Arsenic Trioxide Inhibits The Expression Of Glucocorticoid Receptor In Human Ovarian Cancer And Its Mechanism And Biological Significance

BackgroundArsenic trioxide?As2O3,ATO?is the main component of traditional Chinese medicine arsenic,it has been used as a drug for thousands of years.A large number of in vivo and in vitro experiments have shown that ATO have a strong anti-tumor effect in the treatment of hematological malignancies(such as APL^[1,2],multiple myeloma^[3,4],etc.).It has shown that in a variety of malignant solid tumors such as liver cancer,esophageal cancer,Ovarian cancer,prostate cancer,cervical cancer,etc.ATO also have a strong anti-cancer effect^[5-10].At present,ATO anti-tumor mechanism is more complex,involving the induction of tumor cell apoptosis,induced tumor cell differentiation,inhibition of tumor cell proliferation,inhibition of tumor metastasis and inhibition of angiogenesis and other aspects.Although considerable progress has been made in the study of ATO's anti-tumor mechanism,it has not yet been fully elucidated.In particularly,what kind of molecule plays an important role in ATO's anti-tumor effect still needs to be maintained.Glucocorticoid receptor?GR?is a member of the nuclear receptor superfamily.GR is expressed in almost all tissue cells.The biological effects of glucocorticoid mediated involve in the proliferation,differentiation,apoptosis and immunoregulation of cells,which are closely related to the occurrence and development of tumor.Ovarian cancer is an estrogen-dependent tumor,estrogen receptor through the estrogen receptor?estrogen receptor?mediated signal transduction pathway in the development and progression of ovarian cancer play an important role.However,an interesting phenomenon is that in ovarian cancer tissue specimens,GR has a high abundance of expression,even in the absence of estrogen receptor?ER?and progesterone receptor?PR?expression of ovarian cancer cells still GR expression,suggesting that GR may be involved in the development of ovarian cancer regulation.In the previous study,we found that ATO could induce the apoptosis of human ovarian cancer SKOV3 cells,and could significantly inhibit the expression of GR,which was much higher than that of ER.Based on the earlier findings,we discuss the mechanism of ATO inhibition of GR expression,and further elucidate the effect of GR on the effect of ATO on the proliferation,migration and invasion of ovarian cancer cells.MethodsOur research were made in SKOV3 and HO-8910 cells of ovarian cancer,HepG2 of hepatocellular carcinoma,A549 cells of lung adenocarcinoma in vitro,cells were treated with ATO of different concentration of 0-40?M.The expression of GR mRNA and protein level were dected in these cells.The cell proliferation ability was detected by cell counting and MTT assay.Flow cytometry was used to detect the level of apoptosis and the percentage of cell cycle.The expression of protein level was dected by Western Blot.The Transwell and ibidi cell scratch assay were used to detect the horizontal and vertical migration ability.The Matrigel glue precoated Transwell chamber?Transwell?was to detect cell invasiveness.Results?1?ATO inhibits the expression of GR mRNA and protein in a variety of tumor cells.1.ATO can significantly inhibit the expression of GR in ovarian cancer SKOV3 cells,and this inhibitory effect is dose-dependent and time-dependent.2.ATO inhibits the expression of GR mRNA and protein in human ovarian cancer HO-8910 cells,human hepatocellular carcinoma HepG2 cells and human lung adenocarcinoma A549 cells.ATO inhibits the expression of GR mRNA and protein in human tumor cells.?2?The mechanism of ATO inhibiting the expression of GR mRNA and protein in human ovarian cancer cells.1.ATO can inhibit the expression of GR mRNA in ovarian cancer by methylation of DNA and promote the degradation of GR mRNA.2.ATO can enhance the degradation of GR protein by ubiquitin proteasome pathway.3.ATO inhibits GR protein and P38 activation and activation of reactive oxygen species,but has no inference on JNK activation.?3?The down-regulation of GR expression promoted the inhibitory effect of ATO on the proliferation,migration and invasion in ovarian cancer cells.1.ATO can significantly inhibit the proliferation of ovarian cancer cells and block the cell cycle in G2/M phase.Overexpression of GR reverses this inhibitory effect.The mechanism may be related to the expression of cell cycle related proteins?cyclin D1,cyclin E1,cdc27?.2.ATO can significantly inhibit the migration and invasion of ovarian cancer cells,,while overexpression of GR can reverse this effect.ConclusionIn this study,we can confirm that ATO can down-regulate the expression of GR in a time-dependent and dose-dependent manner,and this inhibition has no obvious cell specificity.ATO promoteing DNA methylation and the degradation of GR mRNA result in ATO's inhibition of the expression of GR mRNA,and its inhibitory effect on GR protein expression may be related to its enhancing degradation by ubiquitin proteasome pathway.By the way,P38 and reactive oxygen species are involved in the down-regulation of GR protein induced by ATO.By inhibiting the expression of GR,ATO could inhibit the proliferation related proteins like cyclin D1,cyclin E1,cdc27 and blocked the cell cycle in G2/M phase,all that result in ATO's inhibitory the proliferation of ovarian cancer cells.ATO could also Significantly inhibite cell migration and invasion process in ovarian cancers by its inhibitory of GR.