Gut microbial profile of treatment-naive patients with primary biliary cholangitis.

Background and aims: The pathogenesis of primary biliary cholangitis (PBC) is associated with alterations of gut microbiota. We compared the gut microbiota of PBC patients and healthy controls from Zhejiang Province and assessed the use of these data for the diagnosis of PBC. Methods: First, 16S rRNA gene sequencing was used to characterize the gut microbiota of treatment-naive PBC patients (n=25) and matched healthy controls (n=25). Then, the value of gut microbiota composition for the diagnosis of PBC and assessment of PBC severity was determined. Results: The gut microbiota of PBC patients had lower diversity based on three different metrics of alpha-diversity (ace, Chao1, and observed features) and fewer overall genera (all p<0.01). PBC patients had significant enrichment of four genera and significant depletion of eight genera. We identified six amplicon sequence variants (Serratia, Oscillospirales, Ruminococcaceae, Faecalibacterium, Sutterellaceae, and Coprococcus) as optimal biomarkers to distinguish PBC patients from controls based on receiver operating characteristic analysis (area under the curve [AUC] = 0.824). PBC patients who were anti-gp210-positive had lower levels of Oscillospiraceae than those who were anti-gp210-negative. KEGG functional annotation suggested the major changes in the gut microbiota of PBC patients were related to lipid metabolism and biosynthesis of secondary metabolites. Conclusion: We characterized the gut microbiota of treatment-naive PBC patients and healthy controls from Zhejiang Province. The PBC patients had significant alterations in their gut microbiota, suggesting that gut microbiota composition could be useful as a non-invasive tool for the diagnosis of PBC.

[The Auxiliary Diagnostic Value of Serum Adenosine Deaminase in Acute Leukemia at Clinical Test].

OBJECTIVE: To investigate the auxiliary diagnostic value of serum adenosine deaminase (ADA) in acute leukemia (AL) at clinical test. METHODS: 123 AL patients hospitalized in Zhejiang hospital from November 2018 to March 2020 were enrolled as the observation group, and 98 healthy people in the same period were randomly enrolled as the control group. AL patients were divided into two groups: 77 acute myeloid leukemia (AML) patients for AML group and 46 acute lymphoblastic leukemia (ALL) patients for ALL group. The levels of adenosine deaminase (ADA), alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutamyl transpeptidase (GGT), lactate dehydrogenase (LDH) and homocysteine (Hcy) in serum of the patients were detected, and the correlation of ADA with these items was analyzed. Receiver operating characteristic curve (ROC) was used to analyze the clinical diagnostic value of ADA, Yoden index was used to confirm the best cut-off point. RESULTS: The serum ADA level in AL patients was significant higher than that in control group (P < 0.05). The results of Pearson correlation analysis showed that there was a significant positive correlation of ADA with Hcy, ALT, AST, GGT, LDH in AML group (r = 0.47, r = 0.28, r = 0.37, r = 0.22, r = 0.55); and also there was a significant positive correlation of ADA with GGT in ALL group (r = 0.54). In AML group, the maximum area under ROC curve was 0.761 (P = 0.00), 95% confidence interval was 0.682-0.841, sensitivity was 54.50%, specificity was 98.90%, and the best cut-off point was 17.1 U/L. In ALL group, the maximum area under ROC curve was 0.785, 95% confidence interval was 0.694-0.877, sensitivity was 65.90%, specificity was 84.00%, and the best cut-off point was 13.45 U/L. CONCLUSION: The detection of ADA in serum can be used as an auxiliary examination in patients with AL, which can provide a certain value for the diagnosis of the disease.

Risk factors for tumor recurrence in patients with pT3N0M0 thoracic esophageal squamous cell carcinoma after esophagectomy.

OBJECTIVE: To analyze the factors contributing to recurrence in patients with pT3N0M0 thoracic esophageal squamous cell carcinoma (ESCC). METHODS: Patients with pT3N0M0 thoracic ESCC who underwent esophagectomy from January 2008 to December 2012 were included retrospectively. The last date of follow-up was 1 December 2016. Multivariate proportional hazard Cox models were used to identify factors associated with total (i.e., any) recurrence (TR), locoregional recurrence (LR), and distant metastasis (DM). RESULTS: A total of 692 patients were included. The median follow-up was 53 months (range: 3-107). The 3- and 5-year TR, LR, and DM rates were 35.8% and 41.0%, 28.7% and 32.1%, and 16.8% and 21.1%, respectively. The Cox analyses showed that the tumor location, number of dissected lymph nodes, and postoperative therapies were significantly associated with LR. The subgroup analysis showed that postoperative therapies could significantly decrease LR in the mediastinum but not in the neck and upper abdomen regions. CONCLUSIONS: The recurrence rate of pT3N0M0 thoracic ESCC patients was high, especially for LR in the mediastinum. Postoperative therapies can significantly reduce the incidence of mediastinal recurrence.

A leptospiral AAA+ chaperone-Ntn peptidase complex, HslUV, contributes to the intracellular survival of Leptospira interrogans in hosts and the transmission of leptospirosis.

Leptospirosis caused by Leptospira is a zoonotic disease of global importance but it is considered as an emerging or re-emerging infectious disease in many areas in the world. Until now, the mechanisms about pathogenesis and transmission of Leptospira remains poorly understood. As eukaryotic and prokaryotic proteins can be denatured in adverse environments and chaperone-protease/peptidase complexes degrade these harmful proteins, we speculate that infection may also cause leptospiral protein denaturation, and the HslU and HslV proteins of L. interrogans may compose a complex to degrade denatured proteins that enhances leptospiral survival in hosts. Here we show that leptospiral HslUV is an ATP-dependent chaperone-peptidase complex containing ATPase associated with various cellular activity (AAA+) and N-terminal nucleophile (Ntn) hydrolase superfamily domains, respectively, which hydrolyzed casein and chymotrypsin-like substrates, and this hydrolysis was blocked by threonine protease inhibitors. The infection of J774A.1 macrophages caused the increase of leptospiral denatured protein aggresomes, but more aggresomes accumulated in hslUV gene-deleted mutant. The abundant denatured leptospiral proteins are involved in ribosomal structure, flagellar assembly, two-component signaling systems and transmembrane transport. Compared to the wild-type strain, infection of cells in vitro with the mutant resulted in a higher number of dead leptospires, less leptospiral colony-forming units and lower growth ability, but also displayed a lower half lethal dose, attenuated histopathological injury and decreased leptospiral loading in lungs, liver, kidneys, peripheral blood and urine in hamsters. Therefore, our findings confirmed that HslUV AAA+ chaperone-Ntn peptidase complex of L. interrogans contributes to leptospiral survival in hosts and transmission of leptospirosis.

3D Printing of Aniline Tetramer-Grafted-Polyethylenimine and Pluronic F127 Composites for Electroactive Scaffolds.

Electroactive hydrogel scaffolds are fabricated by the 3D-printing technique using composites of 30% Pluronic F127 and aniline tetramer-grafted-polyethylenimine (AT-PEI) copolymers with various contents from 2.5% to 10%. The synthesized AT-PEI copolymers can self-assemble into nanoparticles with the diameter of ≈50 nm and display excellent electroactivity due to AT conjugation. The copolymers are then homogeneously distributed into 30% Pluronic F127 solution by virtue of the thermosensitivity of F127, denoted as F/AT-PEI composites. Macroscopic photographs of latticed scaffolds elucidate their excellent printability of F/AT-PEI hydrogels for the 3D-printing technique. The conductivities of the printed F/AT-PEI scaffolds are all higher than 2.0 × 10-3 S cm-1 , which are significantly improved compared with that of F127 scaffold with only 0.94 × 10-3 S cm-1 . Thus, the F/AT-PEI scaffolds can be considered as candidates for application in electrical stimulation of tissue regeneration such as repair of muscle and cardiac nerve tissue.

Enhanced conductivity of rGO/Ag NPs composites for electrochemical immunoassay of prostate-specific antigen.

Electrode materials play a vital role in the development of electrochemical immunosensors (EIs), particularly of label-free EIs. In this study, composites containing reduced graphene oxide with silver nanoparticles (rGO/Ag NPs) were synthesized using binary reductants, i.e. hydrazine hydrate and sodium citrate. Due to the fact that graphene oxide (GO) was fully restored to rGO, and rGO stacking was effectively inhibited by insertion of small Ag NPs between the graphene sheets, the electrical conductivity of rGO/Ag NPs composites was significantly improved compared to rGO alone, with an enhancement factor of 346% at 40wt% of rGO. Moreover, the conducting path between rGO and Ag NPs formed because the structural defects in rGO were effectively repaired by decoration with Ag NPs. Subsequently, based on a screen-printed three-electrode system, a label-free EI for detecting prostate-specific antigen (PSA) was constructed using rGO/Ag NPs composites as a support material. The fabricated EIs demonstrated a wide linear response range (1.0-1000ng/ml), low detection limit (0.01ng/ml) and excellent specificity, reproducibility and stability. Thus, the proposed EIs based on rGO/Ag NPs composites can be easily extended for the ultrasensitive detection of different protein biomarkers.

Effect of intracellular expression of antimicrobial peptide LL-37 on growth of escherichia coli strain TOP10 under aerobic and anaerobic conditions.

Antimicrobial peptides (AMPs) can cause lysis of target bacteria by directly inserting themselves into the lipid bilayer. This killing mechanism confounds the identification of the intracellular targets of AMPs. To circumvent this, we used a shuttle vector containing the inducible expression of a human cathelicidin-related AMP, LL-37, to examine its effect on Escherichia coli TOP10 under aerobic and anaerobic growth conditions. Induction of LL-37 caused growth inhibition and alteration in cell morphology to a filamentous phenotype. Further examination of the E. coli cell division protein FtsZ revealed that LL-37 did not interact with FtsZ. Moreover, intracellular expression of LL-37 results in the enhanced production of reactive oxygen species (ROS), causing lethal membrane depolarization under aerobic conditions. Additionally, the membrane permeability was increased after intracellular expression of LL37 under both aerobic and anaerobic conditions. Transcriptomic analysis revealed that intracellular LL-37 mainly affected the expression of genes related to energy production and carbohydrate metabolism. More specifically, genes related to oxidative phosphorylation under both aerobic and anaerobic growth conditions were affected. Collectively, our current study demonstrates that intracellular expression of LL-37 in E. coli can inhibit growth under aerobic and anaerobic conditions. While we confirmed that the generation of ROS is a bactericidal mechanism for LL-37 under aerobic growth conditions, we also found that the intracellular accumulation of cationic LL-37 influences the redox and ion status of the cells under both growth conditions. These data suggest that there is a new AMP-mediated bacterial killing mechanism that targets energy metabolism.