Study On The Relationships Between PTEN And Peptide Growth Factor Insulin And Cu/Zn SOD

Object: To research the role and the interrelationship of peptide growth factor insulin, H2O2 and PTEN in the regulation to Akt kinase ; To discuss whether the deletion of the PTEN could affect the expression of Cu/Zn SOD and the significance of it; To discuss the relationships between the expression of PTEN and peptide growth factors and Cu/Zn SOD and to discuss the contribution of them to the mechanism of the tumorous formatoin and development, so that we can know the tumor suppressor PTEN more clearly and assist the clinical targeted therapy of tumors more effectively.Method: The control immortalized wild-type mouse embryonic fibroblast cells (PTEN+/+MEFs) and PTEN-null cells (PTEN-/-MEFs) were treated with insulin and/or DPI ,and then the phosphorylation level of Akt kinase was detected by western blot; the diversity of the cell growth was detected by MTT ; The levels of Cu/Zn SOD in PTEN+/+MEFs and PTEN-/-MEFs were detected by Western Blot and Northern Blot ;The level of the superoxide anion was detected by fluorescent probes ; The damage of DNA was detected by single cell alkalescence gel assay; MTT was used to assay the effect of H2O2 on the proliferation of the two types .Result: In the control PTEN+/+MEFs, insulin upregulated the phosphorylation level of Akt kinase, while the DPI could make the phosphorylation level of Akt kinase reversed to the basal-level. However these phenomena were not observed in PTEN-/-MEFs; Insulin facilitated the proliferation of the control PTEN+/+MEFs while DPI could abolish this kind of effect by antagonize the induction of H2O2 by insulin. However either of the treatments could not affect the proliferation of the PTEN-/-MEFs; The expression of Cu/Zn SOD both on the level of protein and mRNA was down regulated and the level of the superoxide anion increased in the PTEN-null cells (PTEN-/-MEFs);The antiproliferative effect of H2O2 to the cells decreased in PTEN-/-MEFs cells; Further more ,the damage of DNA was worse than PTEN+/+MEFs cells both in blank team and H2O2 treated team.Conclusion: The activation of Akt by peptide growth factors depends on the production of H2O2 and the presence of the tumor suppressor PTEN; The deletion of PTEN affected the expression of Cu/Zn SOD. As a result of it, the reactive oxygen species (ROS) kept on a high level, the oxidative damage accumulated and the antiproliferative effect of ROS to the cells decreased;the general conclusion is: the peptide growth factors could active Akt by inducing the production of H2O2 in the cells that PTEN(+), thus promoted the generation of the tumors; on the contrast, Cu/Zn SOD down regulated in the cells that PTEN(-), thus touch off the high level of the ROS and the accumulation of the oxidative damage, the decreased antiproliferative effect of ROS to the cells, and all the factors could promote the generation of the tumors collectively.