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Plexin-B1 silencing inhibits ovarian cancer cell migration and invasion.

Ye S, Hao X, Zhou T, Wu M, Wei J, Wang Y, Zhou L, Jiang X, Ji L, Chen Y, You L, Zhang Y, Xu G, Zhou J, Ma D, Wang S - BMC Cancer (2010)

Bottom Line: Elevated Plexin-B1 expression has been found in diverse human cancers and in non-neoplastic tissues, and it mediates diverse biological and pathological activities.Expression levels of Plexin-B1 protein were significantly higher in serous ovarian carcinomas than in normal ovaries or benign ovarian neoplasms, and in the former, Plexin-B1 expression was positively correlated with lymphatic metastasis, and the membrane and cytoplasm of cancer cells stained positively.Plexin-B1 siRNA significantly suppressed phosphorylation of AKT at Ser473 in SKOV3 cells, but it did not alter total AKT expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China.

ABSTRACT

Background: Elevated Plexin-B1 expression has been found in diverse human cancers and in non-neoplastic tissues, and it mediates diverse biological and pathological activities. However, whether or not Plexin-B1 expression is involved in human ovarian tumors remains unclear. In the present study, Plexin-B1 expression was explored in benign and malignant human ovarian tumor tissues. In addition, the impact of Plexin-B1 expression on ovarian cancer cell proliferation, migration and invasion were investigated in vitro.

Methods: Plexin-B1 expression was analyzed in normal and benign ovarian tissues and serous ovarian tumors (both borderline and malignant) by immunohistochemical staining, as well as in four human ovarian cancer cell lines (A2780, C13*, SKOV3, and OV2008) by RT-PCR and western blot analyses. Furthermore, endogenous Plexin-B1 expression was suppressed by Plexin-B1 siRNA in SKOV3 cells, which overexpress Plexin-B1. Protein levels of Plexin-B1, AKT and AKTSer473 were examined by western blot analysis. Cell proliferation, migration and invasion were measured with MTT, wound healing and boyden chamber assays, respectively, and the cytoskeleton was monitored via F-actin staining.

Results: Expression levels of Plexin-B1 protein were significantly higher in serous ovarian carcinomas than in normal ovaries or benign ovarian neoplasms, and in the former, Plexin-B1 expression was positively correlated with lymphatic metastasis, and the membrane and cytoplasm of cancer cells stained positively. SKOV3 cells displayed the highest Plexin-B1 expression at both the mRNA and protein levels among the four tested human ovarian cancer cell lines and was selected as a cell model for further in vitro experiments. Plexin-B1 siRNA significantly suppressed phosphorylation of AKT at Ser473 in SKOV3 cells, but it did not alter total AKT expression. In addition, silencing of Plexin-B1 in SKOV3 cells inhibited cell migration and invasion and reorganized the cytoskeleton, whereas cell proliferation was not affected.

Conclusion: Plexin-B1 expression correlates with malignant phenotypes of serous ovarian tumors, probably via phosphorylation of AKT at Ser473, suggesting that Plexin-B1 might be a useful biomarker and/or a novel therapeutic target.

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Effect of Plexin-B1 inhibition on cytoskeleton rearrangement. Representative cytoskeletal rearrangements (phalloidin-fluorescein isothiocyanate staining) of untransfected (Blank control) SKOV3 cells and SKOV3 cells at 24 h after a 5-h exposure to Plexin-B1 siRNA2, Plexin-B1 siRNA3 or negative control siRNA. Repression of Plexin-B1 by Plexin-B1 siRNA2 and siRNA3 inhibited F-actin polymerization and filopodia formation in SKOV3 cells. Red indicates F-actin.
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Figure 4: Effect of Plexin-B1 inhibition on cytoskeleton rearrangement. Representative cytoskeletal rearrangements (phalloidin-fluorescein isothiocyanate staining) of untransfected (Blank control) SKOV3 cells and SKOV3 cells at 24 h after a 5-h exposure to Plexin-B1 siRNA2, Plexin-B1 siRNA3 or negative control siRNA. Repression of Plexin-B1 by Plexin-B1 siRNA2 and siRNA3 inhibited F-actin polymerization and filopodia formation in SKOV3 cells. Red indicates F-actin.

Mentions: A previous study proved that Plexin-B1 regulates the actin cytoskeleton by activating Rho activity [22]. To investigate how changes in Plexin-B1 expression affect the reorganization of the cytoskeleton, rhodamine-phalloidin was used to label F-actin in SKOV3 cells 24 h after a 5-h exposure to Plexin-B1 siRNA2 or siRNA3. As shown in Figure 4, SKOV3 cells in the transfected groups shrank without F-actin polymerization or filopodia formation, whereas cells in the blank and control groups showed pointed filopodia or polygonal cell shapes with pointed edges, in which brightly stained longitudinal actin bundles were detected [39].


Plexin-B1 silencing inhibits ovarian cancer cell migration and invasion.

Ye S, Hao X, Zhou T, Wu M, Wei J, Wang Y, Zhou L, Jiang X, Ji L, Chen Y, You L, Zhang Y, Xu G, Zhou J, Ma D, Wang S - BMC Cancer (2010)

Effect of Plexin-B1 inhibition on cytoskeleton rearrangement. Representative cytoskeletal rearrangements (phalloidin-fluorescein isothiocyanate staining) of untransfected (Blank control) SKOV3 cells and SKOV3 cells at 24 h after a 5-h exposure to Plexin-B1 siRNA2, Plexin-B1 siRNA3 or negative control siRNA. Repression of Plexin-B1 by Plexin-B1 siRNA2 and siRNA3 inhibited F-actin polymerization and filopodia formation in SKOV3 cells. Red indicates F-actin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2991310&req=5

Figure 4: Effect of Plexin-B1 inhibition on cytoskeleton rearrangement. Representative cytoskeletal rearrangements (phalloidin-fluorescein isothiocyanate staining) of untransfected (Blank control) SKOV3 cells and SKOV3 cells at 24 h after a 5-h exposure to Plexin-B1 siRNA2, Plexin-B1 siRNA3 or negative control siRNA. Repression of Plexin-B1 by Plexin-B1 siRNA2 and siRNA3 inhibited F-actin polymerization and filopodia formation in SKOV3 cells. Red indicates F-actin.
Mentions: A previous study proved that Plexin-B1 regulates the actin cytoskeleton by activating Rho activity [22]. To investigate how changes in Plexin-B1 expression affect the reorganization of the cytoskeleton, rhodamine-phalloidin was used to label F-actin in SKOV3 cells 24 h after a 5-h exposure to Plexin-B1 siRNA2 or siRNA3. As shown in Figure 4, SKOV3 cells in the transfected groups shrank without F-actin polymerization or filopodia formation, whereas cells in the blank and control groups showed pointed filopodia or polygonal cell shapes with pointed edges, in which brightly stained longitudinal actin bundles were detected [39].

Bottom Line: Elevated Plexin-B1 expression has been found in diverse human cancers and in non-neoplastic tissues, and it mediates diverse biological and pathological activities.Expression levels of Plexin-B1 protein were significantly higher in serous ovarian carcinomas than in normal ovaries or benign ovarian neoplasms, and in the former, Plexin-B1 expression was positively correlated with lymphatic metastasis, and the membrane and cytoplasm of cancer cells stained positively.Plexin-B1 siRNA significantly suppressed phosphorylation of AKT at Ser473 in SKOV3 cells, but it did not alter total AKT expression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China.

ABSTRACT

Background: Elevated Plexin-B1 expression has been found in diverse human cancers and in non-neoplastic tissues, and it mediates diverse biological and pathological activities. However, whether or not Plexin-B1 expression is involved in human ovarian tumors remains unclear. In the present study, Plexin-B1 expression was explored in benign and malignant human ovarian tumor tissues. In addition, the impact of Plexin-B1 expression on ovarian cancer cell proliferation, migration and invasion were investigated in vitro.

Methods: Plexin-B1 expression was analyzed in normal and benign ovarian tissues and serous ovarian tumors (both borderline and malignant) by immunohistochemical staining, as well as in four human ovarian cancer cell lines (A2780, C13*, SKOV3, and OV2008) by RT-PCR and western blot analyses. Furthermore, endogenous Plexin-B1 expression was suppressed by Plexin-B1 siRNA in SKOV3 cells, which overexpress Plexin-B1. Protein levels of Plexin-B1, AKT and AKTSer473 were examined by western blot analysis. Cell proliferation, migration and invasion were measured with MTT, wound healing and boyden chamber assays, respectively, and the cytoskeleton was monitored via F-actin staining.

Results: Expression levels of Plexin-B1 protein were significantly higher in serous ovarian carcinomas than in normal ovaries or benign ovarian neoplasms, and in the former, Plexin-B1 expression was positively correlated with lymphatic metastasis, and the membrane and cytoplasm of cancer cells stained positively. SKOV3 cells displayed the highest Plexin-B1 expression at both the mRNA and protein levels among the four tested human ovarian cancer cell lines and was selected as a cell model for further in vitro experiments. Plexin-B1 siRNA significantly suppressed phosphorylation of AKT at Ser473 in SKOV3 cells, but it did not alter total AKT expression. In addition, silencing of Plexin-B1 in SKOV3 cells inhibited cell migration and invasion and reorganized the cytoskeleton, whereas cell proliferation was not affected.

Conclusion: Plexin-B1 expression correlates with malignant phenotypes of serous ovarian tumors, probably via phosphorylation of AKT at Ser473, suggesting that Plexin-B1 might be a useful biomarker and/or a novel therapeutic target.

Show MeSH
Related in: MedlinePlus