Effects of sporadic inclusion body myositis on skeletal muscle fibre type specific morphology and markers of regeneration and inflammation.

Sporadic inclusion body myositis (sIBM) is a subgroup of idiopathic inflammatory myopathies characterised by progressive muscle weakness and skeletal muscle inflammation. Quantitative data on the myofibre morphology in sIBM remains scarce. Further, no previous study has examined fibre type association of satellite cells (SC), myonuclei number, macrophages, capillaries, and myonuclear domain (MD) in sIBM patients. Muscle biopsies from sIBM patients (n = 18) obtained previously (NCT02317094) were included in the analysis for fibre type-specific myofibre cross-sectional area (mCSA), SCs, myonuclei and macrophages, myonuclear domain, and capillarisation. mCSA (p < 0.001), peripheral myonuclei (p < 0.001) and MD (p = 0.005) were higher in association with type 1 (slow-twitch) than type 2 (fast-twitch) fibres. Conversely, quiescent SCs (p < 0.001), centrally placed myonuclei (p = 0.03), M1 macrophages (p < 0.002), M2 macrophages (p = 0.013) and capillaries (p < 0.001) were higher at type 2 fibres compared to type 1 fibres. In contrast, proliferating (Pax7+/Ki67+) SCs (p = 0.68) were similarly associated with each fibre type. Type 2 myofibres of late-phase sIBM patients showed marked signs of muscle atrophy (i.e. reduced mCSA) accompanied by higher numbers of associated quiescent SCs, centrally placed myonuclei, macrophages and capillaries compared to type 1 fibres. In contrast, type 1 fibres were suffering from pathological enlargement with larger MDs as well as fewer nuclei and capillaries per area when compared with type 2 fibres. More research is needed to examine to which extent different therapeutic interventions including targeted exercise might alleviate these fibre type-specific characteristics and countermeasure their consequences in impaired functional performance.

Raman spectroscopy and one-dimensional convolutional neural network modeling as a real-time monitoring tool for in vitro transaminase-catalyzed synthesis of a pharmaceutically relevant amine precursor.

Raman spectroscopy has been used to measure the concentration of a pharmaceutically relevant model amine intermediate for positive allosteric modulators of nicotinic acetylcholine receptor in a ω-transaminase-catalyzed conversion. A model based on a one-dimensional convolutional neural network was developed to translate raw data augmented Raman spectra directly into substrate concentrations, with which the conversion from ketone to amine by ω-transaminase could be determined over time. The model showed very good predictive capabilities, with R2 values higher than 0.99 for the spectra included in the modeling and 0.964 for an independent dataset. However, the model could not extrapolate outside the concentrations specified by the model. The presented work shows the potential of Raman spectroscopy as a real-time monitoring tool for biocatalytic reactions.

Lack of muscle stem cell proliferation and myocellular hypertrophy in sIBM patients following blood-flow restricted resistance training.

Sporadic inclusion body myositis (sIBM) is characterised by skeletal muscle inflammation, progressive muscle loss and weakness, which is largely refractory to immunosuppressive treatment. Low-load blood-flow restricted (BFR) training has been shown to evoke gains in myofibre cross sectional area (mCSA) in healthy adults. This could partially be due to the activation and integration of muscle satellite cells (SC) resulting in myonuclei addition. Consequently, this study investigated the effect of 12-weeks lower limb low-load BFR resistance training in sIBM patients on SC and myonuclei content, myofibre size and capillarization. Muscle biopsies from sIBM patients randomised to 12-weeks of low-load BFR resistance training (n = 11) or non-exercising controls (CON) (n = 9) were analysed for SC and myonuclei content, myofibre size and capillarization using three-colour immunofluorescence microscopy and computerised quantification procedures. No between-group differences (time-by-group interactions) or within-groups changes were observed for resident SCs (Pax7+/Six1+), proliferating SCs (Pax7+/ Ki67+), myonuclei (Six1+), type 1 mCSA or capillary number (CD31+). However, a time-by-group interaction for type 2 mCSA was observed (p = 0.04). Satellite cell content, myonuclei number, mCSA and capillary density remained unaffected following 12-weeks low-load BFR resistance training, indicating limited myogenic capacity and satellite cell plasticity in long-term sIBM patients.

SARS-CoV-2 detection using reverse transcription strand invasion based amplification and a portable compact size instrument.

Rapid nucleic-acid based tests that can be performed by non-professionals outside laboratory settings could help the containment of the pandemic SARS-CoV-2 virus and may potentially prevent further widespread lockdowns. Here, we present a novel compact portable detection instrument (the Egoo Health System) for extraction-free detection of SARS-CoV-2 using isothermal reverse transcription strand invasion based amplification (RT-SIBA). The SARS-CoV-2 RT-SIBA assay can be performed directly on crude oropharyngeal swabs without nucleic acid extraction with a reaction time of 30 min. The Egoo Health system uses a capsule system, which is automatically sealed tight in the Egoo instrument after applying the sample, resulting in a closed system optimal for molecular isothermal amplification. The performance of the Egoo Health System is comparable to the PCR instrument with an analytical sensitivity of 25 viral RNA copies per SARS-CoV-2 RT-SIBA reaction and a clinical sensitivity and specificity between 87.0-98.4% and 96.6-98.2% respectively.

Influence of Wearing Ballistic Vests on Physical Performance of Danish Police Officers: A Cross-Over Study.

PURPOSE: We aimed to investigate the influence of wearing a ballistic vest on physical performance in police officers. METHODS: We performed a cross-over study to investigate the influence of wearing a ballistic vest on reaction and response time, lumbar muscle endurance and police vehicle entry and exit times. Reaction and response time was based on a perturbation setup where the officers' pelvises were fixed and EMG of lumbar and abdominal muscles was recorded. We used a modified Biering-Sørensen test to assess the lumbar muscle endurance and measured duration of entry and exit maneuvers in a variety of standard-issue police cars. RESULTS: There was a significant difference of 24% in the lumbar muscle endurance test (no vest: 151 s vs. vest: 117 s), and the police officers experienced higher physical fatigue after the test when wearing a vest. Furthermore, officers took longer to both enter and exit police cars when wearing a vest (range: 0.24-0.56 s) depending on the model of the vehicle. There were no significant differences in reaction and response times between the test conditions (with/without vest). DISCUSSION AND CONCLUSION: Wearing of a ballistic vest significantly influenced the speed of movement in entry and exit of police cars and lumbar muscle endurance, although it does not seem to affect reaction or response times. The ballistic vest seems to impair performance of tasks that require maximal effort, which calls for better designs of such vests.

Towards a risk evaluation of workers' exposure to handborne and airborne microbial species as exemplified with waste collection workers.

Bioaerosol exposure is associated with health problems. The aim of this study is to evaluate whether it is possible to assess the risks posed by waste collection workers' exposure through identification and characterization of bacterial and fungal species, to which the workers are exposed. Using MALDI-TOF MS, microorganisms in waste collection workers' exposure through air, hand, and contact with the steering wheel were identified. Fungi found in high concentrations from the workers' exposure were characterized for the total inflammatory potential (TIP), cytotoxicity, and biofilm-forming capacity. In total, 180 different bacterial and 37 different fungal species in the workers' exposure samples were identified. Some of them belong to Risk Group 2, e.g. Escherichia coli, Klebsiella oxytoca, Staphylococcus aureus, and Aspergillus fumigatus, some have been associated with occupational health problems e.g. Penicillium citrinum and P. glabrum and some are described as emerging pathogens e.g. Aureobasidium pullulans. The TIP of fungal species was dose-dependent. High TIP values were found for Penicillium italicum, P. brevicompactum, P. citrinum, and P. glabrum. Several species were cytotoxic, e.g. A. niger and P. expansum, while some, e.g. P. chrysogenum, did not affect the cell viability. Based on waste workers' average inhalation rate, they inhaled up to 2.3 × 104 cfu of A. niger, 7.4 × 104 cfu of P. expansum, and 4.0 × 106 cfu of P. italicum per work day. Some species e.g. A. niger and P. citrinum were able to form biofilm. In conclusion, the workers were exposed to several species of microorganisms of which some to varying degrees can be evaluated concerning risk. Thus, some microorganisms belong to Risk Group 2, and some are described as causing agents of occupational health problems, emerging pathogens, or intrinsically antibiotic resistant. For some other species very little is known. The TIP, cytotoxicity, and ability to form biofilm of the dominating fungi support and expand previous findings. These parameters depended on the species and the dose, thus highlighting the importance of species identification and exposure level in the risk assessment of exposure.

The immune system in sporadic inclusion body myositis patients is not compromised by blood-flow restricted exercise training.

BACKGROUND: Sporadic inclusion body myositis (sIBM) is clinically characterised by progressive proximal and distal muscle weakness and impaired physical function while skeletal muscle tissue displays abnormal cellular infiltration of T cells, macrophages, and dendritic cells. Only limited knowledge exists about the effects of low-load blood flow restriction exercise in sIBM patients, and its effect on the immunological responses at the myocellular level remains unknown. The present study is the first to investigate the longitudinal effects of low-load blood flow restriction exercise on innate and adaptive immune markers in skeletal muscle from sIBM patients. METHODS: Twenty-two biopsy-validated sIBM patients were randomised into either 12 weeks of low-load blood flow restriction exercise (BFRE) or no exercise (CON). Five patients from the control group completed 12 weeks of BFRE immediately following participation in the 12-week control period leading to an intervention group of 16 patients. Muscle biopsies were obtained from either the m. tibialis anterior or the m. vastus lateralis for evaluation of CD3-, CD8-, CD68-, CD206-, CD244- and FOXP3-positive cells by three-colour immunofluorescence microscopy and Visiopharm-based image analysis quantification. A linear mixed model was used for the statistical analysis. RESULTS: Myocellular infiltration of CD3-/CD8+ expressing natural killer cells increased following BFRE (P < 0.05) with no changes in CON. No changes were observed for CD3+/CD8- or CD3+/CD8+ T cells in BFRE or CON. CD3+/CD244+ T cells decreased in CON, while no changes were observed in BFRE. Pronounced infiltration of M1 pro-inflammatory (CD68+/CD206-) and M2 anti-inflammatory (CD68+/CD206+) macrophages were observed at baseline; however, no longitudinal changes in macrophage content were observed for both groups. CONCLUSIONS: Low-load blood flow restriction exercise elicited an upregulation in CD3-/CD8+ expressing natural killer cell content, which suggests that 12 weeks of BFRE training evokes an amplified immune response in sIBM muscle. However, the observation of no changes in macrophage or T cell infiltration in the BFRE-trained patients indicates that patients with sIBM may engage in this type of exercise with no risk of intensified inflammatory activity.

Palladium-Catalyzed Carbonylative α-Arylation of tert-Butyl Cyanoacetate with (Hetero)aryl Bromides.

A three-component coupling protocol has been developed for the generation of 3-oxo-3-(hetero)arylpropanenitriles via a carbonylative palladium-catalyzed α-arylation of tert-butyl 2-cyanoacetates with (hetero)aryl bromides followed by an acid-mediated decarboxylation step. Through the combination of only a stoichiometric loading of carbon monoxide and mild basic reaction conditions such as MgCl2 and dicyclohexylmethylamine for the deprotonation step, an excellent functional group tolerance was ensured for the methodology. Through the use of (13)C-labeled carbon monoxide generated from (13)COgen, the corresponding (13)C-isotopically labeled ß-ketonitriles were obtained, and these products could subsequently be converted into cyanoalkynes and 3-cyanobenzofurans with site specific (13)C-isotope labeling.

Tailoring on-surface supramolecular architectures based on adenine directed self-assembly.

From an interplay of high-resolution scanning tunneling microscopy (STM) imaging and density functional theory (DFT) calculations we demonstrate that by delicately choosing the parent molecule (adenine) we are able to tune the self-assembled nanostructures of adenine derivatives which are directed by the specific intermolecular interactions provided by the adenine moiety.

Steering supramolecular patterns by nucleobase-terminated molecules.

Supramolecular patterns formed by adsorption from a liquid of nucleobase-terminated molecular rods on a graphite surface were investigated by scanning tunneling microscopy.

Single-molecule chemical reactions on DNA origami.

DNA nanotechnology and particularly DNA origami, in which long, single-stranded DNA molecules are folded into predetermined shapes, can be used to form complex self-assembled nanostructures. Although DNA itself has limited chemical, optical or electronic functionality, DNA nanostructures can serve as templates for building materials with new functional properties. Relatively large nanocomponents such as nanoparticles and biomolecules can also be integrated into DNA nanostructures and imaged. Here, we show that chemical reactions with single molecules can be performed and imaged at a local position on a DNA origami scaffold by atomic force microscopy. The high yields and chemoselectivities of successive cleavage and bond-forming reactions observed in these experiments demonstrate the feasibility of post-assembly chemical modification of DNA nanostructures and their potential use as locally addressable solid supports.

Small molecule induced control in duplex and triplex DNA-directed chemical reactions.

Triplex DNA binders can effectively control copper-catalysed alkyne-azide click reactions in DNA architecture, such that either duplex or triplex DNA directed reactions of terminally attached azides and alkynes occur, in the absence or presence of triplex DNA binder, respectively.

Self-assembly of artificial nucleobase 1H-benzimidazole-4,7-dione at the liquid/solid interface.

Self-assembly at the liquid/solid interface of an electrochemically active DNA nucleobase analogue, 1H-benzoimidazole-4,7-dione (Q), has been studied by means of scanning tunneling microscopy (STM). High-resolution STM images revealed the formation of well-ordered two-dimensional (2D) supramolecular nanostructures when the Q molecules are adsorbed onto the graphite surface from a 1-octanol solution. Detailed analysis shows that the observed 2D nanostructures are mainly dominated by hydrogen-bonded Q molecules. Since Q can be considered as a molecule mimicking the nucleobase guanine (G), which is known to form Watson-Crick base pairs with its complementary nucleobase cytosine (C), we have examined the binding ability of Q with C realized by available hydrogen-bonding sites on both Q and C molecules. Upon deposition of a mixture of Q and C molecules onto a graphite surface, one might expect that hydrogen-bonded QC dimers were observed in a new 2D self-assembled structure governed by inter- and intramolecular hydrogen-bonding interactions between Q and C molecules. However, our STM experiments showed that no well-ordered structures are formed and instead phase separation occurs where large-scale homodomains are formed consisting of the individual QQ and CC dimers. To gain further insight into the possible molecular arrangements of the Q and C nucleobases in the mixture phase, the high-resolution STM images are compared with the results from ab initio density functional theory (DFT) calculations.

Synthesis and electrochemical studies of an anthraquinone-conjugated nucleoside and derived oligonucleotides.

The synthesis of a 2'-deoxyuridine nucleoside linked to an anthraquinone moiety, and its incorporation into oligonucleotides are described, including a facile oxidative demethylation with phenyliodine(iii) bis(trifluoroacetate) to reveal the anthraquinone motif. Furthermore, some useful physical and electrochemical properties of the obtained oligonucleotide are also reported, which allow its principal use in electrochemical DNA assays.

Synthesis of rigid homo- and heteroditopic nucleobase-terminated molecules incorporating adenine and/or thymine.

A series of homo- and heteroditopic thymine- and/or adenine-terminated molecules incorporating rigid aryl or oligo(phenylene ethynylene) linkers has been efficiently synthesized. The key steps involved in the synthesis are the construction of the N-arylated nucleobases using the Chan-Lam-Evans-modified Ullman coupling and their further elaboration using the Sonogashira coupling. Furthermore, the synthesis of a rigid tripodal thymine derivative is reported.

Model systems for activation of nucleic acid encoded prodrugs.

The development of more selective chemotherapeutic agents for benign treatments of malicious diseases is highly desirable. In recent years model systems for the release of small molecule drugs from nucleic acid conjugates by templated chemical or photochemical reactions have been designed. Common for these systems is that the stoichiometric or catalytic drug release is controlled by the highly selective hybridization between complementary strands of nucleic acids. Herein, the concepts of the new field of nucleic acid templated release reactions are outlined.

Efficient N-arylation and N-alkenylation of the five DNA/RNA nucleobases.

A general approach to N-arylation and N-alkenylation of all five DNA/RNA nucleobases at the nitrogen atom normally attached to the sugar moiety in DNA or RNA has been developed. Various protected or masked nucleobases engaged readily in the copper-mediated Chan-Lam-Evans-modified Ullmann condensation with a range of different boronic acids at room temperature and were subsequently converted to the corresponding deprotected or unmasked adducts. Different N(3)-protecting groups were examined in the case of thymine, where the benzoyl group afforded the highest yields. A 4-alkylthio-substituted pyrimidin-2(1H)-one served as both a cytosine and a uracil precursor and was N-arylated and N-alkenylated in high yields. Adenine was efficiently and selectively N-arylated and N-alkenylated at the N(9) position by employing a bis-Boc-protected adenine derivative, while a bis-Boc-protected 2-amino-6-chloropurine served as guanine precursor and could also be selectively N(9)-arylated and N(9)-alkenylated.

The total synthesis of spectinabilin and its biomimetic conversion to SNF4435C and SNF4435D.

[reaction: see text] A short synthesis of (+/-)-spectinabilin via a trans-selective Suzuki coupling and subsequent Negishi-type methylation, and its biomimetic conversion to (+/-)-SNF4435C and (+/-)-SNF4435D is described.