N = 120 Normal & Benign Borderline Carcinoma Nodal status Positive Negative Tumour stage I II III IV Age(y) <40 ≥40 Cell differentiation High Medium Low Cases 20 20 80 28 52 20 20 20 20 32 48 32 30 18 3 7 44 21 23 9 10 11 14 18 26 14 18 12 17 13 36 7 29 11 10 9 6 14 22 18 12 6 15.00 35.00 55.00 75.00 44.23 45.00 50.00 55.00 70.00 56.25 54.17 43.75 60.00 66.67 Ye et al. BMC Cancer 2010, 10:611 http://www.biomedcentral.com/1471-2407/10/611 Table 1 Expression of Plexin-B1 in ovarian tissues of different pathological changes and its correlation with clinical characteristics Plexin-B1 expression Positive Negative Positive Percentage (%) <0.001* and n.s.** indicated differences of Plexin-B1 expression in normal ovaries and serous ovarian cystadenomas compared with that in serous borderline ovarian tumours and serous ovarian cystadenocarcinomas respectively. n.s.* indicated difference of Plexin-B1 expression in serous borderline ovarian tumours compared with that in serous ovarian cystadenocarcinomas. n.s. indicated differences without statistical significance. Three different siRNA duplexes targeting Plexin-B1 and a negative control siRNA were separately trans- ferred into SKOV3 cells. At 24 h after transfection, Plexin-B1 mRNA and protein levels were assessed by real-time PCR and western blot analysis, respectively. As shown in Figure 2A-C, each of the three Plexin-B1 siRNA duplexes significantly reduced the amount of Plexin-B1 mRNA and protein. The data are from densi- tometric analyses of Plexin-B1: b-actin; n = 5 indepen- dent experiments. Silencing of Plexin-B1 down-regulats phospho-AKT (Ser473) expression in SKOV3 cells Numerous studies showed that the PI3K/AKT pathway is constitutively overexpressed in ovarian cancers [35] and is involved in regulating malignant cellular responses such as growth, survival, migration, and inva- sion [36]. To investigate the relationship between Plexin-B1 expression and PI3K/AKT signaling, we moni- tored the effect of Plexin-B1 silencing on total AKT level and its phosphorylation level in SKOV3 cells. As shown in Figure 2B, C, inhibition of Plexin-B1 expression decreased the levels of phospho-AKT (Ser473) by more than 30%, but it did not alter total AKT expression. Silencing of Plexin-B1 did not affect the proliferation of SKOV3 cells Increased AKT activity is associated with cell growth. To determine whether the Plexin-B1 siRNA-induced reduction of phospho-AKT (Ser473) would affect the proliferation of SKOV3 cells, the MTT assay was used to monitor the proliferation of untransfected SKOV3 cells and SKOV3 cells treated with Plexin-B1 siRNA2, Plexin-B1 siRNA3 or negative control siRNA. As showed in Figure 2C, there were no significant differ- ences between the siRNA-treated groups and the untreated group (P > 0.05). Silencing of Plexin-B1 inhibited migration and invasion in SKOV3 cells Cancer cell migration and invasion are directly related to metastasis. Plexin-B1 has been implicated in the pro- cesses of cell mobility and cancer metastasis [37,38]. To Page 5 of 11 P <0.001* n.s.** n.s.* 0.008 n.s. n.s. n.s.