Figure 3

Cyr61-promoted EMT is mediated by integrin αvβ5. (A) MG63 cells were pretreated with anti-integrin αvβ3 or αvβ5 mAb (3 μg/mL) for 30 min, followed by stimulation with Cyr61 (10 ng/mL) for 24 h. Cell migration was measured after 24 h using the Transwell assay. The IgG served as the negative control. (B and C) MG63 cells were pretreated with anti-integrin αvβ3 or αvβ5 mAb (3 μg/mL) for 30 min, followed by stimulation with Cyr61 (10 ng/mL) for 24 h. Total mRNA and proteins were extracted from the MG63 cells, and the expression levels of TWIST-1, N-cadherin and E-cadherin were detected using qRT-PCR and western blotting. (D) MG63 cells were transfected with integrin αvβ3, αvβ5 shRNA, or control shRNA for 24 h, followed by treatment with Cyr61 (10 ng/mL) for 24 h, and cell migration was analyzed using the Transwell assay. (E and F) MG63 cells were transfected with integrin αvβ3, αvβ5 shRNA, or control shRNA for 24 h, followed by treatment with Cyr61 (10 ng/mL) for 24 h. Total mRNA and proteins were extracted from the MG63 cells, and expression levels of TWIST-1, N-cadherin and E-cadherin were detected using qRT-PCR and western blotting. (G) MG63 cells were incubated with Cyr61 (10 ng/mL), osteopontin (10 ng/mL) or MFG-E8 (100 ng/mL) for 24 h, and cell migration was measured using the Transwell assay. (H and I) MG63 Cells were incubated with Cyr61, osteopontin or MFG-E8 for 24 h. Total mRNA and proteins were extracted from the MG63 cells, and the expression levels of TWIST-1, N-cadherin, and E-cadherin were detected using qRT-PCR and western blotting. The results are expressed as the mean ± SEM of triplicate samples. *P <0.05 compared with the control group, and ^#P <0.05 compared with Cyr61 treatment.