus of electrons to the respiratory chain. To showcase the potential of this fermentation mode, we direct fermentative flux from glycerol towards isobutanol production. In summary, our design permits using oxygen to selectively re-balance fermentations. This concept is an advance freeing highly efficient microbial fermentation from the limitations imposed by traditional redox balancing.

Abstract Sulfur mustard (SM) is a highly toxic chemical warfare agent. Exposure to SM results in various pathologies including skin lesions with subsequent impaired wound healing. To date, there are no effective treatments available. Here we discover a SM-triggered pathomechanism involving miR-497-5p and its target survivin which contributes to keratinocyte dysfunction. Transcriptome analysis using RNA-seq in normal human epidermal keratinocytes (NHEK) revealed that SM evoked differential expression of 1896 mRNAs and 25 miRNAs with many of these RNAs known to be involved in keratinocyte function and wound healing. We demonstrated that keratinocyte differentiation and proliferation were efficiently regulated by miRNAs induced in skin cells after exposure to SM. The inhibition of miR-497-5p counteracted SM-induced premature differentiation and stimulated proliferation of NHEK. In addition, we showed that microneedle-mediated transdermal application of lipid-nanoparticles containing miR-497-5p inhibitor restored survivin biosynthesis and cellular functionality upon exposure to SM using human skin biopsies. Our findings expand the current understanding of SM-associated molecular toxicology in keratinocytes and highlight miR-497-5p as feasible clinical target for specific skin therapy in SM-exposed patients and beyond.

This interdisciplinary investigation of the human remains of Balduin Helm, one of the most important abbots of the Fürstenfeld monastery, provides novel information on this historic individual. This is particularly interesting since Balduin, during the early 18th century, was involved in the renovation of this large Bavarian monastery. Metastatic prostate cancer was found as evidenced by multiple mixed osteolytic–osteoblastic bone lesions in all available vertebral bodies, fragments of both os coxae, and isolated metastases in skull bones. Distribution, radio- and histomorphology, and especially the immunohistochemical detection of prostate-specific antigens in those metastases definitively confirm this diagnosis. Further investigation, especially by stable isotope analysis, showed a balanced high-level diet with considerable contribution from animal protein and significant freshwater fish. These additional findings suggest a significant radiocarbon reservoir effect as an explanation for a “too old” radiocarbon date. Finally, the obviously high-level protein diet may have contributed to the tumorigenesis which caused the death of the abbot at an advanced age.

Augustinus Bader1, Sabine Ebert1, Shibashish Giri1, Mathias Kremer2, Shuhua Liu2, Andreas Nerlich5, Christina I Günter³, Dagmar U Smith4, Hans-Günther Machens2,31Department of Applied Stem Cell Biology and Cell Techniques, Centre for Biotechnology and Biomedicine, University of Leipzig, Leipzieg, 2Department of Plastic and Hand Surgery, University of Lübeck, Lübeck, 3Department of Plastic and Hand Surgery, Technische Universität München, Munich, 4Münchner Studienzentrum, Technische Universität München, Munich, 5Institute of Pathology, Klinikum München-Bogenhausen, Munich, GermanyBackground: Acceleration of skin regeneration is still an unsolved problem in the clinical treatment of patients suffering from deep burns and scalds. Although erythropoietin (EPO) has a protective role in a wide range of organs and cells during ischemia and after trauma, it has been recently discovered that EPO is not tissue-protective in the common β subunit receptor (βCR) knockout mouse. The protective capacity of EPO in tissue is mediated via a heteroreceptor complex comprising both the erythropoietin receptor (EPOR) and βCR. However, proof of coexpression of these heterogenic receptors in regenerating skin after burns is still lacking.Methods: To understand the role of nanosized recombinant human erythropoietin (rhEPO) in wound healing, we investigated the effects of subcutaneous injections of EPO on skin regeneration after deep second-degree scalding injuries. Our aim was to determine if joint expression of EPOR and βCR is a prerequisite for the tissue-protective effect of rhEPO. The efficiency in wound regeneration in a skin scalding injury mouse model was examined. A deep second-degree dermal scald injury was produced on the backs of 20 female Balb/c mice which were subsequently randomized to four experimental groups, two of which received daily subcutaneous injections of rhEPO. At days 7 and 14, the mice were sacrificed and the effects of rhEPO were analyzed with respect to grade of re-epithelialization (wound closure) and stage of epidermal maturation. This was investigated using different histological parameters of epithelial covering, such as depth of the epidermal layer, epidermal stratification, and presence of conical and hair follicle structures.Results: Expression of EPOR, βCR, and growth hormone receptor at the mRNA and protein levels was demonstrated with reverse transcriptase polymerase chain reaction and Western blot analysis. After rhEPO treatment, the rate of re-epithelialization of the scalding injury was increased and the time to final wound closure was reduced. In addition, the quality of regenerated skin was improved. In this investigation, for the first time, we demonstrated coexpression of EPOR and βCR at the RNA and protein levels in vivo using a deep second-degree scalding injury mouse model. These results highlight the potential role of rhEPO in the improved treatment of burns patients, which might be crucial for the development of innovative new therapy regimes.Conclusion: Local injection of nanosized rhEPO directly to the injury site rather than systemic administration for deep second-degree scalding injuries achieved complete skin regeneration with conical and hair follicle structure via combined expression of EPOR and βCR.Keywords: burns, nanosize, common β subunit, erythropoietin, receptor, local injection

The global push for sustainable energy has heightened the demand for green hydrogen, which is crucial for decarbonizing heavy industry. However, current electrolysis plant capacities are insufficient. This research addresses the challenge through optimizing large-scale electrolysis construction via standardization, modularization, process optimization, and automation. This paper introduces H2Giga, a project for mass-producing electrolyzers, and HyPLANT100, investigating large-scale electrolysis plant structure and construction processes. Modularizing electrolyzers enhances production efficiency and scalability. The integration of AutomationML facilitates seamless information exchange. A digital twin concept enables simulations, optimizations, and error identification before assembly. While construction site automation provides advantages, tasks like connection technologies and handling cables, tubes, and hoses require pre-assembly. This study identifies key tasks suitable for automation and estimating required components. The Enapter Multicore electrolyzer serves as a case study, showcasing robotic technology for tube fittings. In conclusion, this research underscores the significance of standardization, modularization, and automation in boosting the electrolysis production capacity for green hydrogen, contributing to ongoing efforts in decarbonizing the industrial sector and advancing the global energy transition.

The Flood is the first pictorial scene that Michelangelo Buonarroti painted on the ceiling of the Sistine Chapel in the Vatican. On the right side of the fresco a woman with abnormal breast morphology is presented and the nature of her disease is considered using the Guidelines for Iconodiagnosis. A team of experts covering art history, art expertise, medicine, genetics, and pathology undertook the process and concluded that the pathology shown is probably breast cancer, most likely linked to the symbolic significance of an inevitable death as expressed in the Book of Genesis.

In May 2023, The Guardian announced that babies had been born through mitochondrial replacement therapy (MRT). MRT and its ethical and legal implications have been discussed in the media for over a decade. These discussions have been examined by social scientists and communication scholars. In this article, we seek to determine whether and, if so, how social representations have changed since this announcement. Using thematic analysis and social representations theory, we studied a corpus of 70 English language newspaper articles. Results show three thematic dichotomies: (1) moderation vs. sensationalism; (2) ontologising vs. de-ontologising MRT; and (3) using metaphors for persuasion vs. explanation. The societal concern representation largely dominated reporting around 2015 and the hope and opportunity representation was prominent in current reporting. We argue that the hope representation may facilitate greater patient and public acceptance of MRT but that it may also limit awareness, understanding and discussion of its possible risks and limitations.

Poisoning by widow-spider (genus Latrodectus) bites occurs worldwide. The illness, termed latrodectism, can cause severe and persistent pain and can lead to muscle rigidity, respiratory complications, and cardiac problems. It is a global health challenge especially in developing countries. Equine serum-derived polyclonal anti-sera are commercially available as a medication for patients with latrodectism, but the use of sera imposes potential inherent risks related to its animal origin. The treatment may cause allergic reactions in humans (serum sickness), including anaphylactic shock. Furthermore, equine-derived antivenom is observed to have batch-to-batch variability and poor specificity, as it is always an undefined mix of antibodies. Because latrodectism can be extremely painful but is rarely fatal, the use of antivenom is controversial and only a small fraction of patients is treated. In this work, recombinant human antibodies were selected against alpha-latrotoxin of the European black widow (Latrodectus tredecimguttatus) by phage display from a naïve antibody gene library. Alpha-Latrotoxin (α-LTX) binding scFv were recloned and produced as fully human IgG. A novel alamarBlue assay for venom neutralization was developed and used to select neutralizing IgGs. The human antibodies showed in vitro neutralization efficacy both as single antibodies and antibody combinations. This was also confirmed by electrophysiological measurements of neuronal activity in cell culture. The best neutralizing antibodies showed nanomolar affinities. Antibody MRU44–4-A1 showed outstanding neutralization efficacy and affinity to L. tredecimguttatus α-LTX. Interestingly, only two of the neutralizing antibodies showed cross-neutralization of the venom of the Southern black widow (Latrodectus mactans). This was unexpected, because in the current literature the alpha-latrotoxins are described as highly conserved. The here-engineered antibodies are candidates for future development as potential therapeutics and diagnostic tools, as they for the first time would provide unlimited supply of a chemically completely defined drug of constant quality and efficacy, which is also made without the use of animals.

Abstract Background Although histopathological grading systems for disc degeneration are frequently used in research, they are not yet integrated into daily care routine pathology of surgical samples. Therefore, data on histopathological changes in surgically excised disc material and their correlation to clinical parameters such as age, gender or body mass index (BMI) is limited to date. The current study was designed to correlate major physico-clinical parameters from a population of orthopaedic spine center patients (gender, age and BMI) with a quantitative histologic degeneration score (HDS). Methods Excised lumbar disc material from 854 patients (529 men/325 women/mean age 56 (15-96) yrs.) was graded based on a previously validated histologic degeneration score (HDS) in a cohort of surgical disc samples that had been obtained for the treatment of either disc herniation or discogenic back pain. Cases with obvious inflammation, tumor formation or congenital disc pathology were excluded. The degree of histological changes was correlated with sex, age and BMI. Results The HDS (0-15 points) showed significantly higher values in the nucleus pulposus (NP) than in the annulus fibrosus (AF) (Mean: NP 11.45/AF 7.87), with a significantly higher frequency of histomorphological alterations in men in comparison to women. Furthermore, the HDS revealed a positive significant correlation between the BMI and the extent of histological changes. No statistical age relation of the degenerative lesions was seen. Conclusions This study demonstrated that histological disc alterations in surgical specimens can be graded in a reliable manner based on a quantitative histologic degeneration score (HDS). Increased BMI was identified as a positive risk factor for the development of symptomatic, clinically significant disc degeneration.

Abstract Background Curcumin induces apoptosis in many cancer cells and it reduces xenograft growth and the formation of lung metastases in nude mice. Moreover, the plant derived polyphenol has been reported to be able to overcome drug resistance to classical chemotherapy. These features render the drug a promising candidate for tumor therapy especially for cancers known for their high rates concerning therapy resistance like melanoma. Results We show here that the melanoma cell line M14 is resistant to Curcumin induced apoptosis, which correlates with the absence of any effect on NFκB signaling. We show that CXCL1 a chemokine that is down regulated in breast cancer cells by Curcumin in an NFκB dependant manner is expressed at variable levels in human melanomas. Yet in M14 cells, CXCL1 expression did not change upon Curcumin treatment. Following the hypothesis that Curcumin is rapidly removed from the resistant cells, we analyzed expression of known multi drug resistance genes and cellular transporters in M14 melanoma cells and in the Curcumin sensitive breast cancer cell line MDA-MB-231. ATP-binding cassette transporter ABCA1, a gene involved in the cellular lipid removal pathway is over-expressed in resistant M14 melanoma as compared to the sensitive MDA-MB-231 breast cancer cells. Gene silencing of ABCA1 by siRNA sensitizes M14 cells to the apoptotic effect of Curcumin most likely as a result of reduced basal levels of active NFκB. Moreover, ABCA1 silencing alone also induces apoptosis and reduces p65 expression. Conclusion Resistance to Curcumin thus follows classical pathways and ABCA1 expression should be considered as response marker.

Streptococcus canis is a zoonotic agent that causes severe invasive diseases in domestic animals and humans, but little is known about its pathogenesis and virulence mechanisms so far. SCM, the M-like protein expressed by S. canis, is considered one of the major virulence determinants. Here, we report on the two distinct groups of SCM. SCM-1 proteins were already described to interact with its ligands IgG and plasminogen as well as with itself and confer antiphagocytic capability of SCM-1 expressing bacterial isolates. In contrast, the function of SCM-2 type remained unclear to date. Using whole-genome sequencing and subsequent bioinformatics, FACS analysis, fluorescence microscopy and surface plasmon resonance spectrometry, we demonstrate that, although different in amino acid sequence, a selection of diverse SCM-2-type S. canis isolates, phylogenetically representing the full breadth of SCM-2 sequences, were able to bind fibrinogen. Using targeted mutagenesis of an SCM-2 isolate, we further demonstrated that this strain was significantly less able to survive in canine blood. With respect to similar studies showing a correlation between fibrinogen binding and survival in whole blood, we hypothesize that SCM-2 has an important contribution to the pathogenesis of S. canis in the host.

A new mechanism involving intermediate gating states of calcium channels explains how analogue postsynaptic potentials influence neurotransmitter release.

Formation of neutrophil extracellular traps was first described in 2004, showing that NETs are composed of decondensed chromatin fibers and nuclear and granule components. Free DNA is often used to quantify NETs, but to differentiate NETosis from necrotic DNA-release, immunofluorescence microscopy with NET-specific markers is required. Although evaluation by hand is time-consuming and difficult to standardize, it is still widespread. Unfortunately, no standardized method and only limited software tools are available for NET evaluation. This study provides an overview of recent techniques in use and aims to compare two published computer-based methods with hand counting. We found that the selected semi-automated quantification method and fully automated quantification via NETQUANT differed significantly from results obtained by hand and exhibited problems in detection of complex NET structures with partially illogical results. In contrast to that, trained persons were able to adapt to varying settings. Future approaches aimed at developing deep-learning algorithms for fast and reproducible quantification of NETs are needed.

Summary: Direct electrical recordings from conventional boutons in the mammalian central nervous system have proven challenging due to their small size. Here, we provide a protocol for direct whole-cell patch-clamp recordings from small presynaptic boutons of primary dissociated cultured neurons of the rodent neocortex. We describe steps to prepare primary neocortical cultures and recording pipettes, followed by identifying boutons and establishing a whole-cell bouton recording. We then provide details on precise pipette capacitance compensation required for high-resolution current-clamp recordings from boutons.For further details on the use and execution of this protocol, please refer to Ritzau-Jost et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Streptococcus suis is a porcine and zoonotic pathogen in the upper respiratory tract, expressing different capsular serotypes and virulence-associated factors. Given its genomic and phenotypic diversity, the virulence potential of S. suis cannot be attributed to a single factor. Since strong inflammatory response is a hallmark of S. suis infection, the objective of this study was to investigate the differences in transcriptional host responses to two serotype 2 and one serotype 9 strains. Both serotypes are frequently found in clinical isolates. We infected porcine precision-cut lung slices (PCLSs) with two serotype 2 strains of high (strain S10) and low (strain T15) virulence, and a serotype 9 strain 8067 of moderate virulence. We observed higher expression of inflammation-related genes during early infection with strains T15 and 8067, in contrast to infection with strain 10, whose expression peaked late. In addition, bacterial gene expression from infected PCLSs revealed differences, mainly of metabolism-related and certain virulence-associated bacterial genes amongst these strains. We conclude that the strain- and time-dependent induction of genes involved in innate immune response might reflect clinical outcomes of infection in vivo, implying rapid control of infection with less virulent strains compared to the highly virulent strain S10.

Abstract Streptococcus suis, a common member of the porcine respiratory microbiota, can cause life-threatening diseases in pigs as well as humans. A previous study identified the gene trpX as conditionally essential for in vivo survival by intrathecal infection of pigs with a transposon library of S. suis strain 10. Here, we characterized trpX, encoding a putative tryptophan/tyrosine transport system substrate-binding protein, in more detail. We compared growth capacities of the isogenic trpX-deficient mutant derivative strain 10∆trpX with its parent. Growth experiments in chemically defined media (CDM) revealed that growth of 10∆trpX depended on tryptophan concentration, suggesting TrpX involvement in tryptophan uptake. We demonstrated that trpX is part of an operon structure and co-transcribed with two additional genes encoding a putative permease and ATPase, respectively. Bioinformatics analysis identified a putative tryptophan T-box riboswitch in the 5′ untranslated region of this operon. Finally, qRT-PCR and a reporter activation assay revealed trpX mRNA induction under tryptophan-limited conditions. In conclusion, our study showed that TrpX is part of a putative tryptophan ABC transporter system regulated by a T-box riboswitch probably functioning as a substrate-binding protein. Due to the tryptophan auxotrophy of S. suis, TrpX plays a crucial role for metabolic adaptation and growth during infection.

BACKGROUND: We aimed to investigate the self-reported health status and behaviours of 7th-grade adolescents, associations with gender and educational track, as well as health issues addressed during routine school doctor consultations in Switzerland. METHODS: Data on health status and behaviours, specifically general well-being, stimulant and addictive substance use, bullying/violence, exercise, nutrition and health protection, and puberty/sexuality, were drawn from routinely collected self-assessment questionnaires from 1076 (of a total of 1126) students from 14 schools in the Swiss canton of Zug in 2020. Data on health issues addressed in school doctor consultations were collected by nine school doctors (for 595 individual consultations). Multilevel logistic regression analyses were used to investigate the association of gender and educational track with unfavourable health status or behaviours. RESULTS: Although 92% (n = 989) of the students reported being happy or satisfied overall, 21% (n = 215) often or almost always felt sad, and 5-10% had repeatedly been seriously physically hurt (n = 67), sexually harassed with words (n = 88) or experienced uncomfortable physical contact (n = 60). Female gender and a lower educational track were associated with unfavourable health status. In 90% (n = 533) of the school doctor consultations, at least one topic of disease prevention or health promotion was addressed, whereby the topics addressed depended strongly on the individual school doctors. CONCLUSIONS: Our findings revealed that unfavourable health status and behaviours were prevalent among adolescents but the health topics addressed in school doctor consultations were not tailored to students’ self-reported health issues. A school-based approach that strengthens adolescents’ health literacy and provides opportunities for patient-centred counselling has the potential to improve the current and future health of adolescents and, ultimately, adults. To realise this potential, it is essential for school doctors to be sensitised and trained to address students’ health concerns. Emphasis should be placed on the importance of patient-centred counselling, the high prevalence of bullying, and gender and educational differences.

Purpose: Rehabilitation counseling has seen many advances in evidence-based practice in the last decade. The purpose of this study was to provide a review of intervention literature conducted within rehabilitation counseling or related fields from the years 2007-2018. Method: A systematic review of rehabilitation research was conducted, and a total of 163 intervention studies were included in the final sample. Description and analysis include a summary of where and when manuscripts were published, aspects of the intervention (target population, setting, and desired change), and criteria for evaluation of the intervention. Results: Findings show an increase in the use of intervention research during the period studied while also highlighting several patterns in rehabilitation research relating to the targeted samples, outcomes pursued, and the design of research. Conclusions: There is much more intervention research being conducted related to rehabilitation counseling process and outcomes than is demonstrated in rehabilitation counseling journals. This systematic review provides a greater understanding of the progress being made through these international research efforts.

Although common barriers to studying abroad are well documented, this paper proposes a more-overarching barrier is that prevalent evaluation strategies provide little evidence of how study abroad contributes to discipline-specific learning outcomes. Such direct evidence is likely to encourage more employers to view studying abroad as adding value to a graduate resume and hence motivate more students to participate. Studying abroad is often represented in the literature as delivering a transformative impact for all students, regardless of their specific academic programme. While this may be true, it frames the benefits of studying abroad as being generic and supplementary to any discipline-specific curriculum. This paper provides a specific example derived from an Australian national data set and reviews other methodologies that can deliver outcomes data for specific disciplines and for different study programmes, enabling more effective programme evaluation and more effective promotion of the benefits of studying abroad for different students.