The Regulation Effects Of CAMP/PKA Singnal Pathway And TSA On Proliferation Of Thyroid Cancer

ObjectiveTo investigate the regulation effects of cAMP/PKA singnal pathway and TSAon proliferation of thyroid cancer cell line SW579.MethodsSW579cells were treated using dbcAMP(0.5,1.0, and2.0mmol/l), after24h,48h, and72h, PKA activity and proliferation of the treated cells wereinvestigated by ELISA and MTT methods respectively; SW579cells weretreated by1.0mmol/L dbcAMP,50nmol/L TSA, or1.0mmol/LdbcAMP and50nmol/L TSA，after48h,PKA and MPF activities of the treated cells weredetected using ELISA;the mRNAs of CyclinD1,CDK4,and Rb by RT-polymerasechain reaction(PCR); the proteins of PKA,Cyclin1, CDK4and the phosporylatedRB, CDC25B-pS323, and cdc2pTyr15by Western blot.ResultsThe PKA activity of dbcAMP-treated SW579cells increases in aconcentration-and time-dependent manner. Activation of cAMP/PKA pathwayby dbcAMP arrest SW579cells at G1phase, reduces cells at S phase whileG2-phase cells remain unchanged or reduced. Molecularly, Activation ofcAMP/PKA pathway by dbcAMP inhibits MPF activity, downregulates CyclinD1mRNA and protein, reduces phosphorylated Rb protein, and increasephosphorylated CDC25B and cdc2, while we have not deceted any effect on CDK4after Activation of cAMP/PKA pathway. Similar to Activation ofcAMP/PKA pathway, TSA also arrest SW579cells at G1phase, reduces cells atS phase while G2-phase cells remain unchanged or reduced. Molecularly, TSAinhibits MPF activity downregulates CyclinD1mRNA and protein, reducesphosphorylated Rb protein, and increase phosphorylated CDC25B and cdc2,while we have not deceted any effect on CDK4after treating SW579using TSA.Interestingly TSA activates PKA and upregulates PKA protein suggesting TSAinhibits SW579cell growth through activation of PKA pathway. SW579cellstreated using TSA and dbcAMP are more strongly arrested at G1phase thanone treated only using dbcAMP or TSA, and cells at S and G2are morestrongly reduced than one treated only using dbcAMP or TSA demonstrating astrong synergistic effect. Molecularly, TSA and dbcAMP also demonstrating thesynergistic effect.Conclusions1cAMP/PKA signaling pathways show a negative regulation function to cellgrowth proliferation of thyroid cancer;2TSA activates cAMP/PKA pathways by upregulating the PKA protein whichinhibits activities of CDC25B and MPF, downregulate the expression ofCyclinD1and phosphorylated Rb, so inhibit the proliferation of the thyroid scalecancer cell line SW579;3TSA and dbcAMP activate PKA by different ways, and demonstrate asynergistic effect.