Large-Area Flexible Carbon Nanofilms with Synergistically Enhanced Transmittance and Conductivity Prepared by Reorganizing Single-Walled Carbon Nanotube Networks.

Large-area flexible transparent conductive films (TCFs) are highly desired for future electronic devices. Nanocarbon TCFs are one of the most promising candidates, but some of their properties are mutually restricted. Here, a novel carbon nanotube network reorganization (CNNR) strategy, that is, the facet-driven CNNR (FD-CNNR) technique, is presented to overcome this intractable contradiction. The FD-CNNR technique introduces an interaction between single-walled carbon nanotube (SWNT) and Cu─-O. Based on the unique FD-CNNR mechanism, large-area flexible reorganized carbon nanofilms (RNC-TCFs) are designed and fabricated with A3-size and even meter-length, including reorganized SWNT (RSWNT) films and graphene and RSWNT (G-RSWNT) hybrid films. Synergistic improvement in strength, transmittance, and conductivity of flexible RNC-TCFs is achieved. The G-RSWNT TCF shows sheet resistance as low as 69 Ω sq-1 at 86% transmittance, FOM value of 35, and Young's modulus of ≈45 MPa. The high strength enables RNC-TCFs to be freestanding on water and easily transferred to any target substrate without contamination. A4-size flexible smart window is fabricated, which manifests controllable dimming and fog removal. The FD-CNNR technique can be extended to large-area or even large-scale fabrication of TCFs and can provide new insights into the design of TCFs and other functional films.

Gastric duplication cysts: literature review and a case report of rare multiple gastric duplication cysts treated by endoscopic submucosal dissection.

Gastric duplication cysts (GDCs) are rare structural abnormalities, especially in adults. We first report a rare case of small multiple GDCs in a woman, which presents as a submucosal tumor (SMT) at the gastric antrum. In consideration of the patient's request for surgical treatment and minimally invasive resection, endoscopic submucosal dissection (ESD) was performed to remove the cyst. The case provides a reference for ESD surgery to remove small GDCs. So far, there is no consensus or practice guidelines for the diagnosis and management of GDCs. Herein we perform a comprehensive literature review and discussion on GDCs. GDCs are 'repetitive' cystic or tubular structures of gastric mucosa and muscularis mucosae, and share the muscularis propria and serous layer with the normal gastric wall. GDCs protruding into the stomach cavity can be diagnosed by endoscopic ultrasound (EUS), which has higher specificity and accuracy than CT and MRI. Some GDCs may cause complications, even cancerization. Therefore, we suggest that once found, the GDCs could be completely resected. For GDCs protruding into the stomach cavity, endoscopic surgery such as ESD can be adopted to remove the lesion. Endoscopic full-thickness resection (EFTR) may become an option for larger GDCs in the future. For extraluminal GDC, laparoscopic surgery is currently preferred. In this review, we summarized the structural and histopathological characteristics of GDCs and various treatment therapies, in order to provide experience and reference for the diagnosis and treatment of GDCs in the future.

Appropriateness and Comprehensiveness of Using ChatGPT for Perioperative Patient Education in Thoracic Surgery in Different Language Contexts: Survey Study.

BACKGROUND: ChatGPT, a dialogue-based artificial intelligence language model, has shown promise in assisting clinical workflows and patient-clinician communication. However, there is a lack of feasibility assessments regarding its use for perioperative patient education in thoracic surgery. OBJECTIVE: This study aimed to assess the appropriateness and comprehensiveness of using ChatGPT for perioperative patient education in thoracic surgery in both English and Chinese contexts. METHODS: This pilot study was conducted in February 2023. A total of 37 questions focused on perioperative patient education in thoracic surgery were created based on guidelines and clinical experience. Two sets of inquiries were made to ChatGPT for each question, one in English and the other in Chinese. The responses generated by ChatGPT were evaluated separately by experienced thoracic surgical clinicians for appropriateness and comprehensiveness based on a hypothetical draft response to a patient's question on the electronic information platform. For a response to be qualified, it required at least 80% of reviewers to deem it appropriate and 50% to deem it comprehensive. Statistical analyses were performed using the unpaired chi-square test or Fisher exact test, with a significance level set at P<.05. RESULTS: The set of 37 commonly asked questions covered topics such as disease information, diagnostic procedures, perioperative complications, treatment measures, disease prevention, and perioperative care considerations. In both the English and Chinese contexts, 34 (92%) out of 37 responses were qualified in terms of both appropriateness and comprehensiveness. The remaining 3 (8%) responses were unqualified in these 2 contexts. The unqualified responses primarily involved the diagnosis of disease symptoms and surgical-related complications symptoms. The reasons for determining the responses as unqualified were similar in both contexts. There was no statistically significant difference (34/37, 92% vs 34/37, 92%; P=.99) in the qualification rate between the 2 language sets. CONCLUSIONS: This pilot study demonstrates the potential feasibility of using ChatGPT for perioperative patient education in thoracic surgery in both English and Chinese contexts. ChatGPT is expected to enhance patient satisfaction, reduce anxiety, and improve compliance during the perioperative period. In the future, there will be remarkable potential application for using artificial intelligence, in conjunction with human review, for patient education and health consultation after patients have provided their informed consent.

A self-reported inhibitor of metallo-carbapenemases for reversing carbapenem resistance.

Metallo-carbapenemases-mediated carbapenem-resistant Enterobacterales (CREs) has been acknowledged as "urgent threat" by the World Health Organization. The discovery of new strategies that block metallo-carbapenemases activity to reverse carbapenem resistance is an urgent need. In this study, a coumarin copper complex containing a PEG linker and glucose ligand, GluC-Cu, was used to reverse carbapenem resistance. Interestingly, it could effectively inhibit metallo-carbapenemases (NDM-1, IMP-1 and ImiS) with an IC50 value in the range of 0.23-1.21 µM, and simultaneously release the green fluorescence signal (GluC), therefore exhibiting self-reported inhibition performance. The inhibition mechanism of oxidizing Zn(II) thiolate site of NDM-1 from Cu2+ to Cu+ was verified by fluorescence assay, HR-MS, and XPS. Moreover, GluC-Cu in combination with meropenem showed excellent synergistic antibacterial effect to effectively combat E. coli expressing metallo-carbapenemases in vitro and in a mice infection model. This bifunctional metallo-carbapenemases inhibitor provides a novel chemical tool to overcome carbapenem resistance.

Preparing high-concentration individualized carbon nanotubes for industrial separation of multiple single-chirality species.

Industrial production of single-chirality carbon nanotubes is critical for their applications in high-speed and low-power nanoelectronic devices, but both their growth and separation have been major challenges. Here, we report a method for industrial separation of single-chirality carbon nanotubes from a variety of raw materials with gel chromatography by increasing the concentration of carbon nanotube solution. The high-concentration individualized carbon nanotube solution is prepared by ultrasonic dispersion followed by centrifugation and ultrasonic redispersion. With this technique, the concentration of the as-prepared individualized carbon nanotubes is increased from about 0.19 mg/mL to approximately 1 mg/mL, and the separation yield of multiple single-chirality species is increased by approximately six times to the milligram scale in one separation run with gel chromatography. When the dispersion technique is applied to an inexpensive hybrid of graphene and carbon nanotubes with a wide diameter range of 0.8-2.0 nm, and the separation yield of single-chirality species is increased by more than an order of magnitude to the sub-milligram scale. Moreover, with present separation technique, the environmental impact and cost of producing single-chirality species are greatly reduced. We anticipate that this method promotes industrial production and practical applications of single-chirality carbon nanotubes in carbon-based integration circuits.

Comparative efficacy of 5 non-pharmacological therapies for adults with post-stroke cognitive impairment: A Bayesian network analysis based on 55 randomized controlled trials.

Background: As a common sequela after stroke, cognitive impairment negatively impacts patients' activities of daily living and overall rehabilitation. Non-pharmacological therapies have recently drawn widespread attention for their potential in improving cognitive function. However, the optimal choice of non-pharmacological therapies for post-stroke cognitive impairment (PSCI) is still unclear. Hence, in this study, we compared and ranked 5 non-pharmacological therapies for PSCI with a Bayesian Network Meta-analysis (NMA), to offer a foundation for clinical treatment decision-making. Methods: PubMed, EMBASE, Web of Science, Cochrane Central Register of Controlled Trials, Chinese Biomedical Medicine, China National Knowledge Infrastructure, Wangfang Database, and China Science and Technology Journal Database were searched from database inception to December 31, 2021, to collect Randomized Controlled Trials for PSCI. All of the studies were assessed (according to Cochrane Handbook for Systematic Reviews) and then data were extracted by two researchers separately. Pairwise meta-analysis for direct comparisons was performed using Revman. NMA of Bayesian hierarchical model was performed by WinBUGS and ADDIS. STATA was used to construct network evidence plots and funnel plots. Results: A total of 55 trials (53 Two-arm trials and 2 Three-arm trials) with 3,092 individuals were included in this study. In the pair-wise meta-analysis, Transcranial Magnetic Stimulation (TMS), Virtual Reality Exposure Therapy (VR), Computer-assisted cognitive rehabilitation (CA), Transcranial Direct Current Stimulation (tDCS), and Acupuncture were superior to normal cognition training in terms of MoCA, MMSE, and BI outcomes. Bayesian NMA showed that the MoCA outcome ranked Acupuncture (84.7%) as the best therapy and TMS (79.7%) as the second. The MMSE outcome ranked TMS (76.1%) as the best therapy and Acupuncture as the second (72.1%). For BI outcome, TMS (89.1%) ranked the best. Conclusions: TMS and Acupuncture had a better effect on improving cognitive function in post-stroke patients according to our Bayesian NMA. However, this conclusion still needs to be confirmed with large sample size and high-quality randomized controlled trials. Registration: https://inplasy.com (No. INPLASY202260036).

Transcutaneous auricular vagus nerve stimulation in poststroke cognitive impairment: protocol for a randomised controlled trial.

BACKGROUND: As one of the most common stroke sequelae, poststroke cognitive impairment significantly impacts 17.6%-83% of survivors, affecting their rehabilitation, daily living and quality of life. Improving cognitive abilities among patients in stroke recovery is therefore critical and urgent. Transcutaneous auricular vagus nerve stimulation (TAVNS) is a non-invasive, safe, cost-effective treatment with great potential for improving the cognitive function of poststroke patients. This clinical research will evaluate the effectiveness, and help elucidate the possible underlying mechanisms, of TAVNS for improving poststroke cognitive function. METHODS AND ANALYSIS: A single-centre, parallel-group, allocation concealment, assessor-blinded randomised controlled clinical trial. We will allocate 88 recruited participants to the TAVNS or sham group for an intervention that will run for 8 weeks, 5 days per week with twice daily sessions lasting 30 min each. Blood tests will be performed and questionnaires issued at baseline and 8-week and 12 week follow-ups. Primary outcomes will be changes in cognitive function scores. Secondary outcomes will be changes in activities of daily living, quality of life and serum oxidative stress indicators. ETHICS AND DISSEMINATION: The Ethics Committee of the First Affiliated Hospital of Hunan University of Chinese Medicine has approved the protocol (No. HN-LL-YJSLW-2022200). Findings will be published in peer-reviewed academic journals and presented at scientific conferences. TRIAL REGISTRATION NUMBER: ChiCTR2200057808.

Clinical Effect of Hufu Copper Scraping on Shoulder-Hand Syndrome after Stroke.

Objective: To explore the clinical effect of Hufu copper scraping on shoulder-hand syndrome after stroke. Methods: A total of 60 patients with shoulder-hand syndrome after stroke admitted to the First Affiliated Hospital of Hunan University of Traditional Chinese Medicine were enrolled between January 2020 and June 2021. According to the random number table method, they were divided into the control group (n = 30) and the intervention group (n = 30). The control group was given routine rehabilitation intervention, while the intervention group was additionally given Hufu copper scraping. The intervention effect, occurrence of adverse reactions during the intervention, pain, swelling degree of affected hands, serum calcitonin gene-related peptide (CGRP), substance P (SP), and Barthel index before and after 2 months of intervention were compared between the two groups. Results: After intervention, the total response rate of the intervention group was significantly higher than that of the control group (96.67% vs 80.00%) (P < 0.05). After intervention, the score of the visual analogue scale (VAS) and water displacement in the intervention group were lower than those in the control group (P < 0.05). After intervention, levels of CGRP and SP in the intervention group were lower than those in the control group, while the Barthel index was higher compared to the control group (P < 0.05). Conclusion: The intervention effect of Hufu copper scraping is good, which can promote the recovery of related symptoms, relieve pain and swelling of affected hands, downregulate levels of disease-promoting markers, and accelerate recovery of related function in patients with shoulder-hand syndrome after stroke.

Discovery of Quercetin and Its Analogs as Potent OXA-48 Beta-Lactamase Inhibitors.

Carbapenem resistance in Enterobacteriaceae caused by OXA-48 ß-lactamase is a growing global health threat and has rapidly spread in many regions of the world. Developing inhibitors is a promising way to overcome antibiotic resistance. However, there are few options for problematic OXA-48. Here we identified quercetin, fisetin, luteolin, 3',4',7-trihydroxyflavone, apigenin, kaempferol, and taxifolin as potent inhibitors of OXA-48 with IC50 values ranging from 0.47 to 4.54 µM. Notably, the structure-activity relationship revealed that the substitute hydroxyl groups in the A and B rings of quercetin and its structural analogs improved the inhibitory effect against OXA-48. Mechanism studies including enzymatic kinetic assay, isothermal titration calorimetry (ITC), and surface plasmon resonance (SPR) analysis demonstrated that quercetin reversibly inhibited OXA-48 through a noncompetitive mode. Molecular docking suggested that hydroxyl groups at the 3', 4' and 7 positions in flavonoids formed hydrogen-bonding interactions with the side chains of Thr209, Ala194, and Gln193 in OXA-48. Quercetin, fisetin, luteolin, and 3',4',7-trihydroxyflavone effectively restored the antibacterial efficacy of piperacillin or imipenem against E. coli producing OXA-48, resulting in 2-8-fold reduction in MIC. Moreover, quercetin combined with piperacillin showed antimicrobial efficacy in mice infection model. These studies provide potential lead compounds for the development of ß-lactamase inhibitors and in combination with ß-lactams to combat OXA-48 producing pathogen.

Effectiveness of non-pharmacological interventions for treating post-stroke depression: Study protocol of an umbrella review of systematic reviews and meta-analyses.

BACKGROUND: Many systematic reviews and meta-analyses have evaluated the effectiveness of non-pharmacological therapies to improve symptoms of post-stroke depression (PSD) and reduce disability and mortality in patients with PSD. However, no research has appraised the credibility of the evidence. This study aims to summarize and evaluate the current evidence for non-pharmacological treatment of PSD and to seek effective treatment with reference to reliable evidence. METHODS: We searched the electronic databases EMBASE, MEDLINE, Cochrane Central, PubMed, PROSPERO, Web of Science, and CINAHL. We will search articles from the above database for all published meta-analyses to December 2021 to evaluate the effect of non-pharmacological treatment of PSD. Two reviewers will extract the general characteristics of the included articles, as well as participants, interventions, outcome measures, and conclusions. The quality evaluation of each systematic review will be conducted with reference to the AMSTAR 2 tool. The effect size of each review will be recalculated using either a fixed-effects or a random-effects model. Cochrane's Q test and I2 statistics will be used to evaluate the heterogeneity between studies. To determine whether a systematic review had small study effects, we will use the Egger test. We expect to extract valid evidence and classify it from strong to weak. RESULTS: The findings of this umbrella review will provide effective evidence for the non-pharmacological treatment of PSD. CONCLUSION: Our research conclusion will provide clinical staff and PSD patients with appropriate treatment recommendations. ETHICS AND DISSEMINATION: As the data were obtained from published materials, there is no need for ethical approval for this umbrella review. The findings of this umbrella review will be published in a peer-reviewed journal. INPLASY REGISTRATION NUMBER: INPLASY2021100083.

Auricular Point Acupressure Combined with Compound Lidocaine Cream to Manage Arteriovenous Fistula Puncture Pain: A Multicenter Randomized Controlled Trial.

BACKGROUND: Arteriovenous fistula (AVF) puncture pain is an inevitable problem for maintenance hemodialysis (MHD) patients and may seriously endanger the physical and mental health of patients with MHD. Studies have shown that drug or nondrug measures can reduce AVF puncture pain, but much improvement is needed. When combined with compound lidocaine cream (CLC) in the treatment of AVF puncture pain, auricular point acupressure (APA)-a therapeutic method in which specific points on the auricle of the outer ear are stimulated to treat various disorders of the body-and the therapeutic value and synergistic effects of auriculotherapy merit further investigation. METHODS: 120 MHD patients were recruited at blood purification centers in three hospitals between January 2016 and April 2019. After completion of the baseline survey, all patients were randomly divided by the envelope method into a control group, APA group, CLC group, and APA combined with CLC, with 30 subjects per group. The numerical rating scale (NRS) of pain was used to measure the pain before intervention and 1, 4, and 8 weeks after intervention. The State-Trait Anxiety Inventory (STAI), General Comfort Questionnaire (GCQ), blood pressure, and heart rates were obtained before and after the intervention. RESULTS: Pain, anxiety, comfort, blood pressure (BP), and heart rates (HR) of the three groups were better than those of the control group; the difference was statistically significant (P < 0.05). In addition, the APA combined with CLC group was better than the APA group and CLC group, respectively, in those outcomes (P < 0.05). CONCLUSION: Both APA and CLC can effectively relieve AVF puncture pain, and the combined application has more outstanding effects.

Establishment and Clinical Application of an Artificial Intelligence Diagnostic Platform for Identifying Rectal Cancer Tumor Budding.

Tumor budding is considered a sign of cancer cell activity and the first step of tumor metastasis. This study aimed to establish an automatic diagnostic platform for rectal cancer budding pathology by training a Faster region-based convolutional neural network (F-R-CNN) on the pathological images of rectal cancer budding. Postoperative pathological section images of 236 patients with rectal cancer from the Affiliated Hospital of Qingdao University, China, taken from January 2015 to January 2017 were used in the analysis. The tumor site was labeled in Label image software. The images of the learning set were trained using Faster R-CNN to establish an automatic diagnostic platform for tumor budding pathology analysis. The images of the test set were used to verify the learning outcome. The diagnostic platform was evaluated through the receiver operating characteristic (ROC) curve. Through training on pathological images of tumor budding, an automatic diagnostic platform for rectal cancer budding pathology was preliminarily established. The precision-recall curves were generated for the precision and recall of the nodule category in the training set. The area under the curve = 0.7414, which indicated that the training of Faster R-CNN was effective. The validation in the validation set yielded an area under the ROC curve of 0.88, indicating that the established artificial intelligence platform performed well at the pathological diagnosis of tumor budding. The established Faster R-CNN deep neural network platform for the pathological diagnosis of rectal cancer tumor budding can help pathologists make more efficient and accurate pathological diagnoses.

Negative feedback loop of ERK/CREB/miR-212-3p inhibits HBeAg-induced macrophage activation.

The activation of liver macrophages is closely related to liver injury after HBV infection. Our previous results demonstrated that HBeAg played a key role in inducing macrophage activation. As we all know, miRNAs are involved in the regulation of multiple immune cell functions. Meanwhile, we have shown that miR-155 positively regulates HBeAg-induced macrophage activation and accelerates liver injury. Subsequently, based on our previous miRNA sequencing results, we further evaluated the role of miR-212-3p called 'neurimmiR' in HBeAg-induced macrophages in this study. First, miR-212-3p expression was significantly elevated in HBeAg-treated macrophages. Meanwhile, we found up-regulation of miR-212-3p significantly decreased the production of cytokines, whereas knockdown of miR-212-3p held the opposite effect by gains and losses of function. Mechanically, although MAPK signal pathway, including ERK, JNK and p38, was activated in HBeAg-induced macrophages, only ERK promoted the expression of miR-212-3p via transcription factor CREB, which was able to bind to the promoter of miR-212-3p verified by ChIP assay. Moreover, we further indicated that up-regulated miR-212-3p inhibited HBeAg-induced inflammatory cytokine production through targeting MAPK1. In conclusion, miR-212-3p was augmented in HBeAg-stimulated macrophages via ERK/CREB signal pathway and the elevated miR-212-3p suppressed inflammatory cytokine production induced by HBeAg through targeting MAPK1.

Kinetic, Thermodynamic, and Crystallographic Studies of 2-Triazolylthioacetamides as Verona Integron-Encoded Metallo-ß-Lactamase 2 (VIM-2) Inhibitor.

Inhibition of ß-lactamases presents a promising strategy to restore the ß-lactams antibacterial activity to resistant bacteria. In this work, we found that aromatic carboxyl substituted 2-triazolylthioacetamides 1a-j inhibited VIM-2, exhibiting an IC50 value in the range of 20.6-58.6 µM. The structure-activity relationship study revealed that replacing the aliphatic carboxylic acid with aromatic carboxyl improved the inhibitory activity of 2-triazolylthioacetamides against VIM-2. 1a-j (16 mg/mL) restored the antibacterial activity of cefazolin against E. coli cell expressing VIM-2, resulting in a 4-8-fold reduction in MICs. The isothermal titration calorimetry (ITC) characterization suggested that the primary binding 2-triazolylthioacetamide (1b, 1c, or 1h) to VIM-2 was a combination of entropy and enthalpy contributions. Further, the crystal structure of VIM-2 in complex with 1b was obtained by co-crystallization with a hanging-drop vapour-diffusion method. The crystal structure analysis revealed that 1b bound to two Zn(II) ions of the enzyme active sites, formed H-bound with Asn233 and structure water molecule, and interacted with the hydrophobic pocket of enzyme activity center utilizing hydrophobic moieties; especially for the phenyl of aromatic carboxyl which formed π-π stacking with active residue His263. These studies confirmed that aromatic carboxyl substituted 2-triazolylthioacetamides are the potent VIM-2 inhibitors scaffold and provided help to further optimize 2-triazolylthioacetamides as VIM-2 even or broad-spectrum MßLs inhibitors.

The assemblage of covalent and metal binding dual functional scaffold for cross-class metallo-ß-lactamases inhibition.

Aim: The discovery and development of novel broad-spectrum MßLs inhibitors are urgent to overcome antibiotic resistance mediated by MßLs. Methods & results: Herein, the synthesized 21 compounds exhibited potent inhibition to the clinically important MßLs (NDM-1, IMP-1 and ImiS) and effectively restored the antibacterial efficacy of cefazolin and imipenem against Escherichia coli harboring MßLs. 5b was first identified to be dual functional broad-spectrum MßLs inhibitor through assemblage of covalent and metal binding scaffold, which irreversibly inhibited B1, B2 MßLs via forming a Se-S covalent bond, and competitively inhibited B3 MßLs by coordinating the metals at active site. Conclusion: The designed compounds can serve as potent broad-spectrum MßLs inhibitors and combat MßLs-producing 'superbug' in combination with ß-lactams.

Comparison of the results of two chest tube managements during an enhanced recovery program after video-assisted thoracoscopic lobectomy: A randomized trial.

BACKGROUND: This study compared the results of the application of two different chest tube management systems; a drainage ball with low negative pressure and the more commonly used chest tube with water-sealed bottle, after video-assisted thoracoscopic (VATS) lobectomy. METHODS: A total of 60 patients undergoing lobectomy were enrolled into this prospective open label randomized clinical trial and equally divided into two groups. The data collected in the trial included age, gender, forced expiratory volume in 1 second (FEV1), blood loss, operation time, drainage volume, drainage time, length of stay, postoperative pain score according to the Visual Analogue Scale (VAS) within 24 hours after surgery and chest tube removal. This study was registered at ClinicalTrials.gov (NCT03598296). RESULTS: The characteristics of the patients were similar in both groups. Group ball patients had a lower pain score (after operation: 3.47 ± 1.80 vs. 6.20 ± 1.56, P < 0.001; after removal of chest tube: 1.47 ± 1.28 vs. 3.00 ± 1.29, P < 0.001); less analgesic used (2.83 ± 2.09 times vs. 5.00 ± 3.24 times, P = 0.003); less drainage time (upper tube: 3.89 ± 1.63 days vs. 5.10 ± 2.02 days, P = 0.048; lower tube: upper lobe 4.84 ± 1.61 days vs. 5.90 ± 1.52 days, P = 0.041; lower lobe: 3.82 ± 1.08 days vs. 5.70 ± 2.63 days, P = 0.042) and shorter length of stay (5.40 ± 1.65 days vs. 6.37 ± 1.99 days, P = 0.045). All other related parameters were similar in both groups. CONCLUSIONS: For patients undergoing lobectomy, using a drainage ball with negative pressure could reduce hospitalization days and postoperative pain compared with the more commonly used chest tube with water-sealed bottle when a strict postoperative curative procedure was performed.

Real-time monitoring of D-Ala-D-Ala dipeptidase activity of VanX in living bacteria by isothermal titration calorimetry.

The d,d-dipeptidase enzyme VanX is the main cause of vancomycin resistance in gram-positive bacteria because of hydrolysis of the D-Ala-D-Ala dipeptide used in cell-wall biosynthesis. Continuous assay of VanX has proven challenging due to lack of a chromophoric substrate. Here, we report a direct approach for continuous assay of VanX in vitro and in vivo from hydrolysis of D-Ala-D-Ala, based on the heat-rate changes measured with isothermal titration calorimetry (ITC). With the ITC approach, determination of kinetic parameters of VanX hydrolyzing D-Ala-D-Ala and the inhibition constant of d-cysteine inhibitor yielded KM of 0.10 mM, kcat of 11.5 s-1, and Ki of 18.8 µM, which are consistent with the data from ninhydrin/Cd(II) assays. Cell-based ITC studies demonstrated that the VanX expressed in E. coli and in clinical strain VRE was inhibited by d-cysteine with IC50 values of 29.8 and 28.6 µM, respectively. Also, the total heat from D-Ala-D-Ala (4 mM) hydrolysis decreases strongly (in absolute value) from 1.26 mJ for VRE to 0.031 mJ for E. faecalis, which is consistent with the large MIC value of vancomycin of 512 µg/mL for VRE and the much smaller value of 4 µg/mL for E. faecalis. The ITC approach proposed here could be applied to screen and evaluate small molecule inhibitors of VanX or to identify drug resistant bacteria.

Amino Acid Thioesters Exhibit Inhibitory Activity against B1-B3 Subclasses of Metallo-ß-lactamases.

Superbug infection caused by metallo-ß-lactamases (MßLs) is a global public health threat. Previous studies reported that the thioesters specifically inhibited the B3 subclass MßL L1. In this work, nine amino acid thioesters 1-9 were synthesized, the activity evaluation revealed that all of these molecules exhibited broad-spectrum inhibitory efficacy against ImiS, IMP-1, NDM-1, and L1, with IC50 values range of 0.02-54.9 µM (except 5 and 7 on NDM-1), and 1 was found to be the best inhibitor with IC50 range of 0.02-16.63 µM. Minimal inhibitory concentration (MIC) assays showed that thioesters 1, 5 and 9 restored 2-32-fold antibacterial activity of cefazolin and/or imipenem against both Escherichia coli BL21 and DH10B strain expressing ImiS, L1, IMP-1 and NDM-1 (except 5 on NDM-1), and also, thioester 1 increased 2-4-fold antimicrobial activity of cefazolin on two clinical strains Pseudomonas aeruginosa and Klebsiella pneumoniae producing NDM-1. Stability evaluation indicated that thioester 1 was partially hydrolysed by MßLs to be converted into the mercaptoacetic acid, revealing that the thioester and its hydrolysate mercaptoacetic acid jointly inhibit MßLs. Isothermal titration calorimetry (ITC) monitoring showed that thioester 1 exhibited dose-dependent inhibition on four MßLs tested, and the binding of 1/L1 showed mainly enthalpy driven, while 1/NDM-1 was found to be more entropy driven. Docking studies suggested that 1 bound to Zn(II) ion(s) preferentially via its carboxylate group, while other moieties interacted mostly with the conserved active site residues.

Influence of Alloy Substrate Treatment on Microstructure and Surface Performances of Arc-Ion Plated Gold-Like Film.

Three typical surface pretreatment strategies (grind, drawing, polishing) are employed to explore the influence of alloy substrate treatment on microstructure and surface performances of arc-ion plated TiN and ZrN films. The luminance and color of the films are measured by the color coordinate value of CIELab system (a color system which is defined by the International Commission on illumination). The crystal phases, morphology and microstructure are characterized and analyzed by scanning electron microscope (SEM) and X-ray powder diffraction (XRD). In addition, the anti-alkali, salt corrosion and anti-oxidation performances of films are systematically researched. The results show that the films with grinding pretreatment are more like gold color, "L" values are 77.27 cd/m² and 80.30 cd/m². The "b" value of TiN film is 29.96, which is the same as that of pure gold. The "a" value of ZrN film is 0.31, which is the same as pure silver. The density of TiN and ZrN films is the best, and both TiN and ZrN films were crystalline. They have the best anti-alkali and anti-oxidation performance. The films with drawing pretreatment show slant red color and have medium brightness values (74.07⁻76.37 cd/m²), worse compactness, obvious furrows and holes in their microstructures and worse salt corrosion and anti-oxidation performances. However, the TiN films are in amorphous states. The films with polishing pretreatment have the lowest brightness (72.66 cd/m²), gold-like color, superior compactness and best salt corrosion performance, which have a small number of holes. The TiN films with polishing pretreatment are also in amorphous state. Above all, alloy substrate pretreatment by grinding has the best gold-like color, brightness, compactness and corrosion resistance performance. This work exclusively sheds new light on surface pretreatment of alloy substrate by arc-ion plated films and also provides a reference for corrosion resistance performance of gold-like films.