Effects of tiam 1 on invasive capacity of gastric cancer cells in vitro and underlying mechanisms

Abstract

Objective: To investigate changes in the invasive capacity of gastric cancer cells in vitro after expression inhibition of T lymphomainvasion and metastasis inducing factor 1 (Tiam 1) and underlying mechanisms. Methods: Using adhesion selection, two subpopulations with high (MH) or low (ML) invasive capacity were separated from the human gastric cancer cell line MKN-45 (M0). Tiam 1 antisense oligodeoxynucleotide (ASODN) was transfected into MH cells with liposomes, and expression of Tiam 1 mRNA and protein was determined by RT-PCR and quantitative cellular-ELISA. Changes in the cytoskeleton, invasive capacityin vitro and expression of ras-related C3 botulinum toxin substrate 1(Rac 1), integrin β1 and matrix metalloproteinase 2 (MMP 2) between Tiam 1 ASODN transfected MH cells and non-transfected cells were observed by HE staining, cytoskeletal protein staining, scanning electron microscopy, Boyden chamber tests and cyto-immunohistochemistry. Results: A positive correlation existed between the expression level of Tiam l mRNA or protein and the invasion capacity of gastric cancer cells. After ASODN treatment (0.43 μM for 48 h), Tiam 1 mRNA transcription and protein expression in MH cells were decreased by 80% and 24% respectively (P < 0.05), compared with untreated controls, while invasive capacity in vitro was suppressed by 60% (P < 0.05). Morphologic and ultrastructural observation also showed that ASODN-treated MH cellsexhibited smooth surfaces with obviously reduced filopodia and microspikes, which resembled M0 and ML cells. Additionally, cytoskeletal distribution dramatically altered from disorder to regularity with reduced long filament-like structure, projections, pseudopodia on cell surface, and with decreased acitn-bodies in cytoplasm. After Tiam 1 ASODN treatment, the expression of Rac 1 and Integrin β1 in MH cells wasnot affected (P > 0.05), but that of MMP 2 in MH cells was significantly inhibited compared with untreated cells (P < 0.05). Conclusion: Over-expression of Tiam-1 contributes to the invasive phenotype of gastric cancer cells. Inhibition of Tiam 1 expression could impair the invasive capacity of gastric cancer cells through modulating reconstruction of the cytoskeleton and regulating expression of MMP 2.