Effects Of Hypoxia And Metformin On Biological Behaviors In Salivary Adenoid Cystic Carcinoma Cells

Objectives:To investigate the effects of hypoxia and metformin on the biological behavior of salivary adenoid cystic carcinoma(SACC)cells and its mechanisms,providing theoretical and experimental basis for the prevention and treatment of cancer.Methods:1.When SACC-83 cells were cultured under normal oxygen(21% O2)and hypoxia(15%,10%,5% and 1% O2)for 8,24 and 48 h,the proliferation was detected by CCK methods.When SACC-83 cells were treated with 0,5,10,20 and 40 m M of metfromin under normal oxygen and hypoxia(5% O2)for 48 h,the proliferation was examined using CCK methods.2.When SACC-83 cells were treated with 0,20 and 40 m M of metfromin under normal oxygen and hypoxia(5% O2)for 48 h,the migration and invasion were examined using wound healing and transwell assays,respectively.3.When SACC-83 cells were treated with 0,20 and 40 m M of metfromin under normal oxygen and hypoxia(5% O2)for 48 h,the expression of protein and m RNA were evaluated by western blot and real-time PCR assays,respectively.Results:1.The proliferation of SACC-83 cells was decreased with the decrease of oxygen concentration,with the prolongation of time,and with the increase of metformin concentration.The proliferation of cells under normal oxygen was significantly higher than that under hypoxia(5% O2)in the groups treated with 20 and 40 m M of metformin(P<0.05),suggesting that the inhibitory effect of metformin on the proliferation of cells became decrease under hypoxic environment.2.Both migration and invasion of SACC-83 cells under hypoxia were reduced compared with those under normal oxygen.And the migration and invasion of cells were decreased with the increase of metformin concentration under the same oxygen concentration(P<0.01,P<0.05,vs 0 m M).In addition,the migration and invasion of cells only treated with 20 m M of metformin were inhibited under normal oxygen while the migration and invasion of cells just treated with 40 m M of metformin were inhibited under hypoxic environment(5% O2)(P<0.05,vs 0 m M),suggesting that the inhibitory effect of metformin on the migration and invasion of cells reduced under hypoxic environment.3.The protein and m RNA expression of HIF-1?,VEGF,MMP-2 and MMP-9 in SACC-83 cells were all decreased with the increase of metformin concentration(P<0.01,vs 0 m M),but the protein and m RNA expression of these genes under hypoxia were significantly higher than those under normal oxygen(P<0.05).The data showed that hypoxia could up-regulate the expression of HIF-1?,VEGF,MMP-2 and MMP-9 at the levels of protein and m RNA.Conclusion:Hypoxia could antagonize the inhibitory effects of metformin on the proliferation and enhance the invasion in SACC-83 cells.And the underlying mechanisms might depend on regulation the expression of HIF-1?,VEGF,MMP-2 and MMP-9.