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Abstract Cardiac fibrosis is a common pathological feature of cardiovascular diseases that arises from the hyperactivation of fibroblasts and excessive extracellular matrix (ECM) deposition, leading to impaired cardiac function and potentially heart failure or arrhythmia. Extracellular vesicles (EVs) released by cardiomyocytes (CMs) regulate various physiological functions essential for myocardial homeostasis, which are disrupted in cardiac disease. Therefore, healthy CM‐derived EVs represent a promising cell‐free therapy for the treatment of cardiac fibrosis. To this end, we optimized the culture conditions of human adult CMs to obtain a large yield of EVs without compromising cellular integrity by using a defined combination of small molecules. EVs were isolated by ultracentrifugation, and their characteristics were analysed. Finally, their effect on fibrosis was tested. Treatment of TGFβ‐activated human cardiac fibroblasts with EVs derived from CMs using our culture system resulted in a decrease in fibroblast activation markers and ECM accumulation. The rescued phenotype was associated with specific EV cargo, including multiple myocyte‐specific and antifibrotic microRNAs, although their effect individually was not as effective as the EV treatment. Notably, pathway analysis showed that EV treatment reverted the transcription of activated fibroblasts and decreased several signalling pathways, including MAPK, mTOR, JAK/STAT, TGFβ, and PI3K/Akt, all of which are involved in fibrosis development. Intracardiac injection of CM‐derived EVs in an animal model of cardiac fibrosis reduced fibrotic area and increased angiogenesis, which correlated with improved cardiac function. These findings suggest that EVs derived from human adult CMs may offer a targeted and effective treatment for cardiac fibrosis, owing to their antifibrotic properties and the specificity of cargo.

The rising prevalence of antimicrobial resistance (AMR) of bacteria is a global health problem at the human, animal, and environmental interfaces, which necessitates the "One Health" approach. AMR of bacteria in animal feed are a potential cause of the prevalence in livestock; however, the role remains unclear. To date, there is limited research on AMR of bacteria in animal feed in Japan. In this study, a total of 57 complete feed samples and 275 feed ingredient samples were collected between 2018 and 2020. Enterococcus spp. were present in 82.5% of complete feed (47/57 samples), 76.5% of soybean meal (62/81), 49.6% of fish meal (55/111), 33.3% of poultry meal (22/66), and 47.1% of meat and bone meal (8/17) samples. Of 295 isolates, E. faecium (33.2% of total isolates) was the dominant Enterococcus spp., followed by E. faecalis (14.2%), E. hirae (6.4%), E. durans (2.7%), E. casseliflavus (2.4%), and E. gallinarum (1.0%). Of 134 isolates which were tested for antimicrobial susceptibility, resistance to kanamycin was the highest (26.1%), followed by erythromycin (24.6%), tetracycline (6.0%), lincomycin (2.2%), tylosin (1.5%), gentamicin (0.8%), and ciprofloxacin (0.8%). All Enterococcus spp. exhibited susceptibility to ampicillin, vancomycin, and chloramphenicol. Of 33 erythromycin-resistant isolates, only two showed a high minimum inhibitory concentration value (>128 μg/mL) and possessed ermB . These results revealed that overall resistance to antimicrobials is relatively low; however, animal feed is a source of Enterococcus spp. It is essential to elucidate the causative factors related to the prevalence of AMR in animal feed., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Yamagami, Asao, Takahashi, Hashimoto, Okuyama, Arai, Arihara, Masui and Shimazaki.)

Background: Oral function deterioration is related to a variety of factors, including aging, decline in activities of daily living, malnutrition, and cognitive decline. This cross-sectional study examined the effects of aging on oral function in healthy individuals., Methods: A retrospective study was conducted on 175 healthy, independent patients aged 40-89 years, without dementia and with ≥20 teeth, who visited a local dental clinic in Japan. Patients were compared with 92 university students aged 20-29 years. The seven criteria proposed by the Japanese Society of Gerodontology to diagnose "oral hypofunction" were observed and statistically analyzed., Results: Compared with those in the control group, the degree of tongue coating was increased in the group aged over 80 years, occlusal force was decreased in the group aged 70-79 years, tongue motor function was decreased in the groups aged 60-69 years and older, and tongue pressure was decreased in the groups aged 70-79 years and older., Conclusions: Healthy, independent individuals maintained several oral function criteria across aging, including oral mucosal wetness, occlusal force, lip motor function, masticatory function, and swallowing function. Tongue motor function and tongue pressure decreased with aging, indicating that these may be rehabilitation targets., Competing Interests: The authors declare no conflict of interest.

Rhodoferax antarcticus is an Antarctic purple nonsulfur bacterium and the only characterized anoxygenic phototroph that grows best below 20 °C. We present here a high-quality draft genome of Rfx. antarcticus strain ANT.BR T , isolated from an Antarctic microbial mat. The circular chromosome (3.8 Mbp) of Rfx. antarcticus has a 59.1% guanine + cytosine (GC) content and contains 4036 open reading frames. In addition, the bacterium contains a sizable plasmid (198.6 kbp, 48.4% GC with 226 open reading frames) that comprises about 5% of the total genetic content. Surprisingly, genes encoding light-harvesting complexes 1 and 3 (LH1 and LH3), but not light-harvesting complex 2 (LH2), were identified in the photosynthesis gene cluster of the Rfx. antarcticus genome, a feature that is unique among purple phototrophs. Consistent with physiological studies that showed a strong capacity for nitrogen fixation in Rfx. antarcticus , a nitrogen fixation gene cluster encoding a molybdenum-type nitrogenase was present, but no alternative nitrogenases were identified despite the cold-active phenotype of this phototroph. Genes encoding two forms of ribulose 1,5-bisphosphate carboxylase/oxygenase were present in the Rfx. antarcticus genome, a feature that likely provides autotrophic flexibility under varying environmental conditions. Lastly, genes for assembly of both type IV pili and flagella are present, with the latter showing an unusual degree of clustering. This report represents the first genomic analysis of a psychrophilic anoxygenic phototroph and provides a glimpse of the genetic basis for maintaining a phototrophic lifestyle in a permanently cold, yet highly variable, environment.

The anoxygenic phototroph Rhodobacter sphaeroides uses 3-hydroxypropionate as a sole carbon source for growth. Previously, we showed that the gene (RSP&#95;1434) known as acuI, which encodes a protein of the medium-chain dehydrogenase/reductase (MDR) superfamily, was involved in 3-hydroxypropionate assimilation via the reductive conversion to propionyl-coenzyme A (CoA). Based on these results, we speculated that acuI encoded acrylyl-CoA reductase. In this work, we characterize the in vitro enzyme activity of purified, recombinant AcuI using a coupled spectrophotometric assay. AcuI from R. sphaeroides catalyzes the NADPH-dependent acrylyl-CoA reduction to produce propionyl-CoA. Two other members of the MDR012 family within the MDR superfamily, the products of SPO&#95;1914 from Ruegeria pomeroyi and yhdH from Escherichia coli, were shown to also be part of this new class of NADPH-dependent acrylyl-CoA reductases. The activities of the three enzymes were characterized by an extremely low Km for acrylyl-CoA (<3 μM) and turnover numbers of 45 to 80 s(-1). These homodimeric enzymes were highly specific for NADPH (Km = 18 to 33 μM), with catalytic efficiencies of more than 10-fold higher for NADPH than for NADH. The introduction of codon-optimized SPO&#95;1914 or yhdH into a ΔacuI::kan mutant of R. sphaeroides on a plasmid complemented 3-hydroxypropionate-dependent growth. However, in their native hosts, SPO&#95;1914 and yhdH are believed to function in the metabolism of substrates other than 3-hydroxypropionate, where acrylyl-CoA is an intermediate. Complementation of the ΔacuI::kan mutant phenotype by crotonyl-CoA carboxylase/reductase from R. sphaeroides was attributed to the fact that the enzyme also uses acrylyl-CoA as a substrate.

3-Hydroxypropionate is a product or intermediate of the carbon metabolism of organisms from all three domains of life. However, little is known about how carbon derived from 3-hydroxypropionate is assimilated by organisms that can utilize this C(3) compound as a carbon source. This work uses the model bacterium Rhodobacter sphaeroides to begin to elucidate how 3-hydroxypropionate can be incorporated into cell constituents. To this end, a quantitative assay for 3-hydroxypropionate was developed by using recombinant propionyl coenzyme A (propionyl-CoA) synthase from Chloroflexus aurantiacus. Using this assay, we demonstrate that R. sphaeroides can utilize 3-hydroxypropionate as the sole carbon source and energy source. We establish that acetyl-CoA is not the exclusive entry point for 3-hydroxypropionate into the central carbon metabolism and that the reductive conversion of 3-hydroxypropionate to propionyl-CoA is a necessary route for the assimilation of this molecule by R. sphaeroides. Our conclusion is based on the following findings: (i) crotonyl-CoA carboxylase/reductase, a key enzyme of the ethylmalonyl-CoA pathway for acetyl-CoA assimilation, was not essential for growth with 3-hydroxypropionate, as demonstrated by mutant analyses and enzyme activity measurements; (ii) the reductive conversion of 3-hydroxypropionate or acrylate to propionyl-CoA was detected in cell extracts of R. sphaeroides grown with 3-hydroxypropionate, and both activities were upregulated compared to the activities of succinate-grown cells; and (iii) the inactivation of acuI, encoding a candidate acrylyl-CoA reductase, resulted in a 3-hydroxypropionate-negative growth phenotype.

New classes of repetitive DNA elements were effectively identified by isolating small fragments of the elements from the wheat genome. A wheat A genome library was constructed from Triticum monococcum by degenerate cleavage with EcoO109I, the recognition sites of which consisted of 5'-PuGGNCCPy-3' multi-sequences. Three novel repetitive sequences pTm6, pTm69 and pTm58 derived from the A genome were screened and tested for high copy number using a blotting approach. pTm6 showed identity with integrase domains of the barley Ty1-Copia-retrotransposon BARE-1 and pTm58 showed similarity to the barley Ty3-gypsy-like retrotransposon Romani. pTm69, however, constituted a tandem array with useful genomic specificities, but did not share any identity with known repetitive elements. This study also sought to isolate wheat D-genome-specific repetitive elements regardless of the level of methylation, by genomic subtraction. Total genomic DNA of Aegilops tauschii was cleaved into short fragments with a methylation-insensitive 4 bp cutter, MboI, and then common DNA sequences between Ae. tauschii and Triticum turgidum were subtracted by annealing with excess T. turgidum genomic DNA. The D genome repetitive sequence pAt1 was isolated and used to identify an additional novel repetitive sequence family from wheat bacterial artificial chromosomes with a size range of 1 395-1 850 bp. The methods successfully led pathfinding of two unique repetitive families.

Regurgitant flows in valvular diseases were evaluated by a pulsed Doppler flowmeter combined with an electronic beam sector scanning echocardiograph. The apparatus which was newly developed by us allowed the simultaneous demonstration of a sample site on a two-dimensional echocardiogram with flow measurement. Doppler signals of regurgitant flow were recorded as uni-directional or bi-directional wide frequency band signals. The locations, where regurgitant flow signals were detected, were depicted on the corresponding two-dimensional echocardiogram. This procedure was referred to as "a flow mapping technique" for non-invasive visualization of the distribution of regurgitant flow. In 12 patients with mitral regurgitation due to mitral valve prolapse detected by the pulsed Doppler technique, the regurgitant flow was distributed to the opposite side of the prolapsing mitral leaflet. The transmission of the regurgitant murmur was well consistent with the direction of the regurgitant flow. In 14 patients with aortic regurgitation, the distribution of aortic regurgitant flow visualized by the flow mapping technique closely coincided with that obtained by cineaortography. Based on the distribution of the regurgitation, the severity of the regurgitation could be precisely evaluated by the Doppler technique. Regurgitant flow signals were detected in the right atrium in all 13 patients with tricuspid regurgitation diagnosed by right ventriculography. We found 4 patients who did not show Carvallo's sign but in whom were detected regurgitant flow signals by the Doppler technique. In all of them, tricuspid regurgitation was proven at surgery. These results indicate that the Doppler technique presented here has an obvious clinical advantage in detecting and evaluating regurgitant flow in valvular diseases.

1) Intracardiac flows were investigated by bi-directional pulsed Doppler technique with a combined use of the two-dimensional echocardiography. The flow at sampling sites in the heat and great vessels were correctly identified on the two-dimensional echocardiograms. 2) Soundspectrographic analysis of the flow Doppler signals was useful in obtaining information about flow modes and in differentiating flow signals from those of valves. 3) In healthy subjects, intracardiac flow revealed a laminar mode. As a matter of interest, a flow toward the aorta, which is a cranially turned flow, was found at the center and at the outflow tract of the left ventricle from mid to late diastole. 4) In mitral stenosis, the central stream at the stenotic portion exhibited an almost laminar and rapid flow. A good coincidence in flow mode was observed between clinical cases of mitral stenosis and experimental stenosis. 5) Disturbed flows were observed in the following diseased conditions. The diastolic regurgitant flow in the left ventricular outflow tract and mitral inflow in aortic regurgitation, the regurgitant flow into the left atrium and the diastolic mitral inflow in mitral regurgitation, the ejection flow in hypertrophic cardiomyopathy with outflow tract obstruction, and the tricuspid inflow in atrial septal defect. 6) Mechanisms of intracardiac flow dynamics were also discussed.

In this study, transmitral flow velocity during the diastolic period was non-invasively measured to assess diastolic behavior of the left ventricle by pulsed Doppler flowmetry combined with electronic beam sector-scanning echocardiography. The velocity pattern was found to have 2 wave components: one appears in the early diastolic rapid filling phase (R wave) and the other in the late diastolic phase. The peak of the early diastolic inflow velocity (peak EFV), the deceleration rate of the R wave (DC), and the peak of the late diastolic inflow velocity (peak LFV) were compared in healthy subjects as the control, patients with hypertension, hypertrophic cardiomyopathy and definite old myocardial infarction. Normal peak EFV and DC, 61.3 +/- 6.7 cm/sec and 355 +/- 67 cm/sec2, respectively, were markedly reduced in patients with hypertension (50.0 +/- 10.0 cm/sec and 265 +/- 75 cm/sec2), hypertrophic cardiomyopathy (48.8 +/- 10.7 cm/sec and 205 +/- 78 cm/sec2), and myocardial infarction (46.1 +/- 12.0 cm/sec and 240 +/- 84 cm/sec2). Among all disease groups the DC was most significantly reduced in the hypertrophic cardiomyopathy groups. Normal peak LFV, 39.9 +/- 11.0 cm/sec, was significantly increased in patients with hypertension (54.3 +/- 10.7 cm/sec) and myocardial infarction (50.0 +/- 11.0 cm/sec), but not in patients with hypertrophic cardiomyopathy (42.0 +/- 8.4 cm/sec). The ratio peak LFV/peak EFV was significantly greater in all disease groups than in the normal group. These findings indicate that impaired early diastolic filling in all disease groups was compensated by enhanced atrial contraction in patients with hypertension and myocardial infarction, and by prolonged rapid filling in patients with hypertrophic cardiomyopathy. Left ventricular wall thickness was better correlated with DC in patients with hypertension (r = -0.76, p less than 0.01) than in patients in the other groups, which indicates that the left ventricular wall thickening is a more important factor in determining diastolic behavior of the ventricle in hypertension than in hypertrophic cardiomyopathy and myocardial infarction. Thus, the pulsed Doppler technique was proved to be useful in assessing ventricular diastolic events non-invasively.