Effect of regulating miRNA141 expression on malignant biological behaviors of human prostate cancer DU145 cells / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To study the effect of microRNA-141(miR-141) expression regulation on cell proliferation, cell cycle, apoptosis, invasion and migration of human prostate cancer cell line DU145 and its mechanism. Methods: MiR-141 mimics (miR-141 up group) and miR-141 inhibitors (miR-141 down group) were transfected into human prostate cancer DU145 cells by using liposome lipofectamine 2000, and the un-transfection group (Control group) and non-sense miRNA sequence transfection group (NC group) were set. The expression of miRNA-141 in DU145 cells in each group before and after transfection was detected by qPCR. MTT assay was used to detect the proliferation viability and sensitivity to cisplatin (DDP) in DU145 cells of each group. Cell cycle and apoptosis rate of DU145 cells under DDP treatment were detected by Flow cytometry; the changes in cell invasion and migration ability were detected by Transwell method. The protein expressions of VEGF and EGFR in DU145 cells of each group were detected by Western blotting. Results: Compared with the Control and NC group, the level of miRNA-141 expression in the miR-141-down group decreased to (0.18±0.08), the cell proliferation viability decreased significantly while its sensitivity to DDP increased significantly, the cell cycle was blocked in the G0+G1 phase, and the apoptosis rate significantly increased to (46.67±5.86)% while cell invasion rate and migration rate significantly decreased to (44.34±8.32)%, (57.73±6.19)%, and the relative expression levels of VEGF and EGFR decreased to (0.47±0.06), (0.36±0.06), (P<0.05 or P<0.01). But in the miR-141-up group, the level of miRNA-141 expression increased to (4.23±0.53), the cell proliferation viability significantly increased while its sensitivity to DDP decreased significantly, and the cell cycle was promoted into S and G2 phase, the apoptosis rate significantly decreased to (18.77±4.24)% while cell invasion and migration rate significantly increased to (89.94±6.34)%, (94.44±5.84)%, and the relative expression levels of VEGF and EGFR were up to (0.89±0.07), (0.73±0.06),(P<0.05 or P<0.01). Conclusion: miR-141 can act as a growth promoting factor in prostate cancer DU145 cells. miR-141 down-regula‐tion can significantly inhibit the proliferation viability, cell cycle, migration and invasion of DU145 cells, and promote cell apoptosis and DDP-sensitivity, and the mechanism of which may be related with inhibition of VEGF and EGFR protein expressions.

Rapid and sensitive detection of Bordetella bronchiseptica by loop-mediated isothermal amplification (LAMP)

Bordetella bronchiseptica causes acute and chronic respiratory infections in diverse animal species and occasionally in humans. In this study, we described the establishment of a simple, sensitive and cost-efficient loop-mediated isothermal amplification (LAMP) assay for the detection of B. bronchiseptica. A set of primers towards a 235 bp region within the flagellum gene of B. bronchiseptica was designed with online software.. The specificity of the LAMP assay was examined by using 6 porcine pathogens and 100 nasal swabs collected from healthy pigs and suspect infected pigs. The results indicated that positive reactions were confirmed for all B. bronchiseptica and no cross-reactivity was observed from other non-B. bronchiseptica. In sensitivity evaluations, the technique successfully detected a serial dilutions of extracted B. bronchiseptica DNA with a detection limit of 9 copies, which was 10 times more sensitive than that of PCR. Compared with conventional PCR, the higher sensitivity of LAMP method and no need for the complex instrumentation make this LAMP assay a promising alternative for the diagnosis of B. bronchiseptica in rural areas and developing countries where there lacks of complex laboratory services.

Development and evaluation of a loop-mediated isothermal amplification (LAMP) assay for rapid detection of Actinobacillus pleuropneumoniae based the dsbE-like gene

This paper reports on the development and validation of a loop-mediated isothermal amplification assay (LAMP) for the rapid and specific detection of Actinobacillus pleuropneumoniae (A. pleuropneumoniae). A set of six primers were designed derived from the dsbE-like gene of A.pleuropneumoniae and validate the assay using 9 A. pleuropneumoniae reference/field strains, 132 clinical isolates and 9 other pathogens. The results indicated that positive reactions were confirmed for all A. pleuropneumoniae strains and specimens by LAMP at 63ºC for 60 min and no cross-reactivity were observed from other non-A.pleuropneumoniae including Haemophilus parasuis, Escherichia coli, Pasteurella multocida, Bordetella bronchiseptica, Streptococcus suis, Salmonella enterica, Staphylococcus, porcine reproductive and respiratory syndrome virus (PRRSV), and Pseudorabies virus. The detection limit of the conventional PCR was 10² CFU per PCR test tube, while that of the LAMP was 5 copies per tube. Therefore, the sensitivity of LAMP was higher than that of PCR. Moreover, the LAMP assay provided a rapid yet simple test of A. pleuropneumoniae suitable for laboratory diagnosis and pen-side detection due to ease of operation and the requirement of only a regular water bath or heat block for the reaction.