Physiological and transcriptomic analysis reveals the toxicological mechanisms of polystyrene micro- and nano-plastics in Chlamydomonas reinhardtii.

With the accumulation of plastic waste in the environment, the toxicity of micro- and nano-plastics (MNPs) to microalgae has attracted increasing attention. However, the underlying toxic mechanisms of MNPs remain to be elucidated. In this study, we synthesized micro- and nano-scale of polystyrene MNPs (PS MNPs) to investigate their toxicity and toxic mechanisms in Chlamydomonas reinhardtii. We found that PS MNPs significantly inhibit the production of photosynthetic pigments and increase soluble protein content. The detailed analysis of results shows that both materials affect photosynthetic efficiency by damaging the donor side, reaction center, and electron transfer of photosystem II. Moreover, compared to PS MPs, PS NPs have a greater negative impact on algal cells. Analyzing the transcriptome of cells suggests that the most sensitive metabolic pathways in response to PS MNPs involve oxidative phosphorylation, biosynthesis of secondary metabolites, and photosynthesis. Especially, genes related to photosynthesis and oxidative phosphorylation showed significant changes in expression after exposure to PS MNPs. This study provided molecular-level insights into the toxic mechanisms of PS MNPs on microalgae.

Deciphering the Epidemiological Characteristics and Molecular Features of bla KPC-2- or bla NDM-1-Positive Klebsiella pneumoniae Isolates in a Newly Established Hospital.

The emergence of hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) was regarded as an emerging threat in clinical settings. Here, we investigated the prevalence of CRKP strains among inpatients in a new hospital over 1 year since its inception with various techniques, and carried out a WGS-based phylogenetic study to dissect the genomic background of these isolates. The genomes of three representative bla NDM-1-positive strains and the plasmids of four bla KPC-2-positive strains were selected for Nanopore long-read sequencing to resolve the complicated MDR structures. Thirty-five CRKP strains were identified from 193 K. pneumoniae isolates, among which 30 strains (85.7%) harbored bla KPC-2, whereas the remaining five strains (14.3%) were positive for bla NDM-1. The antimicrobial resistance profiles of bla NDM-1-positive isolates were narrower than that of bla KPC-2-positive isolates. Five isolates including two bla NDM-1-positive isolates and three bla KPC-2-positive strains could successfully transfer the carbapenem resistance phenotype by conjugation. All CRKP strains were categorized into six known multilocus sequence types, with ST11 being the most prevalent type. Phylogenetic analysis demonstrated that the clonal spread of ST11 bla KPC-2-positive isolates and local polyclonal spread of bla NDM-1-positive isolates have existed in the hospital. The bla NDM-1 gene was located on IncX3, IncFIB/IncHI1B, and IncHI5-like plasmids, of which IncFIB/IncHI1B plasmid has a novel structure. By contrast, all ST11 isolates shared the similar bla KPC-2-bearing plasmid backbone, and 11 of them possessed pLVPK-like plasmids. In addition, in silico virulome analysis, Galleria mellonella larvae infection assay, and siderophore secretion revealed the hypervirulence potential of most bla KPC-2-positive strains. Given that these isolates also had remarkable environmental adaptability, targeted measures should be implemented to prevent the grave consequences caused by hv-CRKP strains in nosocomial settings.

Characterization of a blaNDM-1-Bearing IncHI5-Like Plasmid From Klebsiella pneumoniae of Infant Origin.

The spread of plasmid-mediated carbapenem-resistant clinical isolates is a serious threat to global health. In this study, an emerging NDM-encoding IncHI5-like plasmid from Klebsiella pneumoniae of infant patient origin was characterized, and the plasmid was compared to the available IncHI5-like plasmids to better understand the genetic composition and evolution of this emerging plasmid. Clinical isolate C39 was identified as K. pneumoniae and belonged to the ST37 and KL15 serotype. Whole genome sequencing (WGS) and analysis revealed that it harbored two plasmids, one of which was a large IncHI5-like plasmid pC39-334kb encoding a wide variety of antimicrobial resistance genes clustered in a single multidrug resistance (MDR) region. The blaNDM-1 gene was located on a ΔISAba125-blaNDM-1-bleMBL-trpF-dsbC structure. Comparative genomic analysis showed that it shared a similar backbone with four IncHI5-like plasmids and the IncHI5 plasmid pNDM-1-EC12, and these six plasmids differed from typical IncHI5 plasmids. The replication genes of IncHI5-like plasmids shared 97.06% (repHI5B) and 97.99% (repFIB-like) nucleotide identity with those of IncHI5 plasmids. Given that pNDM-1-EC12 and all IncHI5-like plasmids are closely related genetically, the occurrence of IncHI5-like plasmid is likely associated with the mutation of the replication genes of pNDM-1-EC12-like IncHI5 plasmids. All available IncHI5-like plasmids harbored 262 core genes encoding replication and maintenance functions and carried distinct MDR regions. Furthermore, 80% of them (4/5) were found in K. pneumoniae from Chinese nosocomial settings. To conclude, this study expands our knowledge of the evolution history of IncHI5-like plasmids, and more attention should be paid to track the evolution pathway of them among clinical, animal, and environmental settings.

First case of infective endocarditis caused by Methylobacterium radiotolerans.

Methylobacterium radiotolerans has only been identified in blood samples from end-stage renal failure or leukaemia patients in clinic. Here, we report a case of infective endocarditis (IE) caused by M. radiotolerans. 16S rRNA sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used to identify the bacteria isolated from cardiac vegetation. A drug sensitivity test was conducted by disk diffusion on blood Mueller-Hinton agar. This isolate was identified as M. radiotolerans, which was susceptible to aminoglycosides and ciprofloxacin. Our findings also suggest that M. radiotolerans can cause infection in a patient with normal immune function.

Metformin attenuates angiotensin II-induced TGFß1 expression by targeting hepatocyte nuclear factor-4-α.

BACKGROUND AND PURPOSE: Metformin, a small molecule, antihyperglycaemic agent, is a well-known activator of AMP-activated protein kinase (AMPK) and protects against cardiac fibrosis. However, the underlying mechanisms remain elusive. TGFß1 is a key cytokine mediating cardiac fibrosis. Here, we investigated the effects of metformin on TGFß1 production induced by angiotensin II (AngII) and the underlying mechanisms. EXPERIMENTAL APPROACH: Wild-type and AMPKα2-/- C57BL/6 mice were injected s.c. with metformin or saline and infused with AngII (3 mg·kg-1 ·day-1 ) for 7 days. Adult mouse cardiac fibroblasts (CFs) were isolated for in vitro experiments. KEY RESULTS: In CFs, metformin inhibited AngII-induced TGFß1 expression via AMPK activation. Analysis using bioinformatics predicted a potential hepatocyte nuclear factor 4α (HNF4α)-binding site in the promoter region of the Tgfb1 gene. Overexpressing HNF4α increased TGFß1 expression in CFs. HNF4α siRNA attenuated AngII-induced TGFß1 production and cardiac fibrosis in vitro and in vivo. Metformin inhibited the AngII-induced increases in HNF4α protein expression and binding to the Tgfb1 promoter in CFs. In vivo, metformin blocked the AngII-induced increase in cardiac HNF4α protein levels in wild-type mice but not in AMPKα2-/- mice. Consequently, metformin inhibited AngII-induced TGFß1 production and cardiac fibrosis in wild-type mice but not in AMPKα2-/- mice. CONCLUSIONS AND IMPLICATIONS: HNF4α mediates AngII-induced TGFß1 transcription and cardiac fibrosis. Metformin inhibits AngII-induced HNF4α expression via AMPK activation, thus decreasing TGFß1 transcription and cardiac fibrosis. These findings reveal a novel antifibrotic mechanism of action of metformin and identify HNF4α as a new potential therapeutic target for cardiac fibrosis. LINKED ARTICLES: This article is part of a themed section on Spotlight on Small Molecules in Cardiovascular Diseases. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.8/issuetoc.

[AMP-activated kinase activation inhibits transforming growth factor-ß1 production in cardiac fibroblasts via targeting C/EBPß].

AMP-activated protein kinase (AMPK) activation has been shown to protect against fibrosis. However, the underlying mechanism remains unclear. Here we explored the effect of AMPK activation on transforming growth factor-ß1 (TGFß1) production induced by angiotensin II (AngII) in cardiac fibroblasts and the underlying mechanisms. Adult mouse cardiac fibroblasts were isolated. TGFß1 and AMPK activity were determined by ELISA and Western blots, respectively. Pretreatment of AMPK activator AICAR inhibited TGFß1 production induced by AngII in cardiac fibroblasts, which was reversed by AMPK inhibitor compound C. Furthermore, bioinformatics predicted a potential CCAAT/enhancer-binding protein ß (C/EBPß) binding site in the promoter region of the mouse Tgfb1 gene. Luciferase reporter with wild type, but not deleted, C/EBPß binding sites transfection in mouse embryonic fibroblasts showed increased TGFß1 transcriptional activity induced by AngII, indicating that C/EBPß mediates AngII-induced TGFß1 transcript expression. Pretreatment of AICAR inhibited C/EBPß expression induced by AngII. In conclusion, AMPK activation inhibited TGFß1 production induced by AngII in cardiac fibroblasts through targeting C/EBPß. This finding provides a new mechanism underlying the anti-fibrogenic effects of AMPK activation.

Metformin is a novel suppressor for transforming growth factor (TGF)-ß1.

Metformin is a widely used first-line antidiabetic drug that has been shown to protect against a variety of specific diseases in addition to diabetes, including cardiovascular disorders, polycystic ovary syndrome, and cancer. However, the precise mechanisms underlying the diverse therapeutic effects of metformin remain elusive. Here, we report that transforming growth factor-ß1 (TGF-ß1), which is involved in the pathogenesis of numerous diseases, is a novel target of metformin. Using a surface plasmon resonance-based assay, we identified the direct binding of metformin to TGF-ß1 and found that metformin inhibits [(125)I]-TGF-ß1 binding to its receptor. Furthermore, based on molecular docking and molecular dynamics simulations, metformin was predicted to interact with TGF-ß1 at its receptor-binding domain. Single-molecule force spectroscopy revealed that metformin reduces the binding probability but not the binding force of TGF-ß1 to its type II receptor. Consequently, metformin suppresses type II TGF-ß1 receptor dimerization upon exposure to TGF-ß1, which is essential for downstream signal transduction. Thus, our results indicate that metformin is a novel TGF-ß suppressor with therapeutic potential for numerous diseases in which TGF-ß1 hyperfunction is indicated.

Cardiac Fibrosis Alleviated by Exercise Training Is AMPK-Dependent.

Regular exercise can protect the heart against external stimuli, but the mechanism is not well understood. We determined the role of adenosine monophosphate-activated protein kinase (AMPK) in regulating swimming exercise-mediated cardiac protection against ß-adrenergic receptor overstimulation with isoproterenol (ISO) in mice. Ten-week-old AMPKα2+/+ and AMPKα2-knockout (AMPKα2-/-) littermates were subjected to 4 weeks of swimming training (50 min daily, 6 days a week) or housed under sedentary conditions. The mice received daily subcutaneous injection of ISO (5 mg/kg/d), a nonselective ß-adrenergic receptor agonist, during the last 2 weeks of swimming training. Swimming training alleviated ISO-induced cardiac fibrosis in AMPKα2+/+ mice but not AMPKα2-/- mice. Swimming training activated cardiac AMPK in AMPKα2+/+ mice. Furthermore, swimming training attenuated ISO-induced production of reactive oxygen species (ROS) and expression of NADPH oxidase and promoted the expression of antioxidant enzymes in AMPKα2+/+ mice but not AMPKα2-/- mice. In conclusion, swimming training attenuates ISO-induced cardiac fibrosis by inhibiting the NADPH oxidase-ROS pathway mediated by AMPK activation. Our findings provide a new mechanism for the cardioprotective effects of exercise.