Suppressive effect of microRNA-449a on the NDRG1/PTEN/AKT axis regulates endometrial cancer growth and metastasis.

Database screening indicated that microRNAs (miRNAs) are involved in pathogenesis of endometrial cancer. Among these miRNAs, miR-449a might be involved in tumorigenesis and lower expression of miR-449a was associated with poor prognosis. However, the role of miR-449a and its underlying molecular mechanism in endometrial cancer (EC) has not been investigated. In this study, our analysis found that miR-449a expression is inversely correlated with the stage of EC. Downregulation of miR-449a was correlated with tumor progression and poor prognosis in the EC patients. Results of functional analyses revealed that overexpression of miR-449a in human EC cells alleviated cell proliferation, invasion and metastasis. Conversely, loss of miR-449a in EC cancer cells facilitated all these cellular activities. Moreover, we identified N-myc downstream-regulated gene 1 (NDRG1) as a direct and functional target gene of miR-449a in EC cells, and the expression of NDRG1 in 87 EC specimens were inversely correlated with that of miR-449a. Additionally, further studies show that the down-regulation of NDRG1 expression inhibited proliferation and metastasis of EC cells through the PTEN/AKT pathway. Therefore, these results suggest that miR-449a suppresses the growth and metastasis of EC by directly targeting the NDRG1 gene and that the activation of miR-449a may represent an effective therapeutic strategy in EC.

Fn14 overcomes cisplatin resistance of high-grade serous ovarian cancer by promoting Mdm2-mediated p53-R248Q ubiquitination and degradation.

BACKGROUND: High-grade serous ovarian cancer (HGSOC) is the most lethal of all gynecological malignancies. Patients often suffer from chemoresistance. Several studies have reported that Fn14 could regulate migration, invasion, and angiogenesis in cancer cells. However, its functional role in chemoresistance of HGSOC is still unknown. METHODS: The expression of Fn14 in tissue specimens was detected by IHC. CCK-8 assay was performed to determine changes in cell viability. Apoptosis was measured by flow cytometry. The potential molecular mechanism of Fn14-regulated cisplatin resistance in HGSOC was investigated using qRT-PCR, western blotting, and Co-IP assays. The role of Fn14 in HGSOC was also investigated in a xenograft mouse model. RESULTS: In this study, we found that Fn14 was significantly downregulated in patients with cisplatin resistance. Patients with low Fn14 expression were associated with shorter progression-free survival and overall survival. Overexpression of Fn14 suppressed cisplatin resistance in OVCAR-3 cells, whereas knockdown of Fn14 did not affect cisplatin resistance in SKOV-3 cells. Interestingly, Fn14 could exert anti-chemoresistance effect only in OVCAR-3 cells harboring a p53-R248Q mutation, but not in SKOV-3 cells with a p53-null mutation. We isolated and identified primary cells from two patients with the p53-R248Q mutation from HGSOC patients and the anti-chemoresistance effect of Fn14 was observed in both primary cells. Mechanistic studies demonstrated that overexpression of Fn14 could reduce the formation of a Mdm2-p53-R248Q-Hsp90 complex by downregulating Hsp90 expression, indicating that degradation of p53-R248Q was accelerated via Mdm2-mediated ubiquitin-proteasomal pathway. CONCLUSION: Our findings demonstrate for the first time that Fn14 overcomes cisplatin resistance through modulation of the degradation of p53-R248Q and restoration of Fn14 expression might be a novel strategy for the treatment of HGSOC.

[Regulation of microRNA-199a on adhesion, migration and invasion ability of human endometrial stromal cells].

OBJECTIVE: To study the regulation of microRNA 199a (miR-199a) on adhesion, migration and invasion ability of human eutopic endometrial stromal cells (ESC) from patients with endometriosis. METHODS: ESC were transfected with miR-199a mimics or negative control (NC) RNA by lipofectamine 2000. The adhesion, migration and invasion ability of ESC were detected by cell adhesion assay, scratch assay, cell migration assay and matrigel invasion assay, respectively. Luciferase reporter assay was used to evaluate whether IKKß was the target gene of miR-199a. The expression of ikappa B kinase beta (IKKß), inhibitory kappa B alpha (IκB-α), phospho-IκB-α(p-IκB-α) and nuclear factor-kappa B (NF-κB) protein were measured by western blot. RESULTS: (1) Adhesion potential: the adhesion inhibitory rates were (14 ± 4)% in miR-199a group and 0 in control group, which showed significant difference (P < 0.01). (2) Migration and invasion: in the scratch assay, ESC transfected with miR-199a exhibited a lower scratch closure rate than that of controls. In migration and invasion assays, the migration and invasion ability of miR-199a group were significantly decreased compared with those of NC group [130 ± 31 vs. 247 ± 36 (P < 0.01); 63 ± 15 vs. 133 ± 17 (P < 0.01), respectively]. (3) The luciferase activity of miR-199a group was significantly lowered than that of control group [0.160 ± 0.006 vs. 0.383 ± 0.083 (P < 0.01)]. The protein levels of IKKß, p-IκB-α, IκB-α and NF-κB of 0.350 ± 0.195, 0.443 ± 0.076, 1.970 ± 0.486 and 0.454 ± 0.147 in miR-199a group were significantly different compared with the NC group in which the protein levels were set at 1.000 (P < 0.01). CONCLUSIONS: miR-199a can inhibit the adhesion, migration and invasion of the ESC. IKKß is the target gene of miR-199a in ESC. One of the mechanisms of the inhibition effect is probably that miR-199a inhibits the activation of NF-κB signaling pathway by targeting IKKß gene.

A study of the efficacy of bacterial biofilm cleanout for gastrointestinal endoscopes.

AIM: To compare the influence and clearance effect of enzymatic and non-enzymatic detergents against Escherichia coli (E. coli) biofilm on the inner surface of gastroscopes. METHODS: Teflon tubes were incubated in a mixture of different detergents and E. coli culture (10(6) CFU/mL) for 72 h at 15 degrees C, and biofilms on the inner surface of the teflon tubes were analyzed by bacterial count and scanning electron microscopy. To evaluate the clearance effect of detergents, after biofilms were formed on the inner surface of Teflon tubes by 72 h lavage with E. coli culture, tubes were lavaged by enzymatic and non-enzymatic detergents at a speed of 250 mL/min, then biofilms on the inner surface were analyzed by bacterial count and scanning electron microscopy. RESULTS: Non-enzymatic detergent had a better inhibition function on biofilm formation than enzymatic detergent as it reduced bacterial burden by 2.4 log compared with the control samples (P = 0.00). Inhibition function of enzymatic detergent was not significantly different to that of control samples and reduced bacterial burden by 0.2 log on average (P > 0.05). After lavaging at 250 mL/min for 3 min, no living bacteria were left in the tubes. Scanning electron microscopy observation showed biofilms became very loose by the high shear force effect. CONCLUSION: Non-enzymatic detergent has a better inhibition effect on biofilm formation at room temperature. High speed pre-lavage and detergents are very important in temporal formed biofilm elimination.