Search

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

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

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.

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.

Dissemination of plasmid-mediated quinolone resistance among pathogenic bacteria is a concern for public health because of decreased sensitivity to fluoroquinolones and increased potentials to develop high fluoroquinolone resistance. Two qnrS1 -positive isolates of Salmonella enterica Corvallis (468) and Typhimurium (484) from imported seafood (Thailand and Vietnam) were tested for quinolone sensitivity using disk agar diffusion and the Sensititre® system. The presence of qnr genes, qnr carrying plasmids, and mutations in the quinolone resistance determining regions were also determined. Minimal inhibitory concentrations of nalidixic acid for isolates 468 and 484 were 8 and 16 µg mL−1, respectively, and those of ciprofloxacin were 1 and 2 µg mL−1, respectively. Disk agar diffusion indicated that isolate 468 was moderately resistant to moxifloxacin, and isolate 484 was resistant to moxifloxacin and moderately resistant to norfloxacin. Isolates 468 and 484 carried a mutation on parC , but not on gyrA , gyrB , or parE . Sequences of qnrS1 -carrying plasmids from isolates 468 and 484, sized 10 039 and 10 047 bp, were nearly identical (> 99% similarity) to each other and to published sequences of plasmids from clinical isolates of Salmonella Typhimurium isolated in the United Kingdom and Taiwan, indicating a dissemination of qnrS1 -carrying plasmids among different serovars of Salmonella from geographically separated sources. This is the first complete sequence of a qnrS1 -carrying plasmid from imported seafood isolate of S. enterica .

血液透析導入時にヘパリン起因性血小板減少症（HIT）を発症し，その後ヘパリンの再投与に成功した症例を経験したので報告する．症例は68歳女性．以前より慢性腎不全を指摘されており，近医に通院していた．末期腎不全にて当院紹介受診し，血液透析導入目的で入院した．入院後，内シャント造設術を施行した．シャント血流は良好であった．透析の抗凝固剤として未分画ヘパリンを使用した．第1回透析から第3回透析までは著変なく施行できた．第4回，第5回透析では若干の静脈圧の上昇を認めたが，明らかな回路内凝固は認めなかった．第6回透析では静脈圧の上昇とチャンバー・ダイアライザーの凝血を認めた．ヘパリン量の不足が原因と考え，ヘパリンの増量を行ったが改善を認めなかった．透析前後で血小板数の減少があり，HITを疑い，抗血小板第4因子/ヘパリン複合体抗体（HIT抗体）を測定した．抗凝固剤をメシル酸ナファモスタットに変更後，回路内凝固と血小板減少は改善した．HIT抗体が陽性であったため，HITと診断し，抗凝固剤をアルガトロバンに変更し，開始時10mg，持続25mg/時で血液透析を行った．以降は回路内凝固を認めず，また，血小板減少も認めなかった．その間，血栓症の合併はなかった．経時的にHIT抗体価を測定し，HIT抗体価が陰性化したことを確認した．ヘパリンの再投与を試みたところ，回路内凝固や血小板減少を認めることなく，血液透析を施行することができた．HIT抗体価の再上昇もなく，現在に至っている．

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.

Bacteroidales are fecal anaerobic bacteria that are common in the digestive systems and feces of warm-blooded animals. Some strains of Bacteroidales have been reported to be host-specific. In this study, Bacteroidales strains from chicken feces were examined for their potential use as indicators of chicken fecal contamination. Bacteroidales 16S rRNA gene sequences from chicken feces were amplified, cloned and sequenced. Phylogenetic analysis was performed using these sequences and published Bacteroidales 16S rRNA gene sequences from human and bovine feces. Primers were designed based on putative chicken feces-specific 16S rRNA gene sequences and the primer pairs were tested for specificity in PCR assays. One set of primers, chBact F1 and chBact R16, specifically amplified DNA from chicken feces in a PCR assay, but did not amplify wild turkey, cat, bovine, or deer fecal DNAs. In addition, DNA from feces contaminated straw-based chicken litter produced a product in the PCR assay. However, DNA from feces contaminated wood shavings-based chicken litter was not amplified. The PCR assay described here may prove a useful tool for the detection of chicken feces and for source tracking in watersheds with fecal contamination.

維持血液透析患者66例について, 新たな動脈硬化性物質として注目されているhomocysteineおよびメチオニン代謝系基質である葉酸の血中濃度を測定した. 血漿homocysteine濃度は12.6-182.0μmol/l (平均45.6±31.9μmol/l) と全例で高値を示すと同時に著しい個体差を認めた. 一方, 血清葉酸濃度は2.7-26.8ng/ml (平均7.5±4.1ng/ml) とほとんどの症例で基準値の範囲内にあった. また, 血清葉酸濃度と血漿homocysteine濃度には負の相関 (r=-0.245; P=0.0445) が認められた. さらに, 血清葉酸濃度15ng/ml以上では40μmol/l以上の高度のhomocysteine高値例は認められなかった. しかし, 通常の食生活で血清葉酸濃度が15ng/ml以上に維持できている症例は3例 (4.5%) に過ぎず, 残りの63例 (95.5%) は血清葉酸濃度15ng/ml以下であった. 今回の結果からacquired hyperhomocysteinemiaの状態にある維持血液透析患者においては通常の食生活のみでは血漿homocysteine濃度を正常に保つに十分な葉酸濃度を得ることはできず, 動脈硬化予防のためには積極的な葉酸投与の必要性が示唆された.

Here we present the draft genome sequence of Janthinobacterium sp. strain CG3, a psychrotolerant non-violacein-producing bacterium that was isolated from the Cotton Glacier supraglacial stream. The genome sequence of this organism will provide insight as to the mechanisms necessary for bacteria to survive in UV-stressed icy environments.

To determine the fluoroquinolone resistance determinants in Salmonella enterica serovar Schwarzengrund from imported foods.Antibiotic susceptibility of Salmonella Schwarzengrund to 16 antibiotics was examined using disc agar diffusion and Etest. Quinolone resistance determinants were examined by sequence analysis of gyrA, gyrB, parC and parE, PCR amplification of qnrA, qnrB and qnrS, and expression of acrB, ramA, marA, soxS and rob using quantitative RT-PCR. The contribution of efflux pump activities to antibiotic resistance was determined by the addition of carbonyl cyanide m-chlorophenylhydrazone (CCCP). The effect of ramR deletion on ciprofloxacin resistance was determined by complementing with wild-type ramR.Salmonella strains 30 and 487 were susceptible to ciprofloxacin and had a single mutation in gyrA as compared with strain 75, which was highly resistant to ciprofloxacin and had a double mutation in gyrA. Increased expression of ramA was associated with high resistance to ciprofloxacin. Strain 75 had a deletion of 315 bp in the ramR gene, which regulates ramA expression. Overexpression of ramA was possibly related to a loss of ramR. Introduction of ramR decreased the MIC of ciprofloxacin from 48 to 24 mg/L. The addition of CCCP did not reduce antibiotic resistance. To our knowledge, this study reports for the first time the natural deletion of ramR in Salmonella Schwarzengrund.This study indicates that fluoroquinolone-resistant Salmonella are prevalent in imported food. Double mutations in gyrA and a loss of ramR were associated with high-level quinolone resistance in multidrug-resistant Salmonella Schwarzengrund strain 75.