Figure 3.

PTEN knockdown attenuates epithelial mesenchymal transition (EMT) and morphological changes in vitro. (A) Schematic representation of the experimental procedure. BEAS-2B cells were transfected with PTEN siRNA (Si-PTEN), followed by HCl stimulation and mechanical stretch using the Flexcell FX500T cell strain loading system. (B) Western blot analysis of PTEN protein expression in BEAS-2B cells transfected with Si-PTEN. GAPDH served as a loading control. (C) Quantification of PTEN expression levels normalized to GAPDH. Data are presented as mean ± SEM. n = 4 /group. (D) Relative mRNA expression of PTEN in BEAS-2B cells transfected with Si-PTEN. Data are presented as mean ± SEM. n = 4 /group. (E) Representative phase-contrast images showing cell morphology in wild-type (WT) and Si-PTEN transfected cells under HCl + Stretch condition. (F) Western blot analysis of E-cadherin, Vimentin, and α-SMA protein expression in WT and Si-PTEN transfected cells under HCl + Stretch condition. (G-I) Quantification of E-cadherin, Vimentin, α-SMA expression levels normalized to GAPDH. Data are presented as mean ± SEM. n = 4 /group. (J) Immunofluorescence staining for E-cadherin (green), Vimentin (red), and α-SMA (red) in WT and Si-PTEN transfected cells under HCl + Stretch condition. Nuclei were counterstained with DAPI (blue). (K) Sequence alignment of human and mouse PTEN showing 99.75% sequence homology (NCBI alignment score: 854; E-value ≈ 0). The table below the alignment score indicates the percentage identity, and the distribution graph shows the alignment scores of the top 1 Blast Hits on 1 subject sequence. Comparisons between two groups were performed using independent sample t tests. **: P < 0.01, ***: P < 0.001, ****: P < 0.0001.