Polyamines In Vitro Microenvironment On Thetransfer Of HepG2Cells

Objective:Polyamine is a long-chain aliphatic compounds.we usually say that polyamines are putrescine (Put),spermine (spm) and spermidine (spd). Polyamines are widely found in nature, which is necessary for mostvital organs. Physiological polyamine content in human tissues and body fluids is very small.But many ofpolyamine content in tumor tissue is more than in normal tissue and is related to tumor growth.This experiment through additional polyamine simulates the human body high,normal and low threedifferent polyamine microenvironment for cell growth,invasion,and metastasis of liver cancer ability andrelated molecular mechanisms.Methods:By HepG2cells for the study, Akt3gene was transfected into HepG2cells by gene transfection. Theexogenous polyamines by high concentrations of polyamines putrescine0.8nmol/mL, spermidine10.0nmol/mL, spermine5.0nmol/mL; normal(middle) polyamine concentration of putrescine0.08nmol/mL, spermidine1.0nmol/mL, spermine0.5nmol/mL; low polyamine concentration of putrescine0.008nmol/mL, spermidine0.1nmol/mL, spermine0.05nmol/mL were added to HepG2cells andHepG2-Akt3cells. HepG2cells and HepG2-Akt3cells were cultured for15and30days in the exogenouspolyamines. Each cell has negative control. To detect the effect of different concentrations of polyamineson two different cell proliferation applys the colony-forming ability of the experiment.To detect the effectof different concentrations of polyamines on two different cell migration applys the Transwell migration assay (unpaved glue); To detect the effects of different concentrations of polyamines on two different cellwound healing capabilities uses cell wound scratch assay.To analyze different polyaminesmicroenvironment on tumor cell adhesion, migration, invasion and various other related protein expression,such as ornithine decarboxylase (ODC), spermidine/spermine acetyltransferase (SSAT), AKT, P27kip1,hypoxia-inducible factor (HIF1α), vascular endothelial growth factor (VEGF), cadherin protein(E-cadherin), integrins (Integrinα6), Akt3, changes in cathepsin D (Cathepsin D) and other proteins usesWestern blot test.Results:Non-transfected HepG2cells under the different polyamine microenvironment show the differentability of proliferation, migration, invasion, and with the increase of concentration of polyamine itsproliferation,migration and invasion ability strengthens gradually in a dose-and time-dependent manner.HepG2cells transfected Akt3under the different polyamine microenvironment also show the differentability of proliferation, migration, invasion, and with the increase of concentration of polyamine itsproliferation, migration and invasion ability strengthens gradually in a dose-and time-dependentmanner.And HepG2cells transfected Akt3of the proliferation, migration and invasion ability are strongerthan Non-transfected HepG2cells under the same concentration of polyamine.Protein immunoblotexperimental results show that polyamine is through the induction of AKt3protein, HIF1α protein andornithine decarboxylase (ODC), spermidine and spermine acetyltransferase (SSAT), vascular endothelialgrowth factor (VEGF) and Integrin α6, Cathepsin D protein increase and P27kip1protein, calciumepithelial adhesion protein (E-cadherin) cut to achieve the high invasion, metastasis and proliferation.Conclusion:Polyamine can promote HepG2cells, HepG2-Akt3cell proliferation, invasion, metastasis, a HepG2 cells transfected Akt3of proliferation,invasion and metastasis ability is better than non-transfected HepG2cells. Akt3gene in the polyamine promoting tumor development has an important role.