The Study Of Diallyl Trisulfide Induced Apoptosis Of Papillary Thyroid Cancer Cells And Molecular Mechanisms

Allicin is the general name of many organic sulfide compounds（OSCs）,which is derived from garlic when the garlic is cut or chewed.All these OSCs have the odiferous nature of garlic.Among the OSCs,diallyl trisulfide（DATS）has many bioactivities in different kinds of cancers.The incidence of thyroid cancer has steadily increased in recent years.Although the prognosis of most thyroid cancers is good,some patients with malignant thyroid cancer is ¹³¹I refractory and resistant to chemotherapy drugs,which eventually leads to death.Therefore,it is needed to investigate that whether DATS could inhibit the progress of thyroid cancer and whether DATS could be used as a therapeutic strategy for thyroid cancer.Hence,the study aimed to investigate the function of DATS on thyroid cancer and its underlying mechanisms.This study will lay the foundations to the application of DATS.This study found that,compared to the effects on BCPAP cells,DATS was less toxic on normal thyroid follicular epithelial cells Nthy-ori-3.1.DATS selectively target thyroid cancer cells rather than thyroid normal cells.Besides,it was found that DATS effectively kill BCPAP cells in a time-and concentration-dependent manner and inhibit cancer cell proliferation and colony formation.The results of Annexin V/PI,Hoechst 33342/PI and JC-1 staining showed that DATS induced apoptosis in BCPAP cells.The results of Western Blot showed that the expressions of anti-apoptotic proteins Bcl-2 and Bcl-X_L were decreased,and the expression of pro-apoptotic protein Bax was increased.Furthermore,Caspase-8 and Caspase-3 were both cleaved.After Caspase-3 was activated,it cleaves its downstream target PARP and finally induced apoptosis in BCPAP cells.However,it was found that DATS-induced apoptosis was not triggered by increasing intracellular ROS levels by DCFH-DA staining assay,but induced by transient overactivation of intracellular MAPK signaling pathway.The results of ELISA showed that H₂S could be released rapidly within 5 min after incubation with different concentrations of DATS.IAM,an inhibitor of H₂S,it can significantly reverse the release of H₂S and increased cell membrane permeability caused by DATS.The quick donor of H₂S,NaHS,and the slow donor of H₂S,GYY4137,can both inhibit cell viability of KTC-1 cells.The above results indicated that DATS was cytotoxic to thyroid cancer cell KTC-1 by releasing H₂S.Western Blot was performed to detect the expression of CTH after DATS treatment and the results found that DATS can upregulate the expression of CTH in KTC-1 cells.IAM,the inhibitor of H₂S,can reverse the upregulation of CTH induced by DATS.This study overexpressed CTH in KTC-1 cells through transfecting CTH-overexpression plasmids.The results were that CTH overexpression reduced thyroid cancer cell activity.All the data indicated that DATS killed KTC-1 cells by releasing H₂S and upregulating the expression of CTH.To investigate the mechanism of DATS-induced CTH upregulation,this study found that DATS upregulated intracellular CTH levels by activating NF-κΒsignaling pathway in KTC-1cells.Using the quick donor of H₂S,NaHS,and the slow donor of H₂S,GYY4137,to treat KTC-1 cells,the results were that high concentrations of both H₂S donors had similar effects with DATS on thyroid cancer cell line KTC-1.PDTC,a kind of NF-κΒinhibitor,reversed DATS-induced NF-κΒactivation and upregulation of CTH protein expression,which indicated that DATS increased the expression of CTH through activating NF-κB signaling pathway.Taken together,all these results further confirmed that H₂S released by DATS did positively upregulate CTH levels through activating NF-κB signaling pathway and consequently induced KTC-1 cell death.In summary,DATS induced apoptosis in papillary thyroid carcinoma cells BCPAP by overactivating MAPK signaling pathway.In addition,DATS can release H₂S,induce CTH expression in KTC-1 cells.The increased CTH,as a positive feedback,further promoted the increase of intracellular H₂S levels and ultimately induced cell death.Therefore,DATS can target CTH in thyroid cancer cells and induce apoptosis of thyroid cancer cells,which is expected to be applied to the treatment of thyroid cancer and further improve the prognosis of thyroid cancer.