Sequestration and efflux largely account for cadmium and copper resistance in the deep-sea Nitratiruptor sp. SB155-2 (phylum Campylobacterota).

In deep-sea hydrothermal vent environments, metal-enriched fluids and sediments abound, making these habitats ideal to study metal resistance in prokaryotes. In this investigation, we employed transcriptomics and shotgun proteomics with scanning transmission electron microscopy and energy-dispersive x-ray spectroscopy (STEM-EDX) to better understand mechanisms of tolerance for cadmium (Cd) and copper (Cu) at stress-inducing concentrations in Nitratiruptor sp. SB155-2 (phylum Campylobacterota). Transcriptomic profiles were remarkably different in the presence of these two metals, displaying 385 (19%) and 629 (31%) differentially transcribed genes (DTG) in the presence of Cd(II) and Cu(II), respectively, while only 7% of differentially transcribed (DT) genes were shared, with genes for non-specific metal transporters and genes involved in oxidative stress-response predominating. Transcriptomic and proteomic analyses confirmed that metal-specific DT pathways under Cu(II) stress, including those involving sulfur, cysteine, and methionine, are likely required for high-affinity efflux systems, while flagella formation and chemotaxis were over-represented under Cd(II) stress. Consistent with these differences, STEM-EDX analysis revealed that polyphosphate-like granules (pPLG), the formation of CdS particles, and the periplasmic space are crucial for Cd(II) sequestration. Overall, this study provides new insights regarding metal-specific adaptations of Campylobacterota to deep-sea hydrothermal vent environments.

Robust Packing of a Self-Assembling Iridium Complex via Endocytic Trafficking for Long-Term Lysosome Tracking.

Live cell imaging of lysosome positioning and motility is critical to studying lysosome status and function for pharmacological interventions. To create a super stable lysosomal probe for long-term live cell imaging, we have designed and synthesized an aromatic-peptide-conjugated cyclometalated iridium(III) complex that emits light via π-π stacking oriented self-assembly in water at extremely low concentration. Through endocytic trafficking, self-assemblies are transformed from nanoparticles into sturdily packed networks that are stabilized in lysosomal acidic environment. Upon short time/low dose treatment of the iridium complex at passage 0, live cell lysosomal tracking is applicable beyond the 14th passage of cells with high labelling rate and a mild decline in luminescence intensity. The illuminated lysosomes are trackable using super-resolution imaging to study their response to cellular processes.

In situ investigation of oxidation across a heterogeneous nanoparticle-support interface during metal support interactions.

Interactions between oxide supports and noble metal nanoparticles (NPs) is an area of intense research interest across all fields of catalysis. Oxygen spillover, metal support interactions (MSIs) and charge transfer are among many mechanisms observed and proposed as to how NP-support interfaces assist and enhance catalysis. This work studies the migration of oxygen across the Pd NP-CuO nanowire (NW) interface and beyond. X-ray photoelectron spectroscopy (XPS) and Kelvin probe force microscopy (KPFM) found an interaction between the Pd NP and CuO NW support, via the formation of PdO at the Pd-CuO interface. It was found, through in situ irradiation at high vacuum transmission electron microscopy (TEM), that oxygen enters the Pd NP lattice from the Pd-CuO interface via amorphization of the NP. Varying the amount of irradiation highlighted the different rates of amorphization of NPs, with full amorphization of a NP leading to the formation of an epitaxially driven PdO across the NPs. Interestingly, in situ heating in XPS observed a reduction to metallic Pd, found to be similarly amorphous during TEM investigation. On comparison with Pd supported on a non-reducible substrate - in which oxidation was found to proceed from the outer surface in, rather than the support interface (resulting in a PdO shell) - it is theorized that the oxidation and reduction of Pd on CuO forms a PdO NP surface full of Pd-PdO sites allowing for synergistic effects, of great use in the oxidation and hydrogenation of organic species.

Defect-assisted electronic metal-support interactions: tuning the interplay between Ru nanoparticles and CuO supports for pH-neutral oxygen evolution.

Electronic metal-support interactions (EMSIs) comprise an area of intense study, the manipulation of which is of paramount importance in the improvement of heterogeneous metal nanoparticle (NP) supported catalysts. EMSI is the transfer of charge from the support to NP, enabling more effective adsorption and interaction of reactants during catalysis. Ru NPs on CuO supports show different levels of EMSI (via charge transfer) depending on their crystal structure, with multiple twinned NPs showing greater potential for EMSI. We use magnetron-assisted gas phase aggregation for the synthesis of batches of Ru NPs with different populations of single crystal and multiple twinned nanoparticles, which were deposited on CuO nanowires (NWs). The surface charging of the Ru-CuO catalysts was investigated by Kelvin probe force microscopy (KPFM) and X-ray photoelectron spectroscopy (XPS). By doubling the population of multiple twinned NPs, the surface potential of the Ru-CuO catalysts increases roughly 4 times, coinciding with a similar increase in the amount of Ru4+. Therefore, tuning the amount of EMSI in a catalyst is possible through changing the population of multiple twinned Ru NPs in the catalyst. Increasing the amount of multiple twin NPs resulted in improved activity in the oxygen evolution reaction (a roughly 2.5 times decrease in the overpotentials when the population of multiple twinned NPs is increased) and better catalyst stability. This improvement is attributed to the fact that the multiple twin NPs maintained a metallic character under oxidation conditions (unlike single crystal NPs) due to the EMSI between the NP and support.

Hidden proteome of synaptic vesicles in the mammalian brain.

Current proteomic studies clarified canonical synaptic proteins that are common to many types of synapses. However, proteins of diversified functions in a subset of synapses are largely hidden because of their low abundance or structural similarities to abundant proteins. To overcome this limitation, we have developed an "ultra-definition" (UD) subcellular proteomic workflow. Using purified synaptic vesicle (SV) fraction from rat brain, we identified 1,466 proteins, three times more than reported previously. This refined proteome includes all canonical SV proteins, as well as numerous proteins of low abundance, many of which were hitherto undetected. Comparison of UD quantifications between SV and synaptosomal fractions has enabled us to distinguish SV-resident proteins from potential SV-visitor proteins. We found 134 SV residents, of which 86 are present in an average copy number per SV of less than one, including vesicular transporters of nonubiquitous neurotransmitters in the brain. We provide a fully annotated resource of all categorized SV-resident and potential SV-visitor proteins, which can be utilized to drive novel functional studies, as we characterized here Aak1 as a regulator of synaptic transmission. Moreover, proteins in the SV fraction are associated with more than 200 distinct brain diseases. Remarkably, a majority of these proteins was found in the low-abundance proteome range, highlighting its pathological significance. Our deep SV proteome will provide a fundamental resource for a variety of future investigations on the function of synapses in health and disease.

Utilizing ballistic nanoparticle impact to reconfigure the metal support interaction in Pt-TiN electrocatalysts.

Tuning the metal support interaction (MSI) in heterogeneous catalysts is of utmost importance for various applications in different catalysis reactions. Pt-TiN systems are strong contenders for commercial catalysts, although the charge screening of Pt and non-involvement of N reduces their effective MSI and limits it to the Pt-Ti interface. Here, the bias driven landing of gas phase synthesized Pt nanoparticles (NPs) is used to change the nature of the MSI and enhance the charge transfer phenomenon. Bias driven landing of the Pt NPs translates their impact energies to the TiN surface, resulting in a weakening of the Ti-N bonds. This facilitates a new interaction between the Pt and N atoms, resulting in an electronic equilibration in the N-Pt-Ti triumvirate, nullifying the charge screening of Pt. This change in the nature of the MSI enables long range charge transfer throughout the catalyst surface and an increase in the electrocatalytic hydrogen evolution reaction (HER) activity of the Pt-TiN system.

Functionalized mesoporous silica nanoparticles for innovative boron-neutron capture therapy of resistant cancers.

Treatment resistance, relapse and metastasis remain critical issues in some challenging cancers, such as chondrosarcomas. Boron-neutron capture therapy (BNCT) is a targeted radiation therapy modality that relies on the ability of boron atoms to capture low energy neutrons, yielding high linear energy transfer alpha particles. We have developed an innovative boron-delivery system for BNCT, composed of multifunctional fluorescent mesoporous silica nanoparticles (B-MSNs), grafted with an activatable cell penetrating peptide (ACPP) for improved penetration in tumors and with gadolinium for magnetic resonance imaging (MRI) in vivo. Chondrosarcoma cells were exposed in vitro to an epithermal neutron beam after B-MSNs administration. BNCT beam exposure successfully induced DNA damage and cell death, including in radio-resistant ALDH+ cancer stem cells (CSCs), suggesting that BNCT using this system might be a suitable treatment modality for chondrosarcoma or other hard-to-treat cancers.

Mechanoresponsive Alignment of Molecular Self-Assembled Negatively Charged Nanofibrils.

Inspired by the mechanoresponsive orientation of actin filaments in cell, we introduce a design paradigm of synthetic molecular self-assembling fibrils that respond to external mechanical force by transforming from a macroscopically disorder state to a highly ordered uniaxial aligned state. The incorporation of aromatic-containing amino acids and negatively charged amino acids lead to self-assembly motifs that transform into uniform nanofibrils in acidic solution. Adjusting the pH level of aqueous solution introduces optimal negative charge to the surface of self-assembling nanofibrils inducing long-range electrostatic repulsion forming a nematic phase. Upon external mechanical force, nanofibrils align in the force direction. Via evaporation casting in capillary confinement, the solvated synthetic self-assembling nanofibrils transform into scalable lamellar domains. Adjusting capillary geometry and drying procedure offers further parameters for tuning the mesoscale alignment of nanofibrils generating a variety of interference colors. The design paradigm of mechanoresponsive alignment of self-assembled nanofibrils as an addition of nanofabrication techniques is potentially employable for realizing biomimetic optical structures.

Self-delivery of N-hydroxylethyl peptide assemblies to the cytosol inducing endoplasmic reticulum dilation in cancer cells.

Inspired by clinical studies on alcohol abuse induced endoplasmic reticulum disruption, we designed a N-hydroxylethyl peptide assembly to regulate the ER stress response in cancer cells. Upon coupling with a coumarin derivative via an ester linkage, a prodrug was synthesized to promote esterase-facilitated self-delivery of N-hydroxylethyl peptide assemblies around the ER, inducing ER dilation. Following this, ER-specific apoptosis was effectively and efficiently activated in various types of cancer cells including drug resistant and metastatic ones.

Self-Assembly-Directed Cancer Cell Membrane Insertion of Synthetic Analogues for Permeability Alteration.

Inspired by the metamorphosis of pore-forming toxins from soluble inactive monomers to cytolytic transmembrane assemblies, we developed self-assembly-directed membrane insertion of synthetic analogues for permeability alteration. An expanded π-conjugation-based molecular precursor with an extremely high rigidity and a long hydrophobic length that is comparable to the hydrophobic width of plasma membrane was synthesized for membrane-inserted self-assembly. Guided by the cancer biomarker expression in vitro, the soluble precursors transform into hydrophobic monomers  forming assemblies inserted into the fluid phase of the membrane exclusively. Membrane insertion of rigid synthetic analogues destroys the selective permeability of the plasma membrane gradually. It eventually leads to cancer cell death, including drug resistant cancer cells.

[Deterioration of Healthcare Service Quality Provided by Occupational Physicians Due to Dumping by Health Checkup and Switching Health Checkup Organizations].

INTRODUCTION: From 2007, competitive bidding for procurement became widely employed by the Japanese Government, and health check providers for government workers are selected every year by this method. Deterioration of health check quality due to excessive price competition is a serious concern. The National Federation of Industrial Health Organization (Zeneiren) conducted an investigative research on the contracting of health check providers and occupational physicians in workplaces in 2015-2016 in an effort to prevent low-cost but low-quality health checks. The report of the research is available on the homepage of Zeneiren. In this paper, we provide a brief overview of the report, and deterioration of health check quality due to dumping by and switching of health check providers is discussed from economic and legal viewpoints. METHOD: Information was obtained from articles in print and on the Internet. RESULTS: A questionnaire survey of health check providers revealed that excessive discounts due to both competitive bidding and demand from companies commissioning health checks occurred on a routine basis, and some providers were concerned about worsening business conditions in the future. In a separate questionnaire survey of occupational physicians, it was discovered that they were able to evaluate the quality of health checks, whereas administrative officials responsible for selecting the providers were seldom able to adequately evaluate the health check quality, resulting in contracting providers of questionable quality, which in turn caused considerable dissatisfaction on the part of occupational physicians. Moreover, when health check providers were switched, the reporting format of health check results changed. The physicians did not favor such a change because of the considerably increased workload involved in coordinating past and current data and the risk of decreased occupational health service quality. DISCUSSION: Dumping makes the management of health check providers very difficult and is a cause of loss of social capital. If health check providers of good quality withdraw from the market, the supply of high-quality health checks decreases. This corresponds to external diseconomy caused by dumping on the part of the health check providers and loss of social surplus (economic surplus). CONCLUSIONS: To avoid deterioration of occupational health service due to low-quality health checks and changes in the reporting format, occupational physicians must actively engage in the selection of health check providers of good quality.

Genetic regulation of mitotic competence in G0 quiescent cells.

Quiescent (G0 phase) cells must maintain mitotic competence (MC) to restart the cell cycle. This is essential for reproduction in unicellular organisms and also for development and cell replacement in higher organisms. Recently, suppression of MC has gained attention as a possible therapeutic strategy for cancer. Using a Schizosaccharomyces pombe deletion-mutant library, we identified 85 genes required to maintain MC during the G0 phase induced by nitrogen deprivation. G0 cells must recycle proteins and RNA, governed by anabolism, catabolism, transport, and availability of small molecules such as antioxidants. Protein phosphatases are also essential to maintain MC. In particular, Nem1-Spo7 protects the nucleus from autophagy by regulating Ned1, a lipin. These genes, designated GZE (G-Zero Essential) genes, reveal the landscape of genetic regulation of MC.

Spin-Coated Crystalline Molecular Monolayers for Performance Enhancement in Organic Field-Effect Transistors.

In organic field-effect transistors, the first few molecular layers at the semiconductor/dielectric interface are regarded as the active channel for charge transport; thus, great efforts have been devoted to the modification and optimization of molecular packing at such interfaces. Here, we report organic monolayers with large-area uniformity and high crystallinity deposited by an antisolvent-assisted spin-coating method acting as the templating layers between the dielectric and thermally evaporated semiconducting layers. The predeposited crystalline monolayers significantly enhance the film crystallinity of upper layers and the overall performance of transistors using these hybrid-deposited semiconducting films, showing a high carrier mobility up to 11.3 cm2 V-1 s-1. Additionally, patterned transistor arrays composed of the templating monolayers are fabricated, yielding an average mobility of 7.7 cm2 V-1 s-1. This work demonstrates a promising method for fabricating low-cost, high-performance, and large-area organic electronics.

Regulating Higher-Order Organization through the Synergy of Two Self-Sorted Assemblies.

The extracellular matrix (ECM) is the natural fibrous scaffold that regulates cell behavior in a hierarchical manner. By mimicking the dynamic and reciprocal interactions between ECM and cells, higher-order molecular self-assembly (SA), mediated through the dynamic growth of scaffold-like nanostructures assembled by different molecular components, was developed. Designed and synthesized were two self-sorted coumarin-based gelators, a peptide molecule and a benzoate molecule, which self-assemble into nanofibers and nanobelts, respectively, with different dynamic profiles. Upon the dynamic growth of the fibrous scaffold assembled from peptide gelators, nanobelts assembled from benzoate gelators transform into a layer-by-layer nanosheet, reaching ninefold increase in height. By using light and an enzyme, the spatial-temporal growth of the scaffold can be modified, leading to in situ height regulation of the higher-order architecture.

Effects of goal-directed fluid therapy on enhanced postoperative recovery: An interventional comparative observational study with a historical control group on oesophagectomy combined with ERAS program.

BACKGROUND AND AIMS: The Enhanced Recovery after Surgery (ERAS) program has been proposed as a postoperative recovery-enhancing strategy. We frequently apply the Modified-ERAS program following oesophagectomy. This study aims to elucidate the impact of goal-directed fluid therapy (GDT) for the perioperative management of oesophageal cancer on the postoperative recovery of patients undergoing oesophagectomy. METHODS: This is an interventional before-after comparative observational study conducted at Kanagawa Cancer Centre, Japan. Patients who underwent elective oesophagectomy for oesophageal cancer were recruited. Group H (retrospectively collected) received intraoperative and postoperative management consisting of fluid administration without haemodynamic monitoring and the M-ERAS program, while Group S prospectively received management consisting of GDT and the M-ERAS program. The primary endpoint was the speed of gastrointestinal functional recovery, while secondary endpoints were the level of postoperative mobilisation, incidence of complications, postoperative length of hospital stay (LOS), and nutritional status after discharge. RESULTS: The proportion of patients who completely egested Gastrografin by postoperative day 4, the level of postoperative mobilisation, and achievement ratio for a 100-m walk on the first postoperative attempt were significantly higher in Group S than in Group H (P = 0.034, P = 0.0197, and P < 0.0001, respectively). No significant differences were observed in the postoperative LOS and incidence of complications within 30 days between the groups. The serum albumin levels at 6 months after discharge was higher in Group S than in Group H (P = 0.0002). CONCLUSIONS: The GDT-ERAS program enhanced postoperative gastrointestinal recovery and mobilisation, as well as postoperative nutritional status and protein synthesis. The program did not affect either postoperative LOS or the incidence of complications. TRIAL REGISTRATION: UMIN registration number: UMIN000013705, https://upload.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000015999.

Co-organizing synthesis of heterogeneous nanostructures through the photo-cleavage of pre-stabilized self-assemblies.

We devised a co-organizing synthesis by targeting metastable assemblies. Applying a photo-cleavage reaction to the pre-stabilized self-assembled nanostructures, we could successfully place physical constraints on the initiating stage of the molecular co-assembly (CA) process for exotic nanostructures that are away from the thermodynamic minima.

Usefulness of Surgical Apgar Score on Predicting Survival After Surgery for Gastric Cancer.

BACKGROUND: Complete surgical resection is essential for a cure for most gastric cancer. Recently it was reported that surgical Apgar score (SAS) can predict postoperative complication and that postoperative complication is associated with poor long-term survival. The aim of this study is to assess whether SAS can predict overall survival (OS) after surgery for gastric cancer. METHODS: We retrospectively compared clinicopathological characteristics and survival between high and low SAS score groups in patients who underwent gastrectomy for gastric cancer. RESULTS: Low-scored SAS group (group L) was significantly more common among ASA-PS 2, open approach, total gastrectomy, D2 lymph node dissection, postoperative complication grade 2-4, deep tumor invasion, lymph node metastases, and advanced pathological TNM stage than high-scored SAS group (group H). The 5-year OS of group H and group L were 81.6 and 55.9 %, respectively (p < .001); OS of group L tended to be poorer than that of group H in stage III patients (p = .060) and in stage IV patients (p < .001). In multivariate analysis, pathological stage and SAS were identified as independent predictors for OS. CONCLUSIONS: SAS is useful for predicting survival after surgery for gastric cancer.

Vectors for Genetically-Encoded Tags for Electron Microscopy Contrast in Drosophila.

BACKGROUND: One of the most notable recent advances in electron microscopy (EM) was the development of genetically-encoded EM tags, including the fluorescent flavoprotein Mini-SOG (Mini-Singlet Oxygen Generator). Mini-SOG generates good EM contrast, thus providing a viable alternative to technically-demanding methods such as immuno-electron microcopy (immuno-EM). Based on the Mini-SOG technology, in this paper, we describe the construction, validation and optimization of a series of vectors which allow expression of Mini-SOG in the Drosophila melanogaster genetic model system. FINDINGS: We constructed a Mini-SOG tag that has been codon-optimized for expression in Drosophila (DMS tag) using PCR-mediated gene assembly. The photo-oxidation reaction triggered by DMS was then tested using these vectors in Drosophila cell lines. DMS tag did not affect the subcellular localization of the proteins we tested. More importantly, we demonstrated the utility of the DMS tag for EM in Drosophila by showing that it can produce robust photo-oxidation reactions in the presence of blue light and the substrate DAB; the resultant electron micrographs contain electron-dense regions corresponding to the protein of interest. The vectors we generated allow protein tagging at both termini, for constitutive and inducible protein expression, as well as the generation of transgenic lines by P-element transformation. CONCLUSIONS: We demonstrated the feasibility of Mini-SOG tagging in Drosophila. The constructed vectors will no doubt be a useful molecular tool for genetic tagging to facilitate high-resolution localization of proteins in Drosophila by electron microscopy.

CNOT3 suppression promotes necroptosis by stabilizing mRNAs for cell death-inducing proteins.

The CCR4-NOT complex is conserved in eukaryotes and is involved in mRNA metabolism, though its molecular physiological roles remain to be established. We show here that CNOT3-depleted mouse embryonic fibroblasts (MEFs) undergo cell death. Levels of other complex subunits are decreased in CNOT3-depleted MEFs. The death phenotype is rescued by introduction of wild-type (WT), but not mutated CNOT3, and is not suppressed by the pan-caspase inhibitor, zVAD-fluoromethylketone. Gene expression profiling reveals that mRNAs encoding cell death-related proteins, including receptor-interacting protein kinase 1 (RIPK1) and RIPK3, are stabilized in CNOT3-depleted MEFs. Some of these mRNAs bind to CNOT3, and in the absence of CNOT3 their poly(A) tails are elongated. Inhibition of RIPK1-RIPK3 signaling by a short-hairpin RNA or a necroptosis inhibitor, necrostatin-1, confers viability upon CNOT3-depleted MEFs. Therefore, we conclude that CNOT3 targets specific mRNAs to prevent cells from being disposed to necroptotic death.

Randomly selected suppressor mutations in genes for NADH : quinone oxidoreductase-1, which rescue motility of a Salmonella ubiquinone-biosynthesis mutant strain.

The primary mobile electron-carrier in the aerobic respiratory chain of Salmonella is ubiquinone. Demethylmenaquinone and menaquinone are alternative electron-carriers involved in anaerobic respiration. Ubiquinone biosynthesis was disrupted in strains bearing deletions of the ubiA or ubiE genes. In soft tryptone agar both mutant strains swam poorly. However, the ubiA deletion mutant strain produced suppressor mutant strains with somewhat rescued motility and growth. Six independent suppressor mutants were purified and comparative genome sequence analysis revealed that they each bore a single new missense mutation, which localized to genes for subunits of NADH : quinone oxidoreductase-1. Four mutants bore an identical nuoG(Q297K) mutation, one mutant bore a nuoM(A254S) mutation and one mutant bore a nuoN(A444E) mutation. The NuoG subunit is part of the hydrophilic domain of NADH : quinone oxidoreductase-1 and the NuoM and NuoN subunits are part of the hydrophobic membrane-embedded domain. Respiration was rescued and the suppressed mutant strains grew better in Luria-Bertani broth medium and could use l-malate as a sole carbon source. The quinone pool of the cytoplasmic membrane was characterized by reversed-phase HPLC. Wild-type cells made ubiquinone and menaquinone. Strains with a ubiA deletion mutation made demethylmenaquinone and menaquinone and the ubiE deletion mutant strain made demethylmenaquinone and 2-octaprenyl-6-methoxy-1,4-benzoquinone; the total quinone pool was reduced. Immunoblotting found increased NADH : quinone oxidoreductase-1 levels for ubiquinone-biosynthesis mutant strains and enzyme assays measured electron transfer from NADH to demethylmenaquinone or menaquinone. Under certain growth conditions the suppressor mutations improved electron flow activity of NADH : quinone oxidoreductase-1 for cells bearing a ubiA deletion mutation.