Study On The Function Of Circadian Gene Per1 And Per2 Besides Its Role In Biological Rhythm Maintaining

ObjectiveInvestigate the signal transmission way and new function of circadian gene by experiments that interfering the circadian gene expression to attenuate animal's drug dependence and raise the circadian gene product level to suppress tumor growth. Investigate the role of circadian gene Period1 on drug reward to find a gene therapy method to drug addiction. Investigate the role of circadian gene Period2 on tumor proliferation and apoptosis.Method and Result1, Research of relationship between mper1 gene and drug dependenceMethod 1,. Plasmid pcDNA 3.1-per1RZ contains coding sequence of ribozyme per1RZ, a hammerhead ribozyme that can selectively cleave the Period1 mRNA, based on eukaryotic expression vector pcDNA 3.1 was constructed. In vitro cleavage experiment prove the target cleave Period1 mRNA ability of per1RZ. Conditional place preference(CPP) paradigm used to investigate the effect of Intracerebroventricular (ICV) injection of pcDNA 3.1-per1RZ on drug reward in mice.Result Quantitative analysis of in vitro cleavage experiment shows the efficacy of ribozyme per1RZ; about 60% of the Period1 mRNA was cleaved by ribozyme per1RZ. CPP test display the block of drug reward inpcDNA 3.1-per1RZ ICV injection group. Period1 expression in brain was attenuated of pcDNA 3.1—per1RZ ICV injection group demonstrated by western blot.2, Research of relationship between mper2 gene and tumor developmentMethod Transfect the eukaryotic mPeriod2 expression vector (pcDNA 3.1 -mPer2) based on pcDNA 3.1 into the in vivo cultured Lewis tumor cell by liposome method. Immunohistochemistry and flowcytometry were used to detect the expression of mPeriod2 in transfected Lewis tumor cell. Flowcytometry was used to examine the proliferation and apoptosis of Lewis tumor cell.Result Expression of PERIOD2 protein in pcDNA 3.1-mper2 transfected Lewis tumor cell was proved by immunohistochemistry and flowcytometry. Flowcytometry results of mPeriod2 expression Lewis tumor cell show less proliferation and high rate of apoptosis.ConclusionInterfere the Period1 expression in brain could attenuate the psychological dependence of drug in mammals. Period2 expression could suppress the proliferation of tumor cell and increase the apoptosis.