Effects Of Proliferation And Apoptosis Of Human Osteosarcoma MG63 Cells Induced By Antioxidant Pyrrolidine Dithiocarbamate

ObjectiveOsteosarcoma is one of the most common malignant bone tumor which is originated from primary mesenehymal tissue. It is frequently occur among in adolescent..lt has high malignant grade and poor prognosis. Only 20% of the patient can live five year.For the past few years.With the development of newly adjunctive chemotherapy, but the patient who show increased resistance to chemotherapy and recur soon have poor effect after chemotherapy and operation.Except traditional operation and chemotherapy, considerable advances have been achieved in the fields of gene therapy for osteosarcoma. Inhibitor of apoptosis protein(IAP) family may play a key role in causing apoptosis resistance and has become the focus of research increasingly. Livin is a novel inhibitor of apoptosis protein(IAP)family member and over expresses in most cancers. High expression of Livin can decrease sensitivity of apoptosis.So it might elevate the effete of osteosarcoma therapy to suppress the expression of Livin and increase the sensitivity of osteosarcoma cell apoptosis.It is reported that antioxidant pyrrolidine dithiocarbamate (PDTC) can inhibit the expression of CIAP1 and CIAP2 and cause apoptosis.But the relationship between PDTC and the expression of Livin is not clear..The aim of this study was to detect the effect of PDTC on the survival and apoptosis of human osteosarcoma MG63,and to explore the effect of Livin in the mechanism of PDTC.MethodsAfter the treatment with PDTC on MG63 cell. MTT assay was used to observe growth inhibition of MG63 cells; Flow cytometry(FCM) was used to detect the cell apoptosis; RT-PCR and western blot assay are applied to detect the expression of Livin and Caspase-3 mRNA and protein respectively.ResultsThere was a dose and time-dependent inhibition of cell proliferation by PDTC(0-400μmol/L) at difference points(24h,48h), As shown in FCM, after different concentrates of PDTC for 48h.the apoptotic rates were 2.80±0.35%,6.78±1.07%,14.45±1.56%,20.23±1.02%,28.65±1.87% respectively. there were significant differences at P<0.01 level compared with control groups.;and after100μmol/L PDTC at difference time points,they were 10.04±1.07%,14.45±1.56%respectively,there were significant differences at P＜0.01 level compared with control groups.The mRNA and protein expression of Livin were down-regulated,but those of caspase-3 were up-regulated by PDTC in a dose-dependent manner. Moreover expression of of Livin had a negative correlation correlation with both PDTC and the apoptosis rate of osteosarcoma cell. which showed a dose-dependent manner. It also had a negative correlation with the expression of caspase-3。Conclusions1.PDTC can inhibit proliferation and the cell cycle distribution changed obviously, induced apoptosis of MG63 cells.The growth of cells was inhibited and cells apoptosis in a dose-dependent manner.2.PDTC can down-regulate of Livin and up-regulate of caspase-3, which suggests that the expression of Livin is controlled by PDTC.3.Expression of of Livin had a negative correlation correlation with both PDTC and the apoptosis rate of osteosarcoma cell. which showed a dose-dependent manner.It also had a negative correlation with the expression of caspase-3.It suggests that PDTC can down-regulate of Livin and up-regulate of caspase-3,which suggests that the expression of Livin is controlled by NF-kB activated state.