Evaluation Of Period2Effect On Human Glioma Cell Growth In Vivo

Objective: Study circadian clock gene Period2(Per2) in U343human gliomacells over-express, Per2gene on human glioma cell growth and proliferationability.Methods: Cultured U343human glioma cells in vitro, divided into three groups:Per2gene over-expression group, empty vector group and U343blank control group.Cloned full-length Per2gene construct pcDNA3.1(+)per2eukaryotic expressionplasmid sequencing (completed). With LipofectamineTM2000liposomes,pcDNA3.1(+)per2and pcDNA3.1(+) were transfected into U343cells respectively,building the Per2gene over-expression of U343-pcDNA3.1(+)per2cells and emptyvector U343-pcDNA3.1(+) cells.Buy SPF level (specific pathogen free) BALB/C Nude genetic background nudemice45, male,4-5weeks old, were randomly divided into three groups for thestatistical principle, each15, cultured①Per2gene over-expression inU343-pcDNA3.1(+)per2cells②empty vector U343-pcDNA3.1(+) cells in③U343cells to logarithmic phase respectively, conventional digestion cells, take the1×107cells (0.2ml), routine disinfection of the skin,1ml syringe inoculatedin nude mice subcutaneously subcutaneously transplanted tumor animal model.Experimental groups of nude mice were fed for21days to observe the groupsof transplanted tumors in nude mice, tumor growth and tumor formation time, toa diameter of about0.2cm x0.2cm when considered as tumor formation, and recordedtumorigenicity days; three groups of nude miceeach group every7days5were killed tumor tissue and compare the size.Observed tumor cases to remove the tumor tissue after dehydration, embeddedin paraffin, HE staining, PCNA immunohistochemical method to detect the Per2geneover-expression group, the proliferation of the tumor tissue of the empty vectorgroup and the control group, were statistically analyzed.Results: The eukaryotic plasmid pcDNA3.1(+)per2and pcDNA3.1(+) into U343glioma cells build Per2gene transiently transfected cells over-expressionU343-pcDNA3.1(+)per2and empty vector U343-pcDNA3.1(+) cells. Successfullyestablished the Per2over-expression Groups, empty vector group and U343blankcontrol group nude mice model.The Per2gene over-expression of tumor formation time was4.7±0.27days;the empty vector aneurysms time was3.2±0.27days; U343blank control tumorformation time was3.5±0.35days, tumor formation rate was100%.7to14daysafter the inoculation of tumor cells, tumor growth was significantly active tumorvolume increases rapidly, the small volume of the Per2gene over-expression grouptumor tissue compared with empty plasmid group and U343blank control group, andpairwise comparison was statistically significant (P <0.05). HE staining fortumor tissue, PCNA immunohistochemistry results show: Per2gene over-expressionof tumor tissue positive expression reduced after pairwise statisticalcomparison results show, Per2gene over-expression group with empty vector group,U343blank control group was significantly different (P <0.05), no significantdifference (P>0.05) between the empty plasmid group and negative control group.Conclusion: This study successfully simulated human glioma cells in vivostate to prove that the circadian clock gene Per2inhibition of U343glioma cellproliferation in vivo. Provide a theoretical basis for targeted therapy of humanglioma as a key point for the follow-up to the circadian clock gene Per2.