Effect Of PI3K Inhibitor WORTMANNIN On Invasion And Migration Of Human Thyroid Follicular Carcinoma Cell Line

PART ONE EFFECT OF PI3K INHIBITOR WORTMANNIN ON INVASION AND MIGRATION OF HUMAN THYROID FOLLICULAR CARCINOMA CELL LINEObjective:To study the effect of wortmannin on migration and invasion of human thyroid follicular carcinoma cell line CGTHW -1,and try to investigate the mechanism of action.Methods:We used MTT colorimetric assay selecting the optimal concentration and time of wortmannin .The trial was divided into two groups,the control group and the experimental group (which was cultured in the case of culture medium with or without calf serum ).CGTHW-1 cells were treated with wortmannin, and then the migration and invasion ability of CGTHW-1 cells was assessed by scratch test and Transwell chamber invasing assay respectively.Using methyl rhodanmine labeled by phalloidine staining microfilament through immunofluorescence to observe the change of morphology of CGTHW-1 cells.The expression of Rac1 was examined by immunocytochemistry staining and semiquantity RT-PCR in CGTHW-1 cells.Results:When CGTHW-1 cells were curltured with 10% calf serum medium ,after treated with wortmannin, the CGTHW-1 cells'migrating speed became slow and the numbers passing through the Transwell chamber were significantly reduced (13.8±3.03), compared with the control group (41.6±6.95, P<0.01). The immunocytochemistry staining and semiquantity RT-PCR experiment demonstrated that the expression of protein and mRNA of Rac1 were significantly downregulated in CGTHW-1 cells after wortmannin treatment (P<0.05). However,when the cells were cultured with culture medium without calf serum,the migration and invasion ability of the two groups had no significant difference(P>0.05).Conclusion:wortmannin could depress migration and invasion of human thyroid follicular carcinoma cells effectively, its mechanism may be due to cause cytoskeleton rearrangement through inhibiting the downstream molecule of PI3K, Rac1. PART TWO EXPRESSION OF INTEGRINβ4 IN THYROID FOLLICULAR CANCER CELL SUBLINES WITH HIGH AND LOW POTENTIAL OF METASTASIS AND ITS SIGNIFICANCEObjective: To investigate the effect to cell invasion ability about integrinβ4 in the two subgroups of human thyroid follicular cancer cells with high- and low-metastatic potentiality.Methods: Continuous in vitro invasion assays in artificial matrigel were applied to screen two subgroups of human thyroid follicular cancer cell line CGTHW-1, characterized by high- and low-metastatic potentiality. The expression of integrinβ4 protein and mRNA were examined by immunocytochemistry, western blot and semiquantitative RT-PCR.Results: Immunocytochemistry, western blot and semiquantitative RT-PCR all revealed that the expression of integrinβ4 at protein and mRNA levels were significantly higher in cells with highly-metastatic potentiality[(0.2775±0.0340)vs(0.1875±0.0222),(0.0997±0.0185)vs(0.0390±0.0102),(0.5550±0.1012)vs(0.2700±0.0299),both p<0.01) ].Conclusion: Integrinβ4 plays an important role in human thyroid cancer invasion.