The Effect Of Id1/Id3 On Invasion And Metastasis Of Colorectal Cancer Cells By Regulating EMT

Objective To study Id1/Id3 expression in different colorectal cancer cells and the effect of Id1/Id3 on cell proliferation, explore the impact of Id1/Id3 on invasion and metastasis of colorectal cancer cells by regulating EMT.Methods The SW620 cell line with Idl/Id3 gene knockdown and the SW480 cell line with over-expression of Idl/Id3 gene were got by using the lentiviral vectors.Idl/Id3 expression was detected by real-time q PCR and Western blot.Cell proliferation was analysed with RTCA, colony formation assay and flow cytometry. Apoptosis was detected with Annexin V-PE/7-AAD by FACS. Cell morphological change was observed under a microscope. The ability of migration and invasion of cells was determined by wound healing assay and transwell migration/invasion assay. The proteins for cell cycle, EMT, PI3K/AKT pathway,invasion and metastasis were measured by western blot assay.Results 1. It was found that Idl/Id3 gene were highly expressed in SW620 cells and lowly expressed in SW480 cells. The SW620 cell lines with stable silence of Idl/Id3 and the SW480 cell lines with stable over-expression of Idl/Id3 were successfully established. 2. Idl/Id3 gene knockdown inhibited proliferation and clonogenicity of colorectal cancer cells, and induced a cell cycle arrest in G0/G1 phase. Idl/Id3 gene over-expression stimulated the proliferative capacity and clonogenicity of colorectal cancer cells, and drove the cell from G0/G1 phase into S phase. But there was no noticeable effect of Idl/Id3 gene on cell apoptosis. 3. Idl/Id3 knockdown synergistically induced MET in the SW620 cells. Conversely, Idl/Id3 over-expression synergistically induced EMT in the SW480 cells. The scratch wound assay and transwell migration/invasion assay showed that the capacity of invasion and metastasis of the cells significantly decreased when Idl/Id3 was knocked down in the SW620 cell. But the capacity of invasion and metastasis of the cells significantly increased when Idl/Id3 overexpressed in the SW480 cell. Idl/Id3 gene knockdown in SW620 cell down-regulated the protein expression of ?-catenin, snail, slug, twist, p-PI3 K,p-AKT and MMP2, and up-regulated the protein expression of E-cadherin, TIMP1 and TIMP2. However, Idl/Id3 gene over-expression down-regulated the protein expression of E-cadherin, TIMP1 and TIMP2 in the SW480 cell, and up-regulated the protein expression of ?-catenin, snail, slug, twist, p-PI3 K, p-AKT and MMP2 in the SW480 cell.Conclusion Id1 and Id3 gene had co-effects on stimulating the proliferation,invasion and migration of colorectal cancer cells. They could drive the cells from G0/G1 phase into S phase. Idl/Id3 gene over-expression could enhance invasion and metastasis ability of colorectal cancer cells by inducing EMT. Idl/Id3 gene silencing could reverse EMT, and than lead to MET. The capacity of invasion and metastasis of the cells significantly decreased. So it was speculated that Idl/Id3 induced EMT through the phosphorylation of PI3K/AKT in colorectal cancer cells,leading to the up-regulation of invasion and metastasis of colorectal cancer cells.