Establishment of the reference interval for high-sensitivity cardiac troponin T in healthy children of Chongqing Nan'an district.

OBJECTIVE: To detect the concentration of high-sensitivity cardiac troponin T (hs-cTnT) in healthy children aged 0-14 years by electrochemiluminescence immunoassay (ECLIA), so as to explore the differences in different ages and genders. The aim of this study is to establish the reference interval for hs-cTnT in healthy children aged 0-14 years. METHODS: After screening, 3463 healthy children, including 1924 boys and 1539 girls, were selected from 4617 children aged 0-14 years. They were divided into nine groups: one day (umbilical cord blood; 'UCB'), one day (venous blood; 'VB'), 2-28 days, 29 days-<3 months, 3-<6 months, 6 months-<1 year old, 1-< 3 years old, 3-< 6 years old and 6-14 years old. A nonparametric test was used to detect the hs-cTnT concentration. The upper limit of the reference interval is the mean of the 99th percentile after bootstrap sampling. RESULTS: Hs-cTnT levels conformed to a non-Gaussian distribution. There was no significant difference in the concentration of hs-cTnT between boys and girls in the general data, but there were differences between boys and girls in the 3-<6 years old and 6-14 years old age groups. Except for UCB and 2-28 days, the concentration of hs-cTnT was significantly different in other age groups. The level of hs-cTnT in neonatal serum (2-28 days) was the highest. In other groups, it decreased gradually with age and dropped to the reference range of adults (0-14ng/L) at one-year old. The upper limit of reference interval of hs-cTnT concentration in each group was, respectively, 60.8, 78.8, 96.6, 58.6, 34.2, 16.2, 11.4, 8.0 (7.8 female), and 7.9 (7.3 female) ng/L. CONCLUSIONS: Referring to WS/T 402-2012 establishment of reference intervals for clinical laboratory testing projects and CLSI (Clinical and Laboratory Standards Institute) C28-A3 documents and the joint expert consensus of ESC (European Society of Cardiology) and ACC (American College of Cardiology) in 2007, we established the reference interval of hs-cTnT concentration in children aged 0-14 years in Chongqing Nan'an district of China which can provide certain reference value for clinical diagnosis and treatment of myocarditis and myocardial (micro) injury in children.

BCG-induced formation of neutrophil extracellular traps play an important role in bladder cancer treatment.

Bacillus Calmette-Guerin (BCG) is one of the most effective treatments for bladder cancer. Little attention has been paid to the possible role of neutrophils in BCG immunotherapy. In this study, we examined neutrophil extracellular traps (NETs) formation induced by BCG stimulation, and found that BCG-induced NETs exerted cytotoxicity, induced apoptosis and cell-cycle arrest, and inhibited migration in bladder tumor cells. BCG-activated tumor cells but not non-activated ones elicited NETs formation, in which IL-8 and TNF-α from activated tumor cells both took effect. Moreover, NETs activated peripheral blood mononuclear cells (PBMCs) exhibited a higher expression of CD4 and Th1 cytokines. Additionally, the role of NETs in vivo contributed to the recruitment of T cells and monocytes-macrophages and tissue damage, thus preventing tumor growth. NETs proteins mainly caused these effects on tumor and cellular immunity. In conclusion, we demonstrated a novel immunoregulatory role for NETs in the early stages of BCG immunotherapy.

Down-regulation of Sphk2 suppresses bladder cancer progression.

Bladder cancer is the second most common urological malignancy around the world and is by far the most frequent urological malignancy in China. The abnormal expression of sphingosine kinase 2 (SphK2) is associated with tumor progression and a poor patient survival rate, however, the effect of SphK2 on the bladder cancer cells remains unclear. The aim of the paper was to study the expression of SphK2 in bladder cancer and the role of SphK2 on the cell proliferation, metastasis, and apoptosis in bladder cancer in vitro. Our results showed that SphK2 is up-regulated in bladder cancer tissues compared with the corresponding adjacent non-neoplastic tissues, and the expression level of SphK2 was significantly higher in human bladder cancer cells in comparison with normal bladder epithelial cells. Silencing of SphK2 could inhibit the proliferation ability of T24 cells in vitro. In addition, SphK2 knockdown could induce a significant increase in the number of apoptotic cells. Furthermore, the transwell assay also showed significant cell migration inhibition in SphK2 siRNA transfectant compared with cell lines transfected with NC. Thus, this study suggested that SphK2 inhibition may provide a promising treatment for bladder cancer patients.

Retraction: Graphene oxide/DNA-decorated electrode for the fabrication of microRNA biosensor.

[This retracts the article DOI: 10.1039/C5RA12373A.].

[Intravesical perfusion of recombinant adeno-associated virus-endostatin in treatment of bladder cancer: experiments with mice].

OBJECTIVE: To study the anti-tumor effect of intravesical perfusion of recombinant adeno-associated virus-endostatin (rAAV-ES) in treatment of bladder cancer. METHODS: Forty-five C57BL/6 mice underwent intravesical perfusion of mouse bladder cancer cells of the line MB49 so as to establish orthotopic murine bladder cancer models and were divided into 3 equal groups, 3 days later to undergo intravesical perfusion of rAAV-ES, rAAV-EYFP, and PBS respectively once per week for 6 times. The anti-tumor effect of rAAV-ES on the tumor bearing mice was studied. RESULTS: The tumor weight of the rAAV-ES group was (145 +/- 30) mg, significantly lighter than those of the rAAV-EYFP and PBS groups [(250 +/- 32) mg and (250 +/- 30) mg respectively, both P < 0.05]. The survival time of the rAAV-ES-treated mice was (46 +/- 7) d, significantly longer than those of the rAAV-EYFP- and PBS-treated groups [(38 +/- 7) d and (38 +/- 6) d respectively, both P < 0.05]. CONCLUSION: An effective biologic agent in bladder cancer gene therapy, intravesical treatment with rAAV-ES inhibits the angiogenesis, thus inhibiting the tumor formation and progression.

Biological effects of eukaryotic recombinant plasmid pReceiver-M61-BAI-1 transfection on T24 cells and HUVECs.

The aim of the current study was to investigate the biological effect on T24 cells and human umbilical vein endothelial cells (HUVECs) of transfection with brain-specific angiogenesis inhibitor-1 (BAI-1). The recombinant plasmid pReceiver-M61-BAI-1 was transfected into human superficial bladder tumor cells (T24) and HUVECs, in parallel with the vector control. mRNA and protein expression levels of BAI­1 were then detected by quantitative polymerase chain reaction (qPCR) and western blotting, respectively. Cell apoptosis of T24 cells and HUVECs prior and subsequent to transfection with BAI­1 was analyzed by flow cytometric analysis. Proliferation of T24 cells and HUVECs prior and subsequent to transfection of BAI-1 was assessed by the MTT method. T24 cells and HUVECs transfected with pReceiver­M61­BA1­1 were classed as the experimental group; T24 cells and HUVECs transfected with p­Receiver­M61 were the control group. qPCR and western blotting methods confirmed that there was positive expression of BAI­1 in T24 cells and HUVECs transfected with pReceiver­M61­BAI­1, however BAI­1 was not expressed in T24 cells and HUVECs transfected with pReceiver­M61. The results of the MTT assay demonstrated that absorbance was markedly reduced in HUVECs at 12, 48 and 72 h subsequent to transfection with pReceiver-M61-BAI-1 when compared with that of the control group and in T24 cells transfected with p­Receiver-M61-BAI-1. Furthermore, flow cytometry results also indicated that the apoptotic rate of HUVECs transfected with p­Receiver­M61­BAI­1 was significantly increased compared with that of the control group and T24 cells transfected with p­Receiver­M61­BAI­1. BAI­1 was observed to markedly inhibit the proliferation of vascular endothelial cells in vitro, however, no direct inhibition by BAI­1 was observed in T24 cells. In conclusion, BAI-1 is suggested to be a potential novel therapautic target for the inhibition of tumor neovascularization.

Antimicrobial susceptibility of hospital acquired Stenotrophomonas maltophilia isolate biofilms.

AIMS: We sought to characterize the antibiotic susceptibility of strains of Stenotrophomonas maltophilia isolated from clinical samples, and the role of Stenotrophomonas maltophilia biofilm in antibiotic resistance. METHODS: Fifty-one clinical Stenotrophomonas maltophilia isolates were obtained from patients with nosocomial infection in the surgical wards and ICUs of six general hospitals in Tianjin, China. In vitro models of Stenotrophomonas maltophilia biofilms were established and confirmed by scanning electron microscopy and fluorescence microscopy with silver staining. The minimal inhibitory concentrations and biofilm inhibitory concentrations of commonly used antibiotics were determined. RESULTS: 47 of 51 strains were resistant to three or more antibiotics. 42 of 51 strains formed Stenotrophomonas maltophilia biofilms in vitro. Stenotrophomonas maltophilia biofilm formation greatly reduced sensitivity to most tested antibiotics, but not to levofloxacin. However, in the presence of erythromycin scanning electron microscopy revealed that levofloxacin inhibited Stenotrophomonas maltophilia biofilm formation. Factorial ANOVA revealed that erythromycin enhanced susceptibility to levofloxacin, cefoperazone/sulbactam, and piperacillin (p<0.05), and an ΔE model revealed that levofloxacin and erythromycin acted synergistically in biofilms, suggesting specific use of combined macrolide therapy may represent an effective treatment for Stenotrophomonas maltophilia infection. CONCLUSIONS: Antibiotics could act synergistically to combat the protection conferred to clinical isolates of Stenotrophomonas maltophilia by biofilms. Macrolide antibiotics may be effective where used in combination.

Expression of brain­specific angiogenesis inhibitor­1 and association with p53, microvessel density and vascular endothelial growth factor in the tissue of human bladder transitional cell carcinoma.

The aim of the present study was to investigate the expression levels of brain­specific angiogenesis inhibitor­1 (BAI­1) in bladder transitional cell carcinoma (BTCC) at different stages and the mechanism by which it inhibits tumor endothelial cell proliferation. Normal bladder mucosa biopsy specimens were obtained as the control group, and human BTCC biopsy specimens were used as the study group. Immunohistochemical assays were used to detect the expression levels of BAI­1, vascular endothelial growth factor (VEGF) and mutant p53, in addition to microvessel density (MVD) in the tissues. Western blotting was used to analyze the differential expression of BAI­1 in the two samples. Statistical analysis was performed, which indicated that BAI­1 expression levels in the normal bladder mucosa group were significantly higher than those in the BTCC group and were associated with clinical staging. BAI­1 levels in the T1 stage BTCC tissues were higher than those in the T2­4 stage BTCC tissues (P<0.05). BAI­1 expression levels were negatively correlated with those of VEGF (r=­0.661, P<0.001), mutant p53 (r=­0.406, P=0.002) and with the MVD (r=­0.675, P<0.001). BAI­1 may be involved in the negative regulation of BTCC microvascular proliferation, and its expression may be associated with a reduction in p53 mutations.

microRNA-21 Regulates Cell Proliferation and Migration and Cross Talk with PTEN and p53 in Bladder Cancer.

This study aimed to determine the molecular mechanism by which the oncogenic micoRNA-21 (miR-21) functions in bladder cancer. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that the expression of miR-21 considerably increased in primary cancer tissue compared with that in the paired adjacent noncancerous tissue and that in normal bladder mucosa. Knockdown of miR-21 by using antisense oligonucleotide significantly suppressed the proliferation and migration of bladder cancer cells (J82 and RT112). Mechanism studies showed that downregulation of miR-21 resulted in cell cycle arrest at the G1 phase and upregulation of the tumor suppressor PTEN (phosphatase and tensin homologue) and p53 phosphorylation at Ser46. The p53 phosphorylation at Ser15 and the whole level of p53 acetylation remained unchanged in response to miR-21 knockdown. MicroRNA-21 regulates proliferation and migration of bladder cancer cells and cross talk with PTEN and p53 in bladder cancer.

Recombinant hIFN-α2b-BCG inhibits tumor growth in a mouse model of bladder cancer.

Bacillus Calmette-Guérin (BCG) reduces the recurrence and progression of non-muscle invasive bladder cancer. The present study aimed to investigate the impact of a recombinant hIFN-α2b-secreting BCG (rBCG) on the mouse bladder MB49 cell line and an orthotopic mouse model of bladder cancer. MB49 cells were cultivated in the presence or absence of rBCG, BCG or BCG+hIFN-α2b. Cellular morphology and viability were assessed by microscopy and CCK-8 assay, respectively. Apoptosis was assessed by acridine orange, Hoechst 33258 staining and flow cytometry. MHC-I expression was assessed by flow cytometry. MB49 cells were transplanted into the bladders of C57BL/6 mice administered BCG, rBCG or BCG+hIFN-α2b. Local tissue Fas expression and T cell subsets were assessed by immunohistochemistry. Peripheral blood TNF-α and IL-12 levels were measured by ELISA, and circulating T lymphocyte subsets by flow cytometry. BCG, rBCG and BCG+hIFN-α2b increased the distortion and death of MB49 cells, yet rBCG reduced the proliferation and enhanced apoptosis most substantially. Apoptosis was increased after a 24-h co-culture with rBCG or BCG+hIFN-α2b. Mice administered rBCG survived longer than mice administered BCG (p<0.001), yet this result was not significantly different from mice administered BCG+hIFN-α2b. The average bladder weight was reduced by administration of rBCG (p<0.001). Fas expression and peripheral blood mTNF-α and mIL-12, cell counts of polymorphonuclear leukocytes, monocytes, T lymphocytes and CD4+/CD8+ ratios were significantly increased by all BCG treatments (p≤0.05), yet monocyte and T lymphocyte counts were higher in mice administered rBCG than in mice treated with BCG or BCG+hIFN-α2b (p=0.000). These results indicate that in an orthotopic murine bladder cancer model rBCG possesses superior antitumor activity to BCG+hIFN-α2b.

Human telomerase reverse-transcriptase promoter-controlled and herpes simplex virus thymidine kinase-armed adenoviruses for renal cell carcinoma treatment.

New treatment strategies are required for renal cell carcinoma (RCC) due to its relative insensitivity to conventional radio- and chemotherapies. The promising strategy of tumor inhibition using human telomerase reverse transcriptase (hTERT)-controlled herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/GCV) in the hTERT promoter-driven HSV-TK/GCV suicide gene system was investigated. Tumor volume, weight, relative proliferation rate, and cell-apoptosis levels were examined in mice injected with adenovirus (Ad)-hTERT-HSV-TK and GCV. Increased cell death occurred following treatment with Ads carrying hTERT-HSV-TK/GCV or cytomegalovirus promoter-controlled (CMV)-HSV-TK/GCV for human RCC 786-0 and fibroblast MRC-5 cells. In mice, Ad-hTERT-HSV-TK/GCV more specifically inhibited tumor and RCC xenograft growth than Ad-CMV-HSV-TK/GCV (P < 0.05). Furthermore, Ad-hTERT-HSV-TK/GCV did not significantly damage normal fibroblasts or organ systems (heart, lung, liver, brain, kidney, and spleen). Thus, Ad-hTERT-HSV-TK/GCV is an effective RCC inhibitor in human cells in vitro and in vivo mouse models, indicating potential usefulness in RCC-targeted gene therapy.

Antimicrobial susceptibility of hospital acquired Stenotrophomonas maltophilia isolate biofilms

Abstract Aims We sought to characterize the antibiotic susceptibility of strains of Stenotrophomonas maltophilia isolated from clinical samples, and the role of Stenotrophomonas maltophilia biofilm in antibiotic resistance. Methods Fifty-one clinical Stenotrophomonas maltophilia isolates were obtained from patients with nosocomial infection in the surgical wards and ICUs of six general hospitals in Tianjin, China. In vitro models of Stenotrophomonas maltophilia biofilms were established and confirmed by scanning electron microscopy and fluorescence microscopy with silver staining. The minimal inhibitory concentrations and biofilm inhibitory concentrations of commonly used antibiotics were determined. Results 47 of 51 strains were resistant to three or more antibiotics. 42 of 51 strains formed Stenotrophomonas maltophilia biofilms in vitro. Stenotrophomonas maltophilia biofilm formation greatly reduced sensitivity to most tested antibiotics, but not to levofloxacin. However, in the presence of erythromycin scanning electron microscopy revealed that levofloxacin inhibited Stenotrophomonas maltophilia biofilm formation. Factorial ANOVA revealed that erythromycin enhanced susceptibility to levofloxacin, cefoperazone/sulbactam, and piperacillin (p < 0.05), and an ΔE model revealed that levofloxacin and erythromycin acted synergistically in biofilms, suggesting specific use of combined macrolide therapy may represent an effective treatment for Stenotrophomonas maltophilia infection. Conclusions Antibiotics could act synergistically to combat the protection conferred to clinical isolates of Stenotrophomonas maltophilia by biofilms. Macrolide antibiotics may be effective where used in combination.