Objective To investigate the effect of the ectopic expression of transmembrane protein PAG1 on the motility of human cutaneous squamous cell carcinoma A431 cells.
Methods The PAG1-EGFP lentiviral vector was constructed. A431 cells were transfected with PAG1-EGFP lentiviral vector (Experimental group) or negative-EGFP lenti-viral vector (Control group), or remained untransfected(Parental group). The transfection efficiency was detected by flow cytometry. To further verify and determine the success of the infection, the expression of PAG1 mRNA and protein were detected by real-time fluorescence quantitative PCR and Western blot, respectively. After additional culture, the wound-healing assay and Transwell chamber assay were performed to evaluate the abilities of migration and invasion.
Results The transfection efficiency in the experimental group and control group were (94.97±0.15)% and (94.60±0.35)%, respectively. After transfected with PAG1-EGFP vector, the expression of PAG1 mRNA was about 1.6 times enhanced than that of untrasfected cells(P=0.000). The PAG1 protein levels were significantly increased after transfection with PAG1-EGFP lentiviral vector (P=0.000). The A431 cell line with PAG1 overexpression was successfully constructed; the ectopic expression of PAG1 significantly decreased the cells healing rate(P=0.000). Moreover, cell migration and invasion abilities were significantly inhibited(P=0.001, P=0.000).
Conclusion The elevated levels of PAG1 could obviously inhibit the mobility of human cutaneous squamous cell carcinoma cells in vitro, which suggesting that the overexpression of PAG1 may influence their metastatic ability in vivo.
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