Effects Of Griseofulvin On Human Small Cell Lung Cancer Cell Line NCI-H446 In Vitro And In Vivo And Its Possible Mechanism

Objective: To study the anti-tumor effects of griseofulvin on human small cell lung cancer cell line NCI-H446 in vitro and in vivo.Methods:①N CI-H446 cells were treated with different concentrations of griseofulvin for 48h.CCK-8 assay was used to determine the cell survival rate;The changes of morphology and ultrastructure were observed by inverted microscope,fluorescence microscope and transmission electron microscope;Cell cycle and apoptosis rate were analysed by flow cytometry(FCM);AnnexinⅤ-FITC apoptosis detection kit was used to detect the apoptosis of cells;The enzyme activity changes of Caspase-3 and Caspase-9 were detected by spectrophotometry;Western blotting was used to detect the changes of Bax,Bc1-2,and cytochrome C in cytosol of NCI-H446 cells in the protein level.②The migration,invasion and adhesion ability of NCI-H446 cells were detected by scratch test,transwell chamber assay and Transwell-Matrigel model respectively.The cell adhesion assay was carried out on 96-well plate precoated with matrigel.③For in vivo study, we first established the NCI-H446 tumor model by grafting NCI-H446 cells into the athymic nude mice,and then injected griseofulvin into the tumors.Two weeks after injection,we sacrificed the mice and removed the tumors,then weighed and calculated the ratios of tumor-suppression.We also detected the expressions of Bcl-2、Bax、Caspase-3、p53 and VEGF with immunohistochemistry.Results:①CCK-8 assay results showed that griseofulvin could significantly inhibit the proliferation of NCI-H446 cells in vitro in a dose-dependant manner,and the IC50 of griseofulvin was 24.58±1.32μM.Typical morphological changes of NCI-H446 cells were observed after induced by griseofulvin. The cell cycle distribution of NCI-H446 were changed,the number of cell increased in the stage of G2/M and decreased in the stage of S and G0/G1, accompanied with the subdiploid apoptotic peak.After 48 hours treatment with griseofulvin,the rates of apoptosis of griseofulvin-treated NCI-H446 cells greatly increased in high concentration group but did not obviously change in low concentration group compared with control group.Colorimetric assay results showed that griseofulvin could activated caspase-3 and caspase-9;Western blotting results indicated that the expression of cytochrome C and Bax were increased but Bcl-2 was decreased by griseofulvin.②The adhesion,metastatic potential and invasiveness of NCI-H446 cells were obviously inhibited by griseofulvin in a dose-dependent manner.The result of scratch test also indicated that the migration ability of NCI-H446 cells in griseofulvin treated group was decreased compared with control group.③The in vivo data showed that griseofulvin inhibited the tumor growth conspicuously through down-regulating the expression of Bcl-2,VEGF and up-regulating the expression of Bax,caspase-3 and p53.The ratios of tumor weight-suppression of griseofulvin group was 42.07 % .Conclusion: Griseofulvin can inhibit the growth and induce apoptosis of NCI-H446 cells;griseofulvin inhibits the adhesion, invasiveness and migration of NCI-H446 cells in vitro;griseofulvin inhibits the tumorigenicity of NCI-H446 cells in vivo.