Fbf1 regulates mouse oocyte meiosis by influencing Plk1.

Fas binding factor 1 (Fbf1) is one of the distal appendage proteins in the centriole, located at its distal and proximal ends. It influences the duplication and separation of centrosomes, thereby affecting the progression of the cell cycle during mitosis. However, the function of Fbf1 in meiosis has remained unclear. To explore the role of Fbf1 in the in vitro maturation of mouse oocyte, immunofluorescence staining was used to examine the Fbf1 location in the oocyte and their phenotype after protein deletion. Western blot was used to examine the protein abundance. This study showed that mouse oocytes express Fbf1 which locates at the spindle poles and around the microtubules. Through taxol and nocodazole treatment, and microinjection of siRNA, it was demonstrated that Fbf1 had an important role in the spindle assembly and chromosome separation during mouse oocyte meiosis In particular, microinjection of Fbf1-siRNA resulted in severe abnormalities in the spindle and chromosome arrangement, decreased aggregation of microtubules, disrupted the first oocyte meiosis, and the extrusion of the first polar body. Furthermore, in the Fbf1-siRNA group, there was reduced expression of Plk1 and its agglutination at the spindle poles, along with retarded chromosome segregation due to the activation of the spindle assembly checkpoint (SAC) component BubR1. These results indicate that Fbf1 may function in microtubule depolymerization and agglutination, control the microtubule dynamics, spindle assembly and chromosome arrangement and, thus, influence the mouse oocyte meiotic maturation.

Antitumor evaluation and multiple analysis on different extracted fractions of the root of Cynanchum auriculatum Royle ex Wight.

The root of Cynanchum auriculatum (C. auriculatum) Royle ex Wight has been shown to possess various pharmacological effects and has recently attracted much attention with respect to its potential role in antitumor activity. The C-21 steroidal glycosides are commonly accepted as the major active ingredients of C. auriculatum. In this study, the antitumor abilities of different extracted fractions of the root bark and the root tuber of C. auriculatum were investigated by using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay in human cancer cell lines HepG2 and SMMC-7721. The results showed that the chloroform and ethyl acetate fractions of the root tuber suppressed tumor cell growth strongly. To identify and characterize the chemical constituents of different active fractions, an ultra high performance liquid chromatography with triple-quadrupole tandem mass spectrometry method was developed for the simultaneous quantitation of eight C-21 steroidal glycosides. The analysis revealed that the C-21 steroidal glycosides were concentrated in the chloroform and ethyl acetate fractions, and the total contents of different fractions in the root tuber were significantly higher than those of corresponding ones in the root bark. Furthermore, the C-21 steroidal glycosides based on different types of aglucones were prone in different medicinal parts of C. auriculatum.

Diagnostic accuracy of transient elastography (FibroScan) in detection of esophageal varices in patients with cirrhosis: A meta-analysis.

AIM: To investigate the diagnostic accuracy of FibroScan (FS) in detecting esophageal varices (EV) in cirrhotic patients. METHODS: Through a systemic literature search of multiple databases, we reviewed 15 studies using endoscopy as a reference standard, with the data necessary to calculate pooled sensitivity (SEN) and specificity (SPE), positive and negative LR, diagnostic odds ratio (DOR) and area under receiver operating characteristics (AUROC). The quality of the studies was rated by the Quality Assessment of Diagnostic Accuracy studies-2 tool. Clinical utility of FS for EV was evaluated by a Fagan plot. Heterogeneity was explored using meta-regression and subgroup analysis. All statistical analyses were conducted via Stata12.0, MetaDisc1.4 and RevMan5. RESULTS: In 15 studies (n = 2697), FS detected the presence of EV with the summary sensitivities of 84% (95%CI: 81.0%-86.0%), specificities of 62% (95%CI: 58.0%- 66.0%), a positive LR of 2.3 (95%CI: 1.81-2.94), a negative LR of 0.26 (95%CI: 0.19-0.35), a DOR of 9.33 (95%CI: 5.84-14.92) and an AUROC of 0.8262. FS diagnosed the presence of large EV with the pooled SEN of 0.78 (95%CI: 75.0%-81.0%), SPE of 0.76 (95%CI: 73.0%-78.0%), a positive and negative LR of 3.03 (95%CI: 2.38-3.86) and 0.30 (95%CI: 0.23-0.39) respectively, a summary diagnostic OR of 10.69 (95%CI: 6.81-16.78), and an AUROC of 0.8321. A meta-regression and subgroup analysis indicated different etiology could serve as a potential source of heterogeneity in the diagnosis of the presence of EV group. A Deek's funnel plot suggested a low probability for publication bias. CONCLUSION: Using FS to measure liver stiffness cannot provide high accuracy for the size of EV due to the various cutoff and different etiologies. These limitations preclude widespread use in clinical practice at this time; therefore, the results should be interpreted cautiously given its SEN and SPE.

Characterization of antigenic domains and epitopes in the ORF3 protein of a Chinese isolate of avian hepatitis E virus.

Avian hepatitis E virus (HEV) is an emerging virus associated with the big liver and spleen disease or hepatitis-splenomegaly syndrome in chickens and subclinical infections by the virus are also common. The complete genome of avian HEV contains three open-reading frames (ORFs) in which ORF2 protein is part of virus particles and thus contains primary epitopes. Antigenic epitopes of avian HEV ORF2 protein have been described but those associated with the ORF3 have not. To analyze the antigenic domains and epitopes in the ORF3 protein of a Chinese isolate of avian HEV (CaHEV), we generated a series of antigens comprised of the complete ORF3 and also five truncated overlapping ORF3 peptides. The antibodies used in this study were mouse antisera and monoclonal antibodies against ORF3, positive chicken sera from Specific Pathogen Free chickens experimentally infected with CaHEV and clinical chicken sera. Using these antigens and antibodies, we identified three antigenic domains at amino acids (aa) 1-28, 55-74 and 75-88 in which aa 75-88 was a dominant domain. The dominant domain contained at least two major epitopes since field chickens infected with avian HEV produced antibodies against the domain and epitopes. These results provide useful information for future development of immunoassays for the diagnosis of avian HEV infection.

RNAi-induced K-Ras gene silencing suppresses growth of EC9706 cells and enhances chemotherapy sensitivity of esophageal cancer.

To analyze the growth, proliferation, apoptosis, invasiveness and chemotherapy sensitivity of EC9706 cells after K-Ras gene silencing, an expression carrier pSilencer-siK-Ras was constructed, and the EC9706 cell line was transfected using a liposome technique. Six groups were established: Control, siRNA NC (transfected with empty vector pSilencer2.1); Ras siRNA (transfected with pSilencer-siK-Ras2); Paclitaxel; Paclitaxel + siRNA NC; and Ras siRNA +Paclitaxel. After the treatment, RT-PCR, Western blotting, MTT assay, flow cytometry and the Transwell technique were used to assess expression of K-Ras mRNA and protein in EC9706 cells, as well as cell growth, proliferation, apoptosis and invasiveness. The effect of Paclitaxel chemotherapy was also tested. pSilencer-siK-Ras2 effectively down-regulated expression of K-Ras mRNA and protein in EC9706 cells, growth being significantly inhibited. Flow cytometry indicated obvious apoptosis of cells in the experimental group, with arrest in the G1 phase; cell migration ability was also reduced. After pSilencer-siK-Ras2 transfection or the addition of Paclitaxel, EC9706 cells were suppressed to different extents; the suppressive effect was strengthened by combined treatment. The results suggested that RNAi-induced K-Ras gene silencing could enhance chemotherapy sensitivity of esophageal cancer.