Evolution of carbapenem resistance in klebsiella pneumoniae and escherichia coli carrying blaNDM-1 gene: imipenem exposure results in sustained resistance memory of strains in vitro.

BACKGROUND: Antibiotics exert an outstanding selective pressure on bacteria, forcing their chromosomal gene mutations and drug resistance genes to spread. The objective of this study is to evaluate the expression of the New Delhi Metallo-ß-Lactamase-1 gene (blaNDM-1) in the clinical isolate (Klebsiella pneumoniae TH-P12158), transformant strains Escherichia coli BL21 (DE3)-blaNDM-1, and Escherichia coli DH5α- blaNDM-1 when exposed to imipenem. METHODS: ß-Lactamase genes (blaSHV, blaTEM-1, blaCTX-M-9, blaIMP, blaNDM-1, blaKPC, blaOXA, blaGES, and blaDHA) from randomly selected carbapenems-sensitive K.pneumoniae (n = 20) and E.coli (n = 20) strains were amplified by PCR. The recombinant plasmid of pET-28a harboring blaNDM-1 was transformed into E.coli BL21 (DE3) and E.coli DH5α by electroporation. The resistance phenotype and higher blaNDM-1 expression in K.pneumoniae TH-P12158, transformant E.coli BL21 (DE3)-blaNDM-1, and E.coli DH5α-blaNDM-1 were observed when exposed to imipenem with grade increasing, decreasing, and canceling doses, respectively. RESULTS: After being exposed to different doses of imipenem, the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of antimicrobial drugs and blaNDM-1 expression of strains increased, which was positively correlated with doses of imipenem. On the contrary, with the decrease or cancellation of imipenem doses, the blaNDM-1 expression was deteriorated, while the MIC and MBC values remained relatively stable. These results demonstrated that low doses of imipenem (˂MIC) could press blaNDM-1 positive strains producing stable drug resistance memory and altered blaNDM-1 expression. CONCLUSIONS: Low doses of imipenem could press blaNDM-1 positive strains producing sustained resistance memory and altered blaNDM-1 expression. In particular, the positive correlation between the resistance genes expression and antibiotics exposure shows promising guiding significance for clinical medication.

Downregulation of the paired box gene 3 inhibits the progression of skin cutaneous melanoma by inhibiting c-MET tyrosine kinase : PAX3 downregulation inhibits melanoma progression.

BACKGROUND: The PAX3 (paired box gene 3) gene is highly expressed in several cancer types. However, its underlying mechanism of action in skin cutaneous melanoma (SKCM) remains unknown. METHODS: In this study, we used the GEPIA database and western blotting to analyze the expression of PAX3. We performed the Kaplan-Meier survival analysis to evaluate the prognostic value of PAX3 in SKCM. Next, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed to evaluate the function of PAX3-related co-expressed genes. Additionally, the function and potential mechanism of action of PAX3 in SKCM were studied through functional experiments. Western blotting was used to detect the changes in the levels of epithelial-mesenchymal transition (EMT)-related and MET (c-MET tyrosine kinase) proteins following PAX3 knockdown. Finally, we assessed the correlation between PAX3 expression and the infiltration of CD4+/CD8+ T cells using the TISIDB database. RESULTS: We found that PAX3 was overexpressed in the SKCM tissues and that these levels were indicative of a poor prognosis of SKCM. The KEGG pathway enrichment analysis showed that PAX3-related co-expressed genes were mainly associated with the oncogenic pathways. Knocking down PAX3 significantly inhibited the proliferation, invasion, and migration of SK-MEL-28 cells. The PAX3 expression was related significantly to the immune infiltration level of CD4+/CD8+ T cells. CONCLUSIONS: Our findings demonstrated that PAX3 knockdown could reverse the EMT of tumor cells, inhibit the growth, and progression of SKCM cells. Therefore, PAX3 may have implications as a potential therapeutic target and promising prognostic biomarker for SKCM.

Validity and reliability of the Chinese version of the Patient-Reported Outcomes Measurement Information System adult profile-57 (PROMIS-57).

BACKGROUND: The PROMIS-57 is a commonly used self-reported instrument to solve the lack of generalizable and universal measures required to evaluate common symptoms and functions from patients' perspectives. This study aimed to translate the PROMIS-57 into Chinese and psychometrically test the translated instrument on patients with breast cancer. METHODS: Translation, cross­cultural adaptation, and psychometric evaluation of the instrument were performed from June 2020 to June 2021. Eligible patients were recruited and completed the PROMIS-57, Functional Assessment of Cancer Therapy-Breast (FACT-B), and a sociodemographic questionnaire. RESULTS: Data from 602 patients with a mean age of 48.83 years were analyzed. Most domains in the PROMIS-57 showed an absence of floor and ceiling effects. Multi-trait scaling analysis demonstrated acceptable convergent and discriminant validity. The correlations between the PROMIS-57 scores and the selected FACT-B scores supported the criterion validity via the Pearson correlation test. Measurement invariance was supported by the absence of differential item functioning for most items. Cronbach's α of the domains ranged from 0.85-0.95. The unidimensional factor structure of all domains was supported using confirmatory factor analyses. Additionally, most items showed acceptable item information curves and item characteristics curve matrices. CONCLUSION: The Chinese version of the PROMIS-57 was found to be a reliable and valid tool for assessing common symptoms and functions among patients with breast cancer.

[PreS1 Antigen of HBV Down-Regulates MHC-â on the Surface of Hepatocytes and Promotes Hepatocarcinogenesis].

Objective: To explore the internal mechanism of hepatocellular carcinoma (HCC) induced by chronic hepatitis B virus (HBV) infection. Methods: L02, HepG2 and Huh7 cells stably overexpressing HBV preS1 antigen were analyzed by flow cytometry, qRT-PCR and tumorigenesis in nude mice to evaluate the effect of preS1 antigen in HBV-related hepatocarcinogenesis. Results: Our results showed that the expression of cancer stem cell (CSCs) related factors and cell surface markers in preS1 overexpressing cells were up-regulated, and the tumorigenicity of these cells was enhanced in nude mice. In addition, preS1 overexpression could down-regulate the expression of major histocompatibility complex Ⅰ (MHC-Ⅰ). The expression of MHC-Ⅰ on the cell surface could be restored by adding interferon gamma (IFN-γ) in the process of cell culturing and the tumorigenicity of cells in nude mice could thus be reduced. Conclusion: Based on the above results, we believe that preS1 is a carcinogen of HBV and that it promotes the formation of liver cancer through down regulating MHC-Ⅰ on the surface of hepatocytes.

Prevalence of Carbapenem-Resistant Klebsiella Pneumoniae (CRKP) and the Distribution of Class 1 Integron in Their Strains Isolated from a Hospital in Central China.

Objective The aim of our study is to investigate the prevalence of Carbapenem-resistant Klebsiella pneumoniae (CRKP) and the genetic characteristics of the class 1 integron in CRKP on multi-drug resistance.Methods Clinical Klebsiella pneumoniae strains were collected from multiple departments of a hospital in central China. CRKP strains were identified among the isolates, and antibiotics susceptibility of CRKP strains was analyzed. The polymerase chain reaction (PCR) was adopted to amplify the class 1 integron variable area. The integron genetic structure was analyzed with enzyme digestion and DNA sequencing technology. The relation between class 1 integron and drug resistance was analyzed statistically.Results Totally 955 strains of Klebsiella pneumoniae were isolated from varied sites of the hospital, and 117(12.3%) of them were identified as CRKP, with a separation rate of 8.9% (26/292) in 2013, 11.3% (38/336) in 2014 and 16.2% (53/327) in 2015, which shows an increasing trend by year. 44.4% (52/117) of CRKP strains were separated from specimen of ICU, and 61.5% (72/117) were from sputum. Over 95% CRKP strains were resistant to ampicillin/sulbactam, aztreonam, imipenem, meropenem, ceftazidme, cefotaxime, cefepime,and piperacillin, while relatively low resistant rates were found in tigecycline (12.8%) and colistin (35.9%). The class 1 integron was detected in 77.8% (91/117) of CRKP strains. Class 1 integron of CRKP was significantly correlated with the antibiotic resistance to the tobramycin, gentamicin and amikacin (all P<0.01). The gene cassette analysis of variable area of class 1 integron showed that aadA2 accounts for 64.8% (59/91), aacA4-catB8-aadA1 23.1% (21/91), and aadA2-dfrA25 12.1% (11/91).Conclusions CRKP has an increasing trend in a clinical setting in China, and most of them were resistant to multiple antibiotics. Class 1 integron in CRKP has strong ability to capture the genes resistant to aminoglycosides antibiotics from environment, with the aadA2 gene as the most popular one.

Effects of Luteolin-7-O-Glucoside on Intestinal Microbiota Dysbiosis and Drug Resistance Transmission Caused by Raoultella ornithinolytica B1645-1: Modulating the Composition of Intestinal Microbiota and Promoting the Transfer of blaNDM-1 Gene from Genus Enterococcus to Lactobacillus in Mice.

Raoultella ornithinolytica is an Enterobacteriaceae bacterium that can infect both humans and animals, while luteolin-7-O-glucoside (IOG) is a flavonoid that has broad effects on the intestinal microbiota of healthy animals. However, current studies lack sufficient data on intestinal microbiota dysbiosis and drug resistance transmission caused by R. ornithinolytica and the possible role of IOG. In this study, BALB/c mice were infected with R. ornithinolytica carrying blaNDM-1 gene and treated with IOG (3 mg/kg·d and 6 mg/kg·d) to analyze the diversity of intestinal microbiota and the transfer of blaNDM-1 between bacteria. The findings indicated that R. ornithinolytica B1645-1 exhibited a significant ability to enhance the Firmicutes/Bacteroidota ratio and increase the relative abundance of Lactobacillus and Bacillus after 48 h, where as 6 mg/kg·d IOG had an opposite effect. Moreover, R. ornithinolytica B1645-1 facilitated the emergence of drug-resistant bacteria and promoted blaNDM-1 gene transfer in Enterococcus, Escherichia, Klebsiella, Acinetobacter, Bacillus, Brevibacterium, and Lactobacillus. Enterococcus was the predominant genus at 48 h. Surprisingly, 6 mg/kg·d IOG significantly inhibited the production of drug-resistant bacteria and promoted blaNDM-1 gene transfer from Enterococcus to Lactobacillus at 144 h. However, the role of Lactobacillus as a recipient for drug-resistant genes should be of more concern.

Classification of symptom subtypes in patients with multiple myeloma during treatment: a cross-sectional survey study in China.

OBJECTIVES: To classify subgroups of cancer-related symptoms in patients with multiple myeloma (MM) during treatment and examine between-group differences in demographic and clinical characteristics in addition to functional status. DESIGN: Cross-sectional survey study. SETTING: Haematology department of two tertiary hospitals affiliated with Guilin Medical University in China. PARTICIPANTS: Using a convenience sampling method, questionnaires were distributed to patients with MM visiting two hospitals in Guilin, China. INTERVENTIONS: The patients were categorised into subgroups based on cancer-related symptoms using a latent class analysis. An analysis of covariance was performed to examine how demographic and clinical characteristics and functional status differed among the subgroups. RESULTS: In total, 216 patients completed the survey, with an average age of 60.3 years. A three-class solution was identified: low symptom burden group (class 1, 36.6%), moderate symptom burden group (class 2, 34.2%) and high symptom burden group (class 3, 29.2%). Patients with low monthly family income (OR=3.14, p=0.010) and complications of MM bone disease (OR=2.95, p=0.029) were more likely to belong to class 2. The predictors of high-burden symptoms were treated with painkillers, antidepressants or hypnotic drugs (OR=3.68, p=0.012) and <5000 daily step counts (OR=2.52, p=0.039) in class 3. Functional status was correlated with symptom burden, with patients in classes 3 and 1 reporting significantly higher and lower functional status, respectively (p<0.05). CONCLUSIONS: Patients with MM experienced varying degrees of symptoms during treatment. The identification of patients with high symptom burden management should focus on the assessment of demographic and clinical characteristics, in addition to functional status.

Heterogeneity of symptoms and functions among women receiving chemotherapy for breast cancer in China: A multicentre, cross-sectional study.

Background: Currently, few studies have explored the heterogeneity of symptoms and functions in patients with breast cancer. This study aimed to identify the subgroups of symptoms and functions in women receiving chemotherapy for breast cancer and determine whether the subgroups differed in demographic and clinical characteristics. Methods: A cross-sectional multicenter survey involving five hospitals in Zhejiang, Shanghai, Shandong, and Guangxi provinces of Mainland China was implemented between August 2020 to December 2021. Participants completed questionnaires that included the PROMIS-57, PROMIS cognitive function short form, and demographic and clinical characteristics. Latent class analysis was performed, followed by chi-square test and analysis of variance. Subsequently, significant variables were included in multinomial logistic regression. Results: A total of 1,180 patients were investigated, with an average age of 48.9 years. Three classes were identified: low symptom burdens and functions group (26.2%, Class 1), moderate symptom burdens and functions group (16.9%, Class 2), and low symptom burdens and high functions group (56.9%, Class 3). Compared with patients in Class 1 and 3, those in Class 2 consistently showed a higher tendency of having urban employee health insurance (odds ratio = 2.506, P < 0.05) and rural health insurance (odds ratio = 2.207, P < 0.05). Additionally, patients in Class 2 tended to be in their fourth cycle of chemotherapy. However, receiving chemotherapy and surgery increased the likelihood of belonging to Class 1. Conclusions: A high proportion of patients experienced varying degrees of symptom and function issues, suggesting that attention is warranted for women with breast cancer undergoing chemotherapy. Patients with the urban employee basic medical system, the new rural cooperative medical system and in the early stage of chemotherapy cycles were more likely to have symptom burdens. Middle-aged postmenopausal women reported varying degrees of cognitive issues. Additionally, surgery increased the presence of potential long-term effects in functional levels.

Occurrence of two NDM-1-producing Raoultella ornithinolytica and Enterobacter cloacae in a single patient in China: probable a novel antimicrobial resistance plasmid transfer in vivo by conjugation.

OBJECTIVES: To identify the general features of acquisition of drug-resistance genes in two multi-drug resistant Enterobacteriaceae strains isolated from a single patient in China. METHODS: The whole-plasmid was sequenced by Illumina Hiseq 4000 and Pacbio RSII procedures. The plasmid conjugation transfer experiment were performed by the mating-out assay. Drug-resistance genes were amplified by PCR assay. RESULTS: We identified two New Delhi metallo-ß-lactamase type 1(NDM-1)-producing isolates, named Raoultella ornithinolytica B1645-1 and Enterobacter cloacae B1645-2, which shared the same sulfonamide-resistant dihydropteroate synthase sul2 gene and aminoglycoside O-phosphotransferase aph(3'')-Ib gene. A novel antimicrobial resistance plasmid pCYNDM01 was first discovered from the multi-drug resistant R. ornithinolytica B1645-1. Interestingly, plasmid pCYNDM01 carried a Gifsy-2 prophage gene. The blaNDM-1 gene was located on a novel complex class 1 integron with a structure of sul1-qacEΔ1-ΔISAba125-blaNDM-1-blaMBL-trpC-ISCR1-catb8-aacA4-IS1-IS6100-dfrA14-intI1. The carrying the blaNDM-1 gene plasmid pCYNDM01 was transferred to the E. cloacae B1645-2 recipient strain. This 149.44 kb plasmid pCYNDM01 belonged to the IncFII type. CONCLUSIONS: A novel antimicrobial resistance plasmid pCYNDM01 was first recovered from a multi-drug resistance R. ornithinolytica B1645-1 isolated from China. The novel complex sul1-type class 1 integron might play an essential role in the mobilization of the blaNDM-1 gene among different enterobacterial species. The occurrence of plasmid pCYNDM01 transfer from R. ornithinolytica to E. cloacae in vitro by conjugation showed that plasmid pCYNDM01 was a self-conjugative plasmid and might cause dissemination of drug-resistance genes within different enterobacterial species from a single patient in vivo by conjugation. The novel variant F-like T4SS of plasmid pCYNDM01 might be as a tool of R. ornithinolytica B1645-1 for resistance genes transfer. The emergence of the two NDM-1-producing Enterobacteriaceae strains should be attracted China attentions and required to prevent its future prevalence.

Reversal of the Warburg effect with DCA in PDGF­treated human PASMC is potentiated by pyruvate dehydrogenase kinase­1 inhibition mediated through blocking Akt/GSK­3ß signalling.

There is accumulating evidence indicating that the growth inhibitory effect of dichloroacetate (DCA) on pulmonary arterial smooth muscle cells (PASMCs) may be associated with the reversal of the Warburg effect and initiation of the mitochondria­dependent apoptotic pathway. Previous studies indicated that platelet­derived growth factor (PDGF) promoted the Warburg effect and resulted in apoptotic resistance of PASMCs, which was attributed to activation of the phosphatidylinositol 3­kinase (PI3K)/protein kinase B (Akt) signalling pathway. However, the mechanism underlying the pro­apoptotic effect of DCA on PDGF­treated PASMCs has not been thoroughly elucidated, and the effect of the Akt/glycogen synthase kinase­3ß (GSK­3ß) pathway inhibition concomitant with the effect of DCA on PASMC proliferation remains unclear. The growth of human PASMCs and the lactate concentration in extracellular medium of PASMCs were detected by Cell Counting Kit­8 assays and a Lactate Colorimetric Assay kit, respectively. Cell apoptosis was evaluated by fluorescence activated cell sorting. The mitochondrial membrane potential (ΔΨm) was assessed with 5,5',6,6'­tetrachloro­1,1',3,3'­tetraethylbenzimidazol­carbocyanine iodide assays. The expression levels of phosphorylated Akt and GSK­3ß, pyruvate dehydrogenase, cleaved caspase­3, pyruvate dehydrogenase kinase­1 (PDK­1), hypoxia inducible factor­1α (HIF­1α) and hexokinase­2 (HK­2) were measured with western blot analysis. Confocal analyses were employed to determine HK­2 co­localisation with the mitochondria. The results indicated that DCA inhibited human PASMC proliferation in a dose­dependent manner. DCA at 10 mM promoted apoptosis and the upregulation of activated caspase­3 in PASMCs pre­treated with 20 ng/ml PDGF­homeodimer BB (BB). Treatment with 5 µM LY294002 produced minimal anti­proliferative effects on human PASMCs and barely induced cellular apoptosis and caspase­3 activation. However, co­administration of 10 mM DCA with LY294002 significantly decreased the cell proliferation index and induced cell apoptosis and caspase­3 activation. The combined administration of LY294002 with DCA significantly decreased lactate concentration, promoted the depolarisation of the ΔΨm and repressed HIF­1α upregulation and HK­2 activation in PASMCs treated with PDGF, which was attributed to the potentiation of DCA­induced PDK­1 inhibition by LY294002 via blockade of the Akt/GSK­3ß/HIF­1α signalling pathway. In conclusion, inhibition of the Akt/GSK­3ß pathway improved the pro­apoptotic effect of DCA on human PASMCs, which may be attributed to a reversal of the Warburg effect by blocking the mutual interaction between HIF­1α and PDK­1, consequently downregulating HK­2. Therefore, combinatory treatment with DCA and PI3K inhibitors may represent a novel therapeutic strategy for the reversal of apoptosis resistance exhibited by PASMCs as a result of mitochondrial bioenergetic abnormalities, as well as the treatment of pulmonary vascular remodelling in pulmonary arterial hypertension.

The photophysical properties of a julolidene-based molecular rotor.

The photophysical properties of 9-dicyanovinyljulolidine are sensitive to solvent viscosity but are little affected by changes in polarity. In fluid solution, the lifetime of the first-excited singlet state is very short and triplet state formation cannot be detected by laser flash photolysis. Decay of the excited singlet state is strongly activated and weak phosphorescence can be observed in a glassy matrix at 77 K. Temperature dependent 1H NMR studies indicate that the molecule undergoes slow internal rotation in solution, for which the activation energy has a value of ca. 35 kJ mol(-1). This process is unlikely to account for the poor fluorescence quantum yield found in fluid solution. Instead, it is considered that the target compound undergoes rapid rotation around the dicyanovinyl double bond from the excited singlet state. The rate of rotation depends weakly on the viscosity of the solvent in a range of linear alcohols at room temperature. This might represent the fact that the rotor is relatively small and can pack into cavities in the solvent structure. In glycerol, the rate of rotation is more sensitive to viscosity effects but a quite complex temperature dependence is observed in ethanol. Here, the rate is almost activationless in a glassy matrix and in fluid solution at high temperature but strongly activated at intermediate temperatures.