Cadherin 6 drives epithelial-mesenchymal transition and oxaliplatin resistance in gastric cancer via transforming growth factor-β/Smad signaling axis

Figure 6 Rescue experiments confirming that cadherin 6 mediates phenotypes via the transforming growth factor-β pathway. A: Images from wound healing assays for cadherin 6 (CDH6)-overexpression (OE-CDH6) and OE-CDH6 + transforming growth factor (TGF)-β inhibitor (10 μM) groups at 0, 24, and 48 hours. Right panel: Quantitative analysis showed that TGF-β inhibitor significantly rescued (reduced) the enhanced migration ability induced by CDH6 overexpression (P < 0.001); B: Images from migration and invasion assays showing that TGF-β inhibitor decreased cell migration (P = 0.0134) and invasion (P = 0.0016); C: Images from colony formation assays showed that TGF-β inhibitor suppressed colony formation in OE-CDH6 cells, suggesting that CDH6-mediated proliferation enhancement depends on the TGF-β pathway (P = 0.0113); D: Quantitative real-time polymerase chain reaction showed that TGF-β inhibitor reduced expression of TβRI (P = 0.0168), Smad2 (P = 0.0293), and Smad3 (P = 0.0486) (upregulated by CDH6 overexpression) and restored expression of epithelial markers E-cadherin (P = 0.0179) and zonula occludens-1 (P = 0.0412) while downregulating that of mesenchymal markers vimentin (P = 0.0443) and α-smooth muscle actin (P = 0.0402); E: TGF-β inhibitor treatment reduced the half-maximal inhibitory concentration of oxaliplatin, indicating reversal of oxaliplatin resistance induced by CDH6 overexpression (P = 0.0017); F: Western blotting showed that TGF-β inhibitor decreased TβRI, Smad2 (total Smad2) and phosphorylated Smad2 levels (indicating suppressed pathway activation), increased expression of epithelial markers (E-cadherin and zonula occludens-1), and decreased expression of mesenchymal markers (N-cadherin, vimentin, and α-smooth muscle actin), confirming reversal of the epithelial-mesenchymal transition phenotype and TGF-β pathway inhibition. Glyceraldehyde-3-phosphate dehydrogenase was used as the normalization control. ^aP < 0.05, ^bP < 0.01, and ^cP < 0.001. CDH6: Cadherin 6; OE-CDH6: Cadherin 6-overexpression; TGF: Transforming growth factor; ZO-1: Zonula occludens-1; SMA: Smooth muscle actin; p-Smad2: Phosphorylated Smad2; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; IC₅₀: Half-maximal inhibitory concentration.