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Background: Because adult cardiomyocytes have little regenerative capacity, resident cardiac fibroblasts (CFs) synthesize extracellular matrix after myocardial infarction (MI) to form fibrosis, leading to cardiac dysfunction and heart failure. Therapies that can regenerate the myocardium and reverse fibrosis in chronic MI are lacking. The overexpression of cardiac transcription factors, including Mef2c/Gata4/Tbx5/Hand2 (MGTH), can directly reprogram CFs into induced cardiomyocytes (iCMs) and improve cardiac function under acute MI. However, the ability of in vivo cardiac reprogramming to repair chronic MI with established scars is undetermined. Methods: We generated a novel Tcf21 iCre /reporter/MGTH2A transgenic mouse system in which tamoxifen treatment could induce both MGTH and reporter expression in the resident CFs for cardiac reprogramming and fibroblast lineage tracing. We first tested the efficacy of this transgenic system in vitro and in vivo for acute MI. Next, we analyzed in vivo cardiac reprogramming and fusion events under chronic MI using Tcf21 iCre /Tomato/MGTH2A and Tcf21 iCre /mTmG/MGTH2A mice, respectively. Microarray and single-cell RNA sequencing were performed to determine the mechanism of cardiac repair by in vivo reprogramming. Results: We confirmed the efficacy of transgenic in vitro and in vivo cardiac reprogramming for acute MI. In chronic MI, in vivo cardiac reprogramming converted ≈2% of resident CFs into iCMs, in which a majority of iCMs were generated by means of bona fide cardiac reprogramming rather than by fusion with cardiomyocytes. Cardiac reprogramming significantly improved myocardial contraction and reduced fibrosis in chronic MI. Microarray analyses revealed that the overexpression of MGTH activated cardiac program and concomitantly suppressed fibroblast and inflammatory signatures in chronic MI. Single-cell RNA sequencing demonstrated that resident CFs consisted of 7 subclusters, in which the profibrotic CF population increased under chronic MI. Cardiac reprogramming suppressed fibroblastic gene expression in chronic MI by means of conversion of profibrotic CFs to a quiescent antifibrotic state. MGTH overexpression induced antifibrotic effects partly by suppression of Meox1, a central regulator of fibroblast activation. Conclusions: These results demonstrate that cardiac reprogramming could repair chronic MI by means of myocardial regeneration and reduction of fibrosis. These findings present opportunities for the development of new therapies for chronic MI and heart failure.

With the increasing importance of power storage devices, demand for the development of supercapacitors possessing both rapid reversible chargeability and high energy density is accelerating. Here we propose a simple process for the room temperature fabrication of pseudocapacitor electrodes consisting of a faradaic redox reaction layer on a metallic electrode with an enhanced surface area. As a model metallic electrode, an Au foil was irradiated with Ar

The effectiveness of antibiotics against bacterial infections has been declining due to the emergence of resistance. Precisely understanding the response of bacteria to antibiotics is critical to maximizing antibiotic-induced bacterial eradication while minimizing the emergence of antibiotic resistance. Cell-to-cell heterogeneity in antibiotic susceptibility is observed across various bacterial species for a wide range of antibiotics. Heterogeneity in antibiotic susceptibility is not always due to the genetic differences. Rather, it can be caused by non-genetic mechanisms such as stochastic gene expression and biased partitioning upon cell division. Heterogeneous susceptibility leads to the stochastic growth and death of individual cells and stochastic fluctuations in population size. These fluctuations have important implications for the eradication of bacterial populations and the emergence of genotypic resistance.

Summary Direct cardiac reprogramming holds great potential for regenerative medicine. However, it remains inefficient, and induced cardiomyocytes (iCMs) generated in vitro are less mature than those in vivo, suggesting that undefined extrinsic factors may regulate cardiac reprogramming. Previous in vitro studies mainly used hard polystyrene dishes, yet the effect of substrate rigidity on cardiac reprogramming remains unclear. Thus, we developed a Matrigel-based hydrogel culture system to determine the roles of matrix stiffness and mechanotransduction in cardiac reprogramming. We found that soft matrix comparable with native myocardium promoted the efficiency and quality of cardiac reprogramming. Mechanistically, soft matrix enhanced cardiac reprogramming via inhibition of integrin, Rho/ROCK, actomyosin, and YAP/TAZ signaling and suppression of fibroblast programs, which were activated on rigid substrates. Soft substrate further enhanced cardiac reprogramming with Sendai virus vectors via YAP/TAZ suppression, increasing the reprogramming efficiency up to ∼15%. Thus, mechanotransduction could provide new targets for improving cardiac reprogramming., Graphical Abstract, Highlights • Hydrogel culture reveals the role of mechanotransduction in cardiac reprogramming • Soft ECM comparable with native myocardium promotes cardiac reprogramming • Soft ECM promotes cardiac reprogramming via YAP/TAZ/fibroblast signaling inhibition • Soft ECM promotes Sendai virus vector-mediated cardiac reprogramming, In this article, Ieda and colleagues showed that a soft matrix, which is comparable with native myocardium, efficiently promoted cardiac reprogramming. This soft matrix enhanced cardiac reprogramming via inhibition of integrin, Rho/ROCK, actomyosin, and YAP/TAZ signaling and subsequent suppression of fibroblast programs, which were activated on conventional rigid substrates, thus demonstrating that mechanotransduction plays a critical role in cardiac reprogramming.

The bacterial heat-shock response is regulated by the alternative sigma factor, σ32 (RpoH), which responds to misfolded protein stress and directs the RNA polymerase to the promoters for genes required for protein refolding or degradation. In P. aeruginosa, RpoH is essential for viability under laboratory growth conditions. Here, we used a transcriptomics approach to identify the genes of the RpoH regulon, including RpoH-regulated genes that are essential for P. aeruginosa. We placed the rpoH gene under control of the arabinose-inducible PBAD promoter, then deleted the chromosomal rpoH allele. This allowed transcriptomic analysis of the RpoH (σ32) regulon following a short up-shift in the cellular concentration of RpoH by arabinose addition, in the absence of a sudden change in temperature. The P. aeruginosa ∆rpoH (PBAD-rpoH) strain grew in the absence of arabinose, indicating that some rpoH expression occurred without arabinose induction. When arabinose was added, the rpoH mRNA abundance of P. aeruginosa ∆rpoH (PBAD-rpoH) measured by RT-qPCR increased five-fold within 15 min of arabinose addition. Transcriptome results showed that P. aeruginosa genes required for protein repair or degradation are induced by increased RpoH levels, and that many genes essential for P. aeruginosa growth are induced by RpoH. Other stress response genes induced by RpoH are involved in damaged nucleic acid repair and in amino acid metabolism. Annotation of the hypothetical proteins under RpoH control included proteins that may play a role in antibiotic resistances and in non-ribosomal peptide synthesis. Phenotypic analysis of P. aeruginosa ∆rpoH (PBAD-rpoH) showed that it is impaired in its ability to survive during starvation compared to the wild-type strain. P. aeruginosa ∆rpoH (PBAD-rpoH) also had increased sensitivity to aminoglycoside antibiotics, but not to other classes of antibiotics, whether cultured planktonically or in biofilms. The enhanced aminoglycoside sensitivity of the mutant strain may be due to indirect effects, such as the build-up of toxic misfolded proteins, or to the direct effect of genes, such as aminoglycoside acetyl transferases, that are regulated by RpoH. Overall, the results demonstrate that RpoH regulates genes that are essential for viability of P. aeruginosa, that it protects P. aeruginosa from damage from aminoglycoside antibiotics, and that it is required for survival during nutrient-limiting conditions.

We conducted a numerical simulation of disaster waste disposal by using a Dynamic Hauling/Transportation model (DHT model) in order to extract the factors to hinder the execution of a disposal management plan of Wakayama city government. The DHT model was developed for estimating number of days required for disaster waste disposal, following parameters are mandisopdatory as the model inputs; (1) disaster waste characteristics (distributions, volume and contents) in damaged area, (2) hauling/transportation route, (3) number/capacities of transportation vehicles, (4) maximum capacities at temporary storage / final disposal sites and (5) abilities of crushing/classification equipment. Almost all parameters were set by referring to a planning document of disaster waste disposal provided by Wakayama city and disposal guidelines published by Ministry of the Environment of Japan. Several parameters without description in these document/guideline, e.g. maximum capacities at temporary storage and distance of hauling/transportation route were estimated by using GIS. Severe shock and tsunami caused by Nankai Trough huge earthquake were adopted as external forces generating disaster waste. By assumption that parameters included in (3) and (5) could be critical parameters for a result of simulation, we performed sensitivity analysis of each parameter in them. As a result, abilities of several crushing / classification at temporary storages were the most sensitive parameters of all evaluated parameters. Therefore, augments of crushing/classification functions at temporary storage might be effective to shorten processing days for disaster waste disposal

Adult hearts have limited regenerative capacity. Hence, after acute myocardial infarction (MI), dead myocardial tissues are digested by immune cells and replaced by fibrosis, leading to ventricular remodeling and heart failure at the chronic stage. Direct reprogramming of the cardiac fibroblasts (CFs) into induced cardiomyocytes (iCMs) with cardiac transcription factors, including Gata4, Mef2c, and Tbx5 (GMT), may have significant potential for cardiac repair. Sendai virus (SeV) vectors expressing GMT have been reported to reprogram the mouse cardiac fibroblasts into iCMs without any risk of insertional mutagenesis. In vivo reprogramming improved the cardiac function after acute MI in immunodeficient mice. However, it is unknown whether the newly generated iCMs could exist in infarct hearts for a prolonged period and SeV-GMT can improve cardiac function after MI at the chronic stage in immunocompetent mice. Here, we show that SeV vectors efficiently infect CFs in vivo and reprogram them into iCMs, which existed for at least four weeks after MI, in fibroblast-linage tracing mice. Moreover, SeV-GMT improved cardiac function and reduced fibrosis and collagen I expression at 12 weeks after MI in immunocompetent mice. Thus, direct cardiac reprogramming with SeV vectors could be a promising therapy for MI.

Hintergrund: Die exzessive Inhalation von Aluminiumstaub führt gelegentlich zu einer überwiegend die Oberlappen betreffenden Lungenfibrose, die einer idiopathischen pleuroparenchymalen Fibroelastose (IPPFE) ähnelt und bei der es sich den Annahmen zufolge um eine sekundäre PPFE handelt. Fallbericht: Ein 67-jähriger Mann, der 50 Jahre lang in einem aluminiumverarbeitenden Unternehmen gearbeitet hatte, stellte sich mit Belastungsdyspnoe in unserer Klinik vor. Die Computertomographie (CT) des Thorax zeigte eine bilaterale dichte subpleurale Konsolidierung in den oberen und mittleren Lungenfeldern entsprechend einer IPPFE; eine etwaige sekundäre PPFE im Zusammenhang mit einer Aluminose wurde jedoch nicht ausgeschlossen. Aufgrund des kritischen Zustands des Patienten erfolgten eine transbronchiale Lungenbiopsie (TBLB) anstatt einer chirurgischen Lungenbiopsie sowie eine Elementaranalyse der entnommenen Gewebeprobe. Da die mittels TBLB gewonnene Probe bedauerlicherweise kein Alveolargewebe enthielt, konnte die Diagnose PPFE nicht pathologisch gesichert werden. Die radiologischen Befunde sprachen jedoch stark für eine PPFE. In der Elementaranalyse wurden hohe Mengen an Aluminium in den Bronchiolarwänden nachgewiesen, so dass die Diagnose einer Aluminose der Atemwege mit wahrscheinlich sekundärer PPFE infolge Aluminiumexposition gestellt wurde. Schlussfolgerungen: Eine TBLB mit Elementaranalyse kann zur Abgrenzung der idiopathischen PPFE von sekundären Ursachen bei staubbedingten Erkrankungen wie Aluminose hilfreich sein. Der vorliegende Fall zeigt, dass die Inhalation von Aluminium eine sekundäre PPFE verursachen kann; ein besonderes Augenmerk sollte auf die Vermeidung einer weiteren Exposition gelegt werden.

Pseudomonas aeruginosa biofilm infections are difficult to treat with antibiotic therapy in part because the biofilms contain subpopulations of dormant antibiotic-tolerant cells. The dormant cells can repopulate the biofilms following alleviation of antibiotic treatments. While dormant, the bacteria must maintain cellular integrity, including ribosome abundance, to reinitiate the de novo protein synthesis required for resuscitation. Here, we demonstrate that the P. aeruginosa gene PA4463 [hibernation promoting factor (HPF)], but not the ribosome modulation factor (PA3049), is required for ribosomal RNA preservation during prolonged nutrient starvation conditions. Single-cell–level studies using fluorescence in situ hybridization (FISH) and growth in microfluidic drops demonstrate that, in the absence of hpf, the rRNA abundances of starved cells decrease to levels that cause them to lose their ability to resuscitate from starvation, leaving intact nondividing cells. P. aeruginosa defective in the stringent response also had reduced ability to resuscitate from dormancy. However, FISH analysis of the starved stringent response mutant showed a bimodal response where the individual cells contained either abundant or low ribosome content, compared with the wild-type strain. The results indicate that ribosome maintenance is key for maintaining the ability of P. aeruginosa to resuscitate from starvation-induced dormancy and that HPF is the major factor associated with P. aeruginosa ribosome preservation.

Background Marfan Syndrome (MFS) is a heritable connective tissue disorder with a high degree of clinical variability including respiratory diseases; a rare case of MFS with massive intrathoracic bleeding has been reported recently. Case presentation A 32-year-old man who had been diagnosed with MFS underwent a Bentall operation with artificial valve replacement for aortic dissection and regurgitation of an aortic valve in 2012. Warfarin was started postoperatively, and the dosage was gradually increased until 2017, when the patient was transported to our hospital due to sudden massive haemoptysis. Computed tomography (CT) with a maximum intensity projection (MIP) revealed several giant pulmonary cysts with fluid levels in the apex of the right lung with an abnormal vessel from the right subclavian artery. Transcatheter arterial embolization was performed with angiography and haemostasis was achieved, which suggested that the bleeding vessel was the lateral thoracic artery (LTA) branch. CT taken before the incident indicated thickening of the cystic wall adjacent to the thorax; therefore, it was postulated that the bleeding originated from fragile anastomoses between the LTA and pulmonary or bronchial arteries. It appears that the vessels exhibited inflammation that began postoperatively, which extended to the cysts. Conclusion We experienced a case of MFS with massive haemoptysis from the right LTA. We have to be aware of the possibility that massive haemoptysis could be induced in MFS with inflamed pulmonary cysts.

Background: Pulmonary hypertension (PH) is defined by a resting mean pulmonary artery pressure (mPAP) ≥ 25 mmHg, on the other hand, the term of mPAP ranging from 21 to 24 mmHg is recognized as “borderline PH”. However, little is known about the clinical importance of borderline PH in interstitial lung disease (ILD). Aims: To evaluate whether borderline PH has an impact on the exercise capacity and the risk of acute exacerbation (AE) in patients with ILD. Methods: Between November 2013 and October 2016, 80 patients with ILD (23 IPF) that underwent right heart catheterization were included. The patients were divided into 3 groups: mPAP ≤ 20mmHg (No-PH group; n = 56), 21 ≤ mPAP ≤ 24mmHg (Bo-PH group; n = 18) and mPAP ≥ 25mmHg (PH group; n = 6). Patients’ characteristics, hemodynamics, 6MWT data, and 1-year incidence of AE were collected. Results: There were no significant differences among 3 groups in patients’ characteristics including pulmonary function. However, both Bo-PH and PH groups had significantly lower 6-min walk distance (p Conclusions: The current study shows that borderline PH is associated with exercise intolerance and increased risk for acute exacerbation in patients with ILD. These results suggest that the presence of borderline PH should be considered the pulmonary vascular disorder in patients with ILD.

Bacterial biofilms contain subpopulations of cells that are dormant and highly tolerant to antibiotics. While dormant, the bacteria must maintain the integrity of macromolecules required for resuscitation. Previously, we showed that hibernation promoting factor (HPF) is essential for protecting Pseudomonas aeruginosa from ribosomal loss during dormancy. In this study, we mapped the genetic components required for hpf expression. Using 5ʹ-RACE and fluorescent protein reporter fusions, we show that hpf is expressed as part of the rpoN operon, but that hpf also has a second promoter (P(hpf)) within the rpoN gene. P(hpf) is active when the cells enter stationary phase, and expression from P(hpf) is modulated, but not eliminated, in mutant strains impaired in stationary phase transition (ΔdksA2, ΔrpoS and ΔrelA/ΔspoT mutants). The results of reporter gene studies and mRNA folding predictions indicated that the 5ʹ end of the hpf mRNA may also influence hpf expression. Mutations that opened or that stabilized the mRNA hairpin loop structures strongly influenced the amount of HPF produced. The results demonstrate that hpf is expressed independently of rpoN, and that hpf regulation includes both transcriptional and post-transcriptional processes, allowing the cells to produce sufficient HPF during stationary phase to maintain viability while dormant.

Cytolethal distending toxin B (cdtB) is a conserved virulence factor in Salmonella enterica serovar Typhi. Here we report the presence and functionality of cdtB in some nontyphoidal Salmonella (NTS) serovars, including Salmonella Javiana (cdtB+wt S. Javiana), isolated from imported food. To understand the role of cdtB in NTS serovars, a deletion mutant (cdtB(-)ΔS. Javiana) was constructed. Macrophages were infected with cdtB+wt S. Javiana (wild type), cdtB(-)Δ S. Javiana (mutant), and cdtB-negative NTS serovar (S. Typhimurium). Cytotoxic activity and transcription level of genes involved in cell death (apoptosis, autophagy, and necrosis) were assessed in infected macrophages. The cdtB+wt S. Javiana caused cellular distension as well as high degree of vacuolization and presence of the autophagosome marker LC3 in infected macrophages as compared with cdtB(-)ΔS. Javiana. The mRNA expression of genes involved in the induction of autophagy in response to toxin (Esr1 and Pik3C3) and coregulators of autophagy and apoptosis (Bax and Cyld) were significantly upregulated in cdtB(+)wt S. Javiana-infected macrophages. As autophagy destroys internalized pathogens in addition to the infected cell, it may reduce the spread of infection.

Pseudomonas aeruginosa is an opportunistic human pathogen that causes severe, life-threatening infections in patients with cystic fibrosis (CF), endocarditis, wounds, or artificial implants. During CF pulmonary infections, P. aeruginosa often encounters environments where the levels of calcium (Ca 2+ ) are elevated. Previously, we showed that P. aeruginosa responds to externally added Ca 2+ through enhanced biofilm formation, increased production of several secreted virulence factors, and by developing a transient increase in the intracellular Ca 2+ level, followed by its removal to the basal submicromolar level. However, the molecular mechanisms responsible for regulating Ca 2+ -induced virulence factor production and Ca 2+ homeostasis are not known. Here, we characterized the genome-wide transcriptional response of P. aeruginosa to elevated [Ca 2+ ] in both planktonic cultures and biofilms. Among the genes induced by CaCl 2 in strain PAO1 was an operon containing the two-component regulator PA2656-PA2657 (here called carS and carR ), while the closely related two-component regulators phoPQ and pmrAB were repressed by CaCl 2 addition. To identify the regulatory targets of CarSR, we constructed a deletion mutant of carR and performed transcriptome analysis of the mutant strain at low and high [Ca 2+ ]. Among the genes regulated by CarSR in response to CaCl 2 are the predicted periplasmic OB-fold protein, PA0320 (here called carO ), and the inner membrane-anchored five-bladed β-propeller protein, PA0327 (here called carP ). Mutations in both carO and carP affected Ca 2+ homeostasis, reducing the ability of P. aeruginosa to export excess Ca 2+ . In addition, a mutation in carP had a pleotropic effect in a Ca 2+ -dependent manner, altering swarming motility, pyocyanin production, and tobramycin sensitivity. Overall, the results indicate that the two-component system CarSR is responsible for sensing high levels of external Ca 2+ and responding through its regulatory targets that modulate Ca 2+ homeostasis, surface-associated motility, and the production of the virulence factor pyocyanin. IMPORTANCE During infectious disease, Pseudomonas aeruginosa encounters environments with high calcium (Ca 2+ ) concentrations, yet the cells maintain intracellular Ca 2+ at levels that are orders of magnitude less than that of the external environment. In addition, Ca 2+ signals P. aeruginosa to induce the production of several virulence factors. Compared to eukaryotes, little is known about how bacteria maintain Ca 2+ homeostasis or how Ca 2+ acts as a signal. In this study, we identified a two-component regulatory system in P. aeruginosa PAO1, termed CarRS, that is induced at elevated Ca 2+ levels. CarRS modulates Ca 2+ signaling and Ca 2+ homeostasis through its regulatory targets, CarO and CarP. The results demonstrate that P. aeruginosa uses a two-component regulatory system to sense external Ca 2+ and relays that information for Ca 2+ -dependent cellular processes.

Here, we present the draft genome sequence for the violacein-producing Janthinobacterium sp. CG23_2 isolated from an Antarctic supraglacial stream. The genome is ~7.85 Mb, with a G+C content of 63.5%. The genome includes 7,247 candidate protein coding genes, which may provide insight into UV tolerance mechanisms.

Bacteria have traditionally been studied as single-cell organisms. In laboratory settings, aerobic bacteria are usually cultured in aerated flasks, where the cells are considered essentially homogenous. However, in many natural environments, bacteria and other microorganisms grow in mixed communities, often associated with surfaces. Biofilms are comprised of surface-associated microorganisms, their extracellular matrix material, and environmental chemicals that have adsorbed to the bacteria or their matrix material. While this definition of a biofilm is fairly simple, biofilms are complex and dynamic. Our understanding of the activities of individual biofilm cells and whole biofilm systems has developed rapidly, due in part to advances in molecular, analytical, and imaging tools and the miniaturization of tools designed to characterize biofilms at the enzyme level, cellular level, and systems level.