MYC-regulated lncRNA NEAT1 promotes B cell proliferation and lymphomagenesis via the miR-34b-5p-GLI1 pathway in diffuse large B-cell lymphoma.

BACKGROUND: LncRNA NEAT1 has been identified as a tumour driver in many human cancers. However, the underlying mechanism of lncRNA NEAT1 in diffuse large B-cell lymphoma (DLBCL) progression is unclear. METHODS: The expression levels of NEAT1, GLI1 and miR-34b-5p were detected by RT-qPCR and Western blotting in DLBCL tissues and cell lines. MTT and colony formation assays were performed to examine cell proliferation, while annexin-V staining and TUNEL assays were performed to measure cell apoptosis. The effect of NEAT1, GLI1 and miR-34b-5p on cell cycle-associated proteins was evaluated by Western blotting. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were employed to investigate the interaction between NEAT1 and miR-34b-5p or GLI1 and miR-34b-5p. Moreover, chromatin immunoprecipitation (ChIP) was performed to demonstrate the interaction between MYC and NEAT1. RESULTS: NEAT1 and GLI1 were upregulated while miR-34b-5p was downregulated in DLBCL tissues and cell lines compared to normal controls. Knockdown of NEAT1 or overexpression of miR-34b-5p inhibited cell proliferation but promoted cell apoptosis. Overexpression of NEAT1 reversed GLI1-knockdown induced attenuation of cell proliferation. In other words, NEAT1 acted as a competing endogenous RNA (ceRNA), regulating the miR-34b-5p-GLI1 axis, further affecting the proliferation of DLBCL. Moreover, MYC modulated NEAT1 transcription by directly binding to the NEAT1 promoter. CONCLUSION: We revealed that MYC-regulated NEAT1 promoted DLBCL proliferation via the miR-34b-5p-GLI1 pathway, which could provide a novel therapeutic target for DLBCL.

Stool vs. Serum Hepatitis B Virus DNA in Patients with Chronic Hepatitis B.

BACKGROUND Serum hepatitis B virus (HBV) DNA and hepatitis B e antigen (HBeAg) liver function in patients with chronic hepatitis B (CHB) are significantly associated. A comparison of clinical significance of fecal HBV DNA and serum HBV DNA has not yet been reported. MATERIAL AND METHODS Stool and serum samples were collected from 66 patients with CHB. Fecal HBV DNA, serum HBV DNA, and intestinal microbiota DNA were detected by real-time quantitative fluorescence polymerase chain reaction (PCR). Liver function and HBeAg were analyzed. RESULTS The stool and serum HBV DNA were positively correlated (r=0.57, P=0.001). Fecal HBV DNA was higher in the HBeAg-positive group than in the HBeAg-negative group (P=0.02). Fecal HBV DNA was negatively correlated with alkaline phosphatase (ALP) (r=-0.41, P=0.001) and TBIL (r=-0.29, P=0.02), and was positively correlated with Enterococcus (r=0.38, P=0.002). Serum HBV DNA was negatively correlated with alanine aminotransferase (ALT) (r=-0.30,P=0.02), aminotransferase (AST) (r=-0.26, P=0.049), and Lactobacillus (r=-0.31, P=0.01). CONCLUSIONS These observations suggest that fecal HBV DNA and serum HBV DNA in patients with CHB have different effects. Fecal HBV DNA might be associated with changes in Enterococcus concentrations, but serum HBV DNA is not.

Antimicrobial resistance patterns and characterization of integrons in clinical isolates of Shigella from China.

One hundred fifty-three Shigella isolates were examined for multiple antibiotic resistance phenotypes and prevalence of class 1 and class 2 integron sequences. The gene cassettes dfrA17-aadA5, dfrA12-orfF-aadA2, and arr-3-aacA4 were found in typical class 1 integrons. The gene cassettes blaOXA-1-aadA1 and dfrA1-sat1-aadA1 were detected in atypical class 1 integrons and in class 2 integrons, respectively. This is the first report of arr-3-aacA4 cassette detected in typical class 1 integrons among Shigella isolates. Rates of antibiotic resistance were different between integron-positive and integron-negative strains (P < 0.05), and all integron-positive isolates were resistant to at least 3 different antimicrobial agents. Typical class 1 integron-positive isolates showed higher resistance rates to cefotaxime and ciprofloxacin than did integron-negative ones (P < 0.05). Typical class 1 integrons and ß-lactamase genes were found in conjugative plasmids, otherwise class 2 and atypical class 1 integrons were located on chromosome. This study demonstrated the wide distribution of class 1 integrons in Shigella spp., which may lead resistance to cefotaxime and ciprofloxacin in China.

Palladium-Catalyzed Regio- and Diastereoselective Hydro(hetero)arylation for Rapid Construction of Quaternary Center Containing Cyclobutanes.

Herein we report a Pd-catalyzed regio- and diastereoselective hydro(hetero)arylation of inactivated alkylidenecyclobutanes. This protocol provides a rapid and atom-economical route to access 3-cyclobutyl (hetero)arenes with good functionalities toleration. With the assistance of the directing group, nucleophilic attack happened on the bulkier γ-position to form the quaternary carbon center. Furthermore, the selected products exhibited antitumor bioactivities.

Crash injury severity prediction considering data imbalance: A Wasserstein generative adversarial network with gradient penalty approach.

For each road crash event, it is necessary to predict its injury severity. However, predicting crash injury severity with the imbalanced data frequently results in ineffective classifier. Due to the rarity of severe injuries in road traffic crashes, the crash data is extremely imbalanced among injury severity classes, making it challenging to the training of prediction models. To achieve interclass balance, it is possible to generate certain minority class samples using data augmentation techniques. Aiming to address the imbalance issue of crash injury severity data, this study applies a novel deep learning method, the Wasserstein generative adversarial network with gradient penalty (WGAN-GP), to investigate a massive amount of crash data, which can generate synthetic injury severity data linked to traffic crashes to rebalance the dataset. To evaluate the effectiveness of the WGAN-GP model, we systematically compare performances of various commonly-used sampling techniques (random under-sampling, random over-sampling, synthetic minority over-sampling technique and adaptive synthetic sampling) with respect to dataset balance and crash injury severity prediction. After rebalancing the dataset, this study categorizes the crash injury severity using logistic regression, multilayer perceptron, random forest, AdaBoost and XGBoost. The AUC, specificity and sensitivity are employed as evaluation indicators to compare the prediction performances. Results demonstrate that sampling techniques can considerably improve the prediction performance of minority classes in an imbalanced dataset, and the combination of XGBoost and WGAN-GP performs best with an AUC of 0.794 and a sensitivity of 0.698. Finally, the interpretability of the model is improved by the explainable machine learning technique SHAP (SHapley Additive exPlanation), allowing for a deeper understanding of the effects of each variable on crash injury severity. Findings of this study shed light on the prediction of crash injury severity with data imbalance using data-driven approaches.

The prognostic significance of circulating plasma cells in newly diagnosed multiple myeloma patients.

Objective: Multiple myeloma (MM) is a highly characteristic tumor that is influenced by numerous factors that determine its prognosis. Studies indicate that the presence of circulating plasma cells (cPCs) is a detrimental factor that significantly impacts the prognosis of patients with MM. Methods: This study retrospectively analyzed the prognostic value of cPCs quantified by 10-color flow cytometry in 145 newly diagnosed MM (NDMM) cases in the First Affiliated Hospital of Soochow University from November 2018 to February 2021. The study was approved by the Ethics Committee of the hospital (2021 No. 93). Results: Of the 145 patients, 99 (68.2%) were detected cPCs. Through receiver operating characteristics (ROC) analysis, an optimal threshold of 0.165% was identified as a predictor for overall survival (OS). The median progression-free survival (PFS) was 33 months in patients with cPCs ≥0.165%, whereas those with cPCs <0.165% had a PFS of <33 months (p=0.001). The median OS was not reached for two groups; the 3-year OS for patients with cPCs ≥0.165% was 71% compared with 87% for those with cPCs <0.165% (p=0.003). In transplant patients, cPCs ≥0.165% also predicted worse prognosis. Similarly, when considering cytogenetic risk factors in conjunction with cPC levels, comparable results were obtained. To evaluate whether the Revised International Staging System (R-ISS) groups could be further stratified based on different prognostic factors related to cPCs, our study revealed similar median PFS and OS rates in R-ISS II stage patients with cPCs ≥0.165% compared to those in the III stage (p=0.659 and 0.249, respectively). Conclusion: This study demonstrates that a high ratio of cPCs serves as a reliable indicator for predicting a poorer prognosis in MM cases. Furthermore, incorporating the R-ISS system and cytogenetic risk factors alongside the level of cPCs enhances the accuracy of prognostic predictions for patients with MM.

Plasmid-mediated quinolone resistance in extended-spectrum-ß-lactamase- and AmpC ß-lactamase-producing Serratia marcescens in China.

We investigated the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants and examined the association of these determinants with extended-spectrum ß-lactamases (ESBLs) and/or plasmid-mediated AmpC ß-lactamases (pAmpCs) in Serratia marcescens isolates in China. In this study, the presence of PMQR determinants was significantly related to the coproduction of ESBLs and/or pAmpCs (CTX-M-14, SHV-5, DHA-1, and ACT-1), among which CTX-M-14 was the most common gene type.

Efficacy and toxicity of SEAM (semustine, etoposide, cytarabine, and melphalan) conditioning regimen followed by autologous stem cell transplantation in lymphoma.

OBJECTIVES: The aim of this retrospective study was to evaluate the safety and efficacy of SEAM regimen followed by auto-SCT in lymphoma. PATIENTS AND METHODS: We retrospectively reviewed the records of patients with lymphoma who underwent auto-SCT with SEAM conditioning regimen from January 2010 to June 2018 at our centre. In total, 97 patients were analysed. RESULTS: The median time to neutrophil engraftment and platelet engraftment was 9.5 days (range, 7-15 days) and 12 days (range, 7-25 days), respectively. Grade 3-4 nausea/vomiting, mucositis and diarrhoea were observed in 21.6%, 36.1%, and 11.3% of patients, respectively. Treatment-related mortality at 100 days occurred in 2 patients (2.1%). After a median follow-up time of 53.9 months, the 3-year incidence of disease relapse or progression was 34%. The estimated progression-free survival and overall survival at 3 years were 62% and 75%, respectively. Compared with previous studies using BEAM as the conditioning regimen, this study shows that the SEAM regimen has a comparable efficacy and safety profile. CONCLUSIONS: The SEAM regimen is feasible and might be an ideal alternative to BEAM regimen for lymphoma auto-SCT.

Enhanced efficacy of imipenem-colistin combination therapy against multiple-drug-resistant Enterobacter cloacae: in vitro activity and a Galleria mellonella model.

BACKGROUND/PURPOSE: To investigate the in vitro and in vivo activity of imipenem-colistin combination against multidrug-resistant Enterobacter cloacae infections in order to determine whether it should be explored further. METHODS: The antimicrobial activity of colistin alone and in combination with imipenem was assessed versus an imipenem-susceptible isolate, E. cloacae GN1059, or an imipenem-resistant strain, E. cloacae GN0791, isolated in Anhui, China. The potential synergy of imipenem-colistin was evaluated using a checkerboard assay, as well as static time-kill experiments at 1× and 2× minimum inhibitory concentration (MIC). A simple invertebrate model (Galleria mellonella) was developed to assess the in vivo efficacy of imipenem-colistin in treating E. cloacae infection. RESULTS: In checkerboard assays, synergy (defined as a fractional inhibitory concentration index of ≤ 0.5) was observed between imipenem and colistin for both isolates tested. In time-kill assays, the combination of imipenem-colistin at 1× or 2× MIC resulted in complete killing of both strains. In the G. mellonella larvae model infected with lethal doses of E. cloacae, the combination therapy led to significantly increased survival of the larvae as compared with imipenem or colistin monotherapy alone (p < 0.05). CONCLUSION: This is the first report demonstrating the efficacy of antimicrobial agents in the G. mellonella larvae model of infections caused by E. cloacae. Our study suggested that imipenem-colistin combination was highly active against E. cloacae both in vitro and in the simple invertebrate model, and provided preliminary in vivo evidence that such combination might be useful therapeutically.

Synergy against extensively drug-resistant Acinetobacter baumannii in vitro by two old antibiotics: colistin and chloramphenicol.

Combination antimicrobial therapy is an important option in the fight against Gram-negative 'superbugs'. This study systematically investigated the synergistic effect of colistin (CST) and chloramphenicol (CHL) in combination against extensively drug-resistant Acinetobacter baumannii (XDR-AB). The microtitre plate chequerboard assay was used to test synergy against 50 XDR-AB clinical strains. Then, three XDR-AB clinical isolates and the type strain A. baumannii ATCC 19606 were chosen for further synergy studies using time-kill assay, mutant prevention concentration (MPC) assay and real-time population analysis profile (PAP) assay. In the chequerboard assays, synergistic or additive effects [defined as a fractional inhibitory concentration index (FICI) of ≤0.5 and 0.5 < FICI < 1, respectively] were observed in all 50 isolates. In further synergy testing, the results of time-kill assays indicated that CST monotherapy produced rapid bacterial killing followed by rapid re-growth, with the emergence of CST resistance; CHL monotherapy was largely ineffective. The combination CST/CHL, however, showed a synergistic effect and enhanced bacterial killing in the four tested strains. It also significantly delayed re-growth and suppressed the emergence of CST resistance. In the MPC assay, a decrease in MPCs for CST was observed in the two CST-susceptible strains. PAP assay showed that both CST-resistant strains were heteroresistant.

Activity of levofloxacin in combination with colistin against Acinetobacter baumannii: In vitro and in a Galleria mellonella model.

BACKGROUND/PURPOSE: Treatment of Acinetobacter baumannii infections is challenging owing to widespread multidrug-resistant A. baumannii (MDR-AB) and the lack of novel agents. Although recent data suggest that levofloxacin (LVX) may have unique activity against MDR-AB in combination with colistin (CST), further preclinical work is needed. METHODS: We used a A. baumannii type strain ATCC19606, a CST-resistant strain AB19606R, and two clinical isolates (GN0624 and GN1115) of MDR-AB to investigate the in vitro and in vivo efficacy of LVX-CST combination. Synergy studies were performed using the microtiter plate chequerboard assay and time-kill methodology. Inhibitory activity of antibiotics against biofilms and the mutant prevention concentrations were also studied in vitro. A simple invertebrate model (Galleria mellonella) has been used to assess the in vivo activity of antimicrobial therapies. RESULTS: The LVX-CST combination was bactericidal against the CST-susceptible clinical isolate (GN0624). In checkerboard assays, synergy (defined as a fractional inhibitory concentration index of < 0.5) was observed between CST and LVX in GN0624. The combination had antibiofilm properties on the preformed biofilms of four tested strains and could prevent the emergence of CST-resistant A. baumanni. Treatment of G. mellonella larvae infected with lethal doses of A. baumannii resulted in significantly enhanced survival rates when LVX was given with CST compared with CST treatment alone (p < 0.05). CONCLUSION: In summary, a synergistic or additive effect between CST and LVX was observed in vitro and in vivo against CST-susceptible A. baumannii strains, although not against CST-resistant ones.

Characterization of Extended-Spectrum ß-Lactamase Genes of Shigella flexneri Isolates With Fosfomycin Resistance From Patients in China.

BACKGROUND: The emergence of fosfomycin resistance and extended-spectrum ß-lactamase (ESBL) genes is a serious threat to public health and a new challenge in shigellosis treatment. The purpose of this study was to identify fosfomycin resistance and characterize ß-lactamase genes in fos-carrying isolates of Shigella flexneri from patients in China. METHODS: A total of 263 S. flexneri isolates were collected from 34 hospitals in the Anhui Province of China during September 2012-September 2015 and screened for fosA3, fosA, and fosC2 by PCR amplification and sequencing. The fos-carrying isolates were then screened for ß-lactamase genes. The clonal relationships between fosA3-carrying isolates, the transmissibility of fosfomycin resistance, replicon types of plasmids carrying fosfomycin resistance genes and other associated resistance genes were investigated. RESULTS: Twenty-five of the 263 isolates (9.5%) showed resistance to fosfomycin, and 18 (6.8%) were positive for fosA3. None of the isolates was positive for fosA or fosC2. Seventeen of the isolates carrying fosA3 (94%) were CTX-M producers (seven CTX-M-55, five CTX-M-14, and five CTX-M-123), while three (16.7%) were TEM producers (TEM-1).Sixteen (88.9%) fosA3-carrying isolates exhibited multi-drug resistance. The replicon types of the 13 fosA3-carrying plasmids were IncF (n=13), IncHI2 (n=3), IncIl-Ir (n=2), and IncN (n=1). CONCLUSIONS: Our results indicated that fosA3 could spread through plasmids in S. flexneri isolates, along with the bla(CTX-M) and bla(TEM), which facilitate its quick dispersal. To the best of our knowledge, this is the first report of CTX-M-123-type ESBLs in S. flexneri isolates from patients in China.

Galleria mellonella as an in vivo model for assessing the efficacy of antimicrobial agents against Enterobacter cloacae infection.

BACKGROUND/PURPOSE: Enterobacter cloacae is a well-recognized nosocomial pathogen. Use of a rapid, in vivo infection model for E. cloacae that can determine the efficacy of antibiotic therapies could help facilitate screening for new treatments. Nonmammalian model systems of infection, such as Galleria mellonella, have significant logistical and ethical advantages over mammalian models. MATERIALS AND METHODS: We utilized G. mellonella larvae to determine the utility of this infection model to study antibacterial efficacy. G. mellonella killing with heat-killed or live clinical isolates (E. cloacae GN1059 and GN0791) was tested. We also investigated the effect of postinoculation incubation temperature on the survival of infected larvae. The protection of administration of antibiotics to infected larvae was investigated. Finally, we determined the G. mellonella hemolymph burden of E. cloacae after administration of different antibiotics. RESULTS: With live bacterial inocula, G. mellonella killing was significantly dependent on the number of E. cloacae cells injected in a dose-dependent manner. Further, we observed that survival was reduced with increasing the postinoculation temperature. Treatment of a lethal E. cloacae infection with antibiotics that had in vitro activity significantly prolonged the survival of larvae compared with treatment with antibiotics to which the bacteria were resistant. The therapeutic benefit arising from administration of antibiotic correlated with a reduced burden of E. cloacae cells in the hemolymph. CONCLUSION: The G. mellonella infection model has the potential to be used to facilitate the in vivo study of host-pathogen interactions in E. cloacae and the efficacy of antibacterial agents.

Detection of the plasmid-mediated quinolone resistance determinants in clinical isolates of Acinetobacter baumannii in China.

We examined the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants among Acinetobacter baumannii in Anhui, China. And ß-lactamase genes and mutations in quinolone resistance-determining regions (QRDRs) were also investigated among the PMQR-positive isolates. Among the 39 A. baumannii isolates, 3 (7.7%) isolates harbored qnrB and 1 (2.6%) harbored qnrS. Mutations in the QRDRs of gyrA were identified in 2 isolates amongst the 4 PMQR-positive isolates. Quinolone resistance could be transferred by conjugation from all 4 PMQR-positive donors. In conclusion, more attention should be taken to prevent the transmission of PMQR genes among A. baumannii.

In vitro synergy of colistin combinations against extensively drug-resistant Acinetobacter baumannii producing OXA-23 carbapenemase.

Fifty extensively drug-resistant Acinetobacter baumannii (XDRAB) were isolated from patients. The chequerboard microdilution method was used to determine the in vitro activities of five colistin (COL)-based combinations including COL+fosfomycin (FOS), COL+rifampicin (RIF), COL+imipenem (IMP), COL+sulbactam (SUP) and COL+levofloxacin (LVX). The synergistic activity was evaluated by the fractional inhibitory concentration index (FICI). According to our results, the combination of COL was synergistic with FOS, RIF, IMP, SUP and LVX with the ratios of 50, 72, 88, 92 and 64%, respectively. When combined with COL, the other five agents showed increased antimicrobial activities. In addition, two of the combinations, COL+RIF and COL+IMP, were more active than the combinations of COL+FOS, COL+SUP and COL+LVX. More importantly, these combination regimens could exert synergistic effects at the sub-minimum inhibitory concentration (MIC) levels against XDRAB strains.

In vivo activity of vancomycin combined with colistin against multidrug-resistant strains of Acinetobacter baumannii in a Galleria mellonella model.

BACKGROUND: With increasing antibiotic resistance, the selection of effective treatment of A. baumannii infections is particularly challenging. METHODS: This study assessed the activities of the combination of vancomycin and colistin combination in vitro and in vivo using a Galleria mellonella model against four colistin-susceptible or colistin-resistant A. baumannii strains. RESULTS: In checkerboard assays, synergy was observed between vancomycin and colistin for all four strains tested (0.156 ≤ Fractional inhibitory concentration indices [FICI] ≤ 0.281). In time-kill assays, the combination showed continued bactericidal activity and synergy after 24 h for colistin-susceptible strains. For colistin-resistant strains, the combination resulted in bactericidal activity within 8 h, but sustained bacterial re-growth was then observed. Treatment of G. mellonella larvae infected with lethal doses of A. baumannii (except 19606R) resulted in significantly increased survival rates when vancomycin was given with colistin compared to colistin treatment alone (p < 0.05). CONCLUSIONS: These findings suggest that regimens containing vancomycin may be useful for infections due to multidrug-resistant Acinetobacter baumannii.

New Delhi Metallo-ß-Lactamase-Mediated Carbapenem Resistance: Origin, Diagnosis, Treatment and Public Health Concern.

OBJECTIVE: To review the origin, diagnosis, treatment and public health concern of New Delhi metallo-ß-lactamase (NDM)-producing bacteria. DATA SOURCES: We searched database for studies published in English. The database of PubMed from 2007 to 2015 was used to conduct a search using the keyword term "NDM and Acinetobacter or Enterobacteriaceae or Pseudomonas aeruginosa." STUDY SELECTION: We collected data including the relevant articles on international transmission, testing methods and treatment strategies of NDM-positive bacteria. Worldwide NDM cases were reviewed based on 22 case reports. RESULTS: The first documented case of infection caused by bacteria producing NDM-1 occurred in India, in 2008. Since then, 13 blaNDM variants have been reported. The rise of NDM is not only due to its high rate of genetic transfer among unrelated bacterial species, but also to human factors such as travel, sanitation and food production and preparation. With limited treatment options, scientists try to improve available therapies and create new ones. CONCLUSIONS: In order to slow down the spread of these NDM-positive bacteria, a series of measures must be implemented. The creation and transmission of blaNDM are potentially global health issues, which are not issues for one country or one medical community, but for global priorities in general and for individual wound care practitioners specifically.

In vivo activity of daptomycin/colistin combination therapy in a Galleria mellonella model of Acinetobacter baumannii infection.

Antimicrobial treatment of multidrug-resistant Acinetobacter baumannii (MDR-AB) infections continues to pose significant challenges. With limited options, clinicians have been pushed towards using unorthodox combinations of licensed antibiotics. Although daptomycin/colistin combination appears to be a promising treatment option based on in vitro data, further preclinical work is needed. In this study, the A. baumannii-Galleria mellonella system was employed to study the in vivo efficacy of this combination in order to determine whether it should be explored further for the treatment of MDR-AB infections. The antimicrobial activity of colistin alone and in combination with daptomycin was assessed versus an A. baumannii type strain (ATCC 19606) and a MDR-AB clinical strain (GN2231) isolated in Anhui, China. Synergy studies were performed using the microtitre plate chequerboard assay and time-kill methodology. The in vivo activity of daptomycin/colistin combination was assessed using a G. mellonella larvae model. The combination of daptomycin and colistin was bactericidal against both strains tested. In chequerboard assays, daptomycin was highly active against A. baumannii when combined with colistin [fractional inhibitory concentration index (FICI) of <0.5]. Treatment of G. mellonella larvae infected with lethal doses of A. baumannii resulted in significantly enhanced survival rates when daptomycin was given with colistin compared with colistin treatment alone (P<0.05). This work suggests that daptomycin/colistin combination is highly active against A. baumannii both in vitro and in a simple invertebrate model of infection.