Azine‐containing biaryls are ubiquitous scaffolds in many areas of chemistry, and efficient methods for their synthesis are continually desired. Pyridine rings are prominent amongst these motifs. ...Transition‐metal‐catalysed cross‐coupling reactions have been widely used for their synthesis and functionalisation as they often provide a swift and tuneable route to related biaryl scaffolds. However, 2‐pyridine organometallics are capricious coupling partners and 2‐pyridyl boron reagents in particular are notorious for their instability and poor reactivity in Suzuki–Miyaura cross‐coupling reactions. The synthesis of pyridine‐containing biaryls is therefore limited, and methods for the formation of unsymmetrical 2,2′‐bis‐pyridines are scarce. This Review focuses on the methods developed for the challenging coupling of 2‐pyridine nucleophiles with (hetero)aryl electrophiles, and ranges from traditional cross‐coupling processes to alternative nucleophilic reagents and novel main group approaches.
This Review focuses on methods developed as solutions for the challenging coupling of 2‐pyridyl nucleophiles with (hetero)aryl electrophiles. The topic encompasses traditional cross‐coupling processes, modifications to the Suzuki–Miyaura coupling, as well as alternative nucleophilic reagents and recent developments in C−H activation.
OBJECTIVES: To investigate the association between antipsychotic drug use and risk of pneumonia in elderly people.
DESIGN: A nested case‐control analysis.
SETTING: Data were used from the PHARMO ...database, which collates information from community pharmacies and hospital discharge records.
PARTICIPANTS: A cohort of 22,944 elderly people with at least one antipsychotic prescription; 543 cases of hospital admission for pneumonia were identified. Cases were compared with four randomly selected controls matched on index date.
MEASUREMENTS: Antipsychotic drug use in the year before the index date was classified as current, recent, or past use. No prescription for an antipsychotic in the year before the index date was classified as no use. The strength of the association between use of antipsychotics and the development of pneumonia was estimated using multivariate logistic regression analysis and expressed as odds ratios (ORs) with 95% confidence intervals (CIs).
RESULTS: Current use of antipsychotics was associated with an almost 60% increase in the risk of pneumonia (adjusted OR=1.6, 95% CI=1.3–2.1). The risk was highest during the first week after initiation of an antipsychotic (adjusted OR=4.5, 95% CI=2.8–7.3). Similar associations were found after exclusion of elderly people with a diagnosis of delirium. Current users of atypical agents showed a higher risk of pneumonia (adjusted OR=3.1, 95% CI=1.9–5.1) than users of conventional agents (adjusted OR=1.5, 95% CI=1.2–1.9). There was no clear dose‐response relationship.
CONCLUSION: Use of antipsychotics in elderly people is associated with greater risk of pneumonia. This risk is highest shortly after the initiation of treatment, with the greatest increase in risk found for atypical antipsychotics.
Disease-associated genetic variants that lie in non-coding regions found by genome-wide association studies are thought to alter the functionality of transcription regulatory elements and target gene ...expression. To uncover causal genetic variants, variant regulatory elements and their target genes, here we cross-reference human transcriptomic, epigenomic and chromatin conformation datasets. Of 104 genetic variant regions associated with atrial fibrillation candidate target genes are prioritized. We optimize EMERGE enhancer prediction and use accessible chromatin profiles of human atrial cardiomyocytes to more accurately predict cardiac regulatory elements and identify hundreds of sub-threshold variants that co-localize with regulatory elements. Removal of mouse homologues of atrial fibrillation-associated regions in vivo uncovers a distal regulatory region involved in Gja1 (Cx43) expression. Our analyses provide a shortlist of genes likely affected by atrial fibrillation-associated variants and provide variant regulatory elements in each region that link genetic variation and target gene regulation, helping to focus future investigations.
T‐tubules (TT) form a complex network of sarcolemmal membrane invaginations, essential for well‐co‐ordinated excitation–contraction coupling (ECC) and thus homogeneous mechanical activation of ...cardiomyocytes. ECC is initiated by rapid depolarization of the sarcolemmal membrane. Whether TT membrane depolarization is active (local generation of action potentials; AP) or passive (following depolarization of the outer cell surface sarcolemma; SS) has not been experimentally validated in cardiomyocytes. Based on the assessment of ion flux pathways needed for AP generation, we hypothesize that TT are excitable. We therefore explored TT excitability experimentally, using an all‐optical approach to stimulate and record trans‐membrane potential changes in TT that were structurally disconnected, and hence electrically insulated, from the SS membrane by transient osmotic shock. Our results establish that cardiomyocyte TT can generate AP. These AP show electrical features that differ substantially from those observed in SS, consistent with differences in the density of ion channels and transporters in the two different membrane domains. We propose that TT‐generated AP represent a safety mechanism for TT AP propagation and ECC, which may be particularly relevant in pathophysiological settings where morpho‐functional changes reduce the electrical connectivity between SS and TT membranes.
Key points
Cardiomyocytes are characterized by a complex network of membrane invaginations (the T‐tubular system) that propagate action potentials to the core of the cell, causing uniform excitation–contraction coupling across the cell.
In the present study, we investigated whether the T‐tubular system is able to generate action potentials autonomously, rather than following depolarization of the outer cell surface sarcolemma.
For this purpose, we developed a fully optical platform to probe and manipulate the electrical dynamics of subcellular membrane domains.
Our findings demonstrate that T‐tubules are intrinsically excitable, revealing distinct characteristics of self‐generated T‐tubular action potentials.
This active electrical capability would protect cells from voltage drops potentially occurring within the T‐tubular network.
figure legend Methodological framework employed to dissect the electrical properties of isolated t‐tubules (TT). Isolated cardiomyocytes with specific expression of ChR2 (ChR2 CM) are initially stained with a voltage‐sensitive dye (VSD) suitable for two‐photon imaging, thus allowing TT labelling and imaging. Subsequently, TTs are disconnected from the superficial sarcolemma using a formamide‐based osmotic shock, and the electrical activity of isolated TTs is probed using a random‐access two‐photon microscope during optogenetic stimulation. This approach allowed the first recording of self‐generated T‐tubular action potentials (TT AP) in cardiac cells.
Mesenchymal stem cells (MSCs) show unexplained differences in differentiation potential. In this study, differentiation of human (h) MSCs derived from embryonic, fetal and adult sources toward ...cardiomyocytes, endothelial and smooth muscle cells was investigated. Labeled hMSCs derived from embryonic stem cells (hESC-MSCs), fetal umbilical cord, bone marrow, amniotic membrane and adult bone marrow and adipose tissue were co-cultured with neonatal rat cardiomyocytes (nrCMCs) or cardiac fibroblasts (nrCFBs) for 10 days, and also cultured under angiogenic conditions. Cardiomyogenesis was assessed by human-specific immunocytological analysis, whole-cell current-clamp recordings, human-specific qRT-PCR and optical mapping. After co-culture with nrCMCs, significantly more hESC-MSCs than fetal hMSCs stained positive for α-actinin, whereas adult hMSCs stained negative. Furthermore, functional cardiomyogenic differentiation, based on action potential recordings, was shown to occur, but not in adult hMSCs. Of all sources, hESC-MSCs expressed most cardiac-specific genes. hESC-MSCs and fetal hMSCs contained significantly higher basal levels of connexin43 than adult hMSCs and co-culture with nrCMCs increased expression. After co-culture with nrCFBs, hESC-MSCs and fetal hMSCs did not express α-actinin and connexin43 expression was decreased. Conduction velocity (CV) in co-cultures of nrCMCs and hESC-MSCs was significantly higher than in co-cultures with fetal or adult hMSCs. In angiogenesis bioassays, only hESC-MSCs and fetal hMSCs were able to form capillary-like structures, which stained for smooth muscle and endothelial cell markers.Human embryonic and fetal MSCs differentiate toward three different cardiac lineages, in contrast to adult MSCs. Cardiomyogenesis is determined by stimuli from the cellular microenvironment, where connexin43 may play an important role.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Atrial fibrillation (AF) is the most common clinical tachyarrhythmia with a strong tendency to progress in time. AF progression is driven by derailment of protein homeostasis, which ultimately causes ...contractile dysfunction of the atria. Here we report that tachypacing-induced functional loss of atrial cardiomyocytes is precipitated by excessive poly(ADP)-ribose polymerase 1 (PARP1) activation in response to oxidative DNA damage. PARP1-mediated synthesis of ADP-ribose chains in turn depletes nicotinamide adenine dinucleotide (NAD
), induces further DNA damage and contractile dysfunction. Accordingly, NAD
replenishment or PARP1 depletion precludes functional loss. Moreover, inhibition of PARP1 protects against tachypacing-induced NAD
depletion, oxidative stress, DNA damage and contractile dysfunction in atrial cardiomyocytes and Drosophila. Consistently, cardiomyocytes of persistent AF patients show significant DNA damage, which correlates with PARP1 activity. The findings uncover a mechanism by which tachypacing impairs cardiomyocyte function and implicates PARP1 as a possible therapeutic target that may preserve cardiomyocyte function in clinical AF.
Background
Optogenetics could offer a solution to the current lack of an ambulatory method for the rapid automated cardioversion of atrial fibrillation (AF), but key translational aspects remain to ...be studied.
Objective
To investigate whether optogenetic cardioversion of AF is effective in the aged heart and whether sufficient light penetrates the human atrial wall.
Methods
Atria of adult and aged rats were optogenetically modified to express light‐gated ion channels (i.e., red‐activatable channelrhodopsin), followed by AF induction and atrial illumination to determine the effectivity of optogenetic cardioversion. The irradiance level was determined by light transmittance measurements on human atrial tissue.
Results
AF could be effectively terminated in the remodeled atria of aged rats (97%, n = 6). Subsequently, ex vivo experiments using human atrial auricles demonstrated that 565‐nm light pulses at an intensity of 25 mW/mm2 achieved the complete penetration of the atrial wall. Applying such irradiation onto the chest of adult rats resulted in transthoracic atrial illumination as evidenced by the optogenetic cardioversion of AF (90%, n = 4).
Conclusion
Transthoracic optogenetic cardioversion of AF is effective in the aged rat heart using irradiation levels compatible with human atrial transmural light penetration.
Cardiomyocytes (CMs) from human induced pluripotent stem cells (hiPSCs) are functionally immature, but this is improved by incorporation into engineered tissues or forced contraction. Here, we showed ...that tri-cellular combinations of hiPSC-derived CMs, cardiac fibroblasts (CFs), and cardiac endothelial cells also enhance maturation in easily constructed, scaffold-free, three-dimensional microtissues (MTs). hiPSC-CMs in MTs with CFs showed improved sarcomeric structures with T-tubules, enhanced contractility, and mitochondrial respiration and were electrophysiologically more mature than MTs without CFs. Interactions mediating maturation included coupling between hiPSC-CMs and CFs through connexin 43 (CX43) gap junctions and increased intracellular cyclic AMP (cAMP). Scaled production of thousands of hiPSC-MTs was highly reproducible across lines and differentiated cell batches. MTs containing healthy-control hiPSC-CMs but hiPSC-CFs from patients with arrhythmogenic cardiomyopathy strikingly recapitulated features of the disease. Our MT model is thus a simple and versatile platform for modeling multicellular cardiac diseases that will facilitate industry and academic engagement in high-throughput molecular screening.
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•Cardiac fibroblasts and endothelial cells induce hiPSC-cardiomyocyte maturation•CX43 gap junctions form between cardiac fibroblasts and cardiomyocytes•cAMP-pathway activation contributes to hiPSC-cardiomyocyte maturation•Patient-derived hiPSC-cardiac fibroblasts cause arrhythmia in microtissues
Orlova, Bellin, Mummery, and colleagues combined three hiPSC-derived cardiac cell types in 3D microtissues. Cardiomyocytes matured structurally and functionally. Replacing healthy hiPSC-cardiac fibroblasts with patient fibroblasts recapitulated aspects of arrhythmogenic cardiomyopathy. Single-cell transcriptomics, electrophysiology, metabolomics, and ultrastructural analysis revealed roles for CX43 gap junctions and cAMP signaling in the tri-cell-type dialog.
Homologous recombination (HR) is a highly accurate mechanism of DNA repair that can be exploited for homology-directed gene targeting. Since in most cell types HR occurs very infrequently (∼10−6 to ...10−8), its practical application has been largely restricted to specific experimental systems that allow selection of the few cells that become genetically modified. HR-mediated gene targeting has nonetheless revolutionized genetics by greatly facilitating the analysis of mammalian gene function. Recent studies showed that generation of double-strand DNA breaks at specific loci by designed endonucleases greatly increases the rate of homology-directed gene repair. These findings opened new perspectives for HR-based genome editing in higher eukaryotes. Here, we demonstrate by using donor DNA templates together with the adeno-associated virus (AAV) Rep78 and Rep68 proteins that sequence- and strand-specific cleavage at a native, predefined, human locus can also greatly enhance homology-directed gene targeting. Our findings argue for the development of other strategies besides direct induction of double-strand chromosomal breaks to achieve efficient and heritable targeted genetic modification of cells and organisms. Finally, harnessing the cellular HR pathway through Rep-mediated nicking expands the range of strategies that make use of AAV elements to bring about stable genetic modification of human cells.
Anti-Hu antibodies (Hu-Ab) and anti-CV2/CRMP5 antibodies (CV2/CRMP5-Ab) have been identified in association with paraneoplastic neurological disorders. However, it is not clear whether these ...antibodies are associated with specific neurological symptoms or are only markers of anti-cancer immune reaction.
To address this question, 37 patients with CV2/CRMP5-Ab and 324 patients with Hu-Ab were compared.
Whereas the age and sex ratio were the same between the two groups, the distribution of neurological symptoms was not. Patients with CV2/CRMP5-Ab presented more frequently cerebellar ataxia, chorea, uveo/retinal symptoms and myasthenic syndrome (Lambert-Eaton myasthenic syndrome LEMS or myasthenia gravis). They also had a better Rankin score. In contrast, dysautonomia, brainstem encephalitis and peripheral neuropathy were more frequent in patients with Hu-Ab. Limbic encephalitis occurred similarly in both groups. Small-cell lung cancer was the most frequently associated tumour in both groups of patients, while malignant thymoma was observed only in patients with CV2/CRMP5-Ab. In particular, patients with CV2/CRMP5-Ab and thymoma developed myasthenic syndrome more frequently, while patients with SCLC developed neuropathies more frequently. Chorea and myasthenic syndrome were only seen in patients with CV2/CRMP5-Ab. The median survival time was significantly longer in patients with CV2/CRMP5-Ab, and this effect was not dependent on the type of tumour.
The data demonstrate that in patients with paraneoplastic neurological syndromes, the neurological symptoms and survival vary with both the type of associated onco-neural antibody and the type of tumour.
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