As shown in Figure 2, compared with empty vector, cell cycle was arrested at the G1 phase when cells were transfected with pEGFP N1 MT1G. The percentage of G1 phase was increased from selleck chemicals 55. 9% to 62. 1% at 60 h post transfection, and from 59. 1% to 65. 9% at 84 h post transfection in K1 cells, and from 61. 0% to 67. 7% at 48 h post transfection, and from 62. 4% to 68. 0% at 72 h post transfection in FTC133 cells, respectively. In addition, characteristic morphologies of apoptotic nuclei, such as chromatin condensation, margination and nuclear fragmentation, were more frequently observed in cells transfected with pEGFP N1 MT1G compared with empty vector. As shown in Figure 3, the apoptotic cell number increased in MT1G transfected cells compared with empty vector transfected cells, particularly in K1 cells.
MT1G inhibits thyroid cancer cell migration and invasion In the present study, promoter methylation of MT1G was shown to increase the risk of lymph node metastasis in PTC patients. Thus, we next attempted to explore the ef fect of MT1G restoration on the migration and invasion of thyroid cancer cells. As shown in Figure 4A, for K1 cells, there was a significantly lower number of migrated cells in MT1G transfected cells than empty vector transfected cells, indicating that MT1G inhibited cancer cell migration. Furthermore, the Matrigel assays showed that the number of cells that passed through Matrigel coated membrane into the lower chamber was significantly lower in MT1G transfected K1 cells than empty vector transfected K1 cells.
Cell migration and invasion assays were also performed in FTC133 cells using the same protocols. However, we failed to find any migrating or invading cells in both MT1G and empty vector transfected cells. Thus, scratch wound healing assay was performed to evaluate cell migration in FTC133 cells. As shown in Figure 4C, the wound healing was markedly inhibited in MT1G transfected cells as com pared to empty vector transfected cells. These observa tions suggest that MT1G inhibits the invasive potential of thyroid cancer cells. MT1G acts as a tumor suppressor via modulating the activity of PI3K Akt pathway To gain insights into the downstream signaling pathways modulated by MT1G in tumor inhibition, we investi gated the effect of MT1G on the activities of PI3K Akt and MAPK pathways, which play a key role in cell pro liferation and survival in human cancers, including thy roid cancer.
Our data showed that ectopic expression of MT1G inhibited phosphorylation of Akt in both K1 and FTC133 cells. However, we did not find its effect on phosphorylation of Erk1 2. Next, we investigated the effect of MT1G on the expres sion of Mdm2, which can be regulated by the PI3K Akt pathway. As also shown in Figure 5A, we selleck inhibitor indeed observed that MT1G restoration decreased Mdm2 ex pression in thyroid cancer cells.