Study On The Effect Of Time Rhythm Gene On Leukemia And Its Mechanism

Objective The possible mechanism of the gene family in leukemia is studied by analyzing the expression of CLOCK,Bmal1,Per1,Per2,Per3,Cry1,C-myc,CyclinD1 and Wee1 in leukemia.Through the analysis of circadian gene Per2 in resistant CML cell line KCL22 cells proliferation,apoptosis and differentiation effect,to further explore the Per2 gene play a role in the process of drug resistance in leukemia,in order to find a new target for the treatment of leukemia.Method The expression level of circadian gene CLOCK,Bmal1,Per1,Per2,Per3,Cry1 and cell cycle gene C-myc,CyclinD1 and Wee1 in 30 cases of CML,50 cases of AML and 30 normal controls were detected by qRT-PCR.Construction of Per2 expression lentivirus transfected CML cell line KCL22 cells,and screening stable strains.The proliferation of KCL22 cells was detected by CCK-8,and apoptosis and differentiation of KCL22 cells were detected by flow cytometry.Results Compared with normal people,the expression of circadian gene CLOCK and cell cycle gene C-myc and CyclinD1 increased in AML and CML,while the expression of rhythmic genes Bmal1,Per1,Per2,Per3 and cell cycle gene Wee1 expression decreased(P<0.05).There is a linear correlation between CLOCK,Bmal1,Per1,Per2,Per3,Cry1,Wee1,C-myc,CyclinD1 in AML and CML(P<0.05).In AML,Per1 was positively correlated with the expression level of Per2(r=0.538,P<0.01).Per1 was positively correlated with the expression level of Per3(r=0.497,P<0.01).Per2 was positively correlated with the expression level of Per3(r=0.376,P<0.05).In CML,Per1 was positively correlated with the expression level of Per2(r=0.658,P<0.01),Per1 was positively correlated with the expression level of Per3(r=0.592,P<0.01),and Per2 was positively correlated with Per3 expression level(r=0.385,P<0.05).We have successfully constructed the expression of Per2 lentivirus and transfected CML cell line KCL22 cells and screened the KCL22/Per2 stable overexpression of Per2 strains and used it as the experimental group.Comparison with control group(KCL22/untreated)and no-load group(KCL22/empty),the overexpression of Per2 causes the proliferation of KCL22 cells to be inhibited,induces cell cycle G1 cell growth[experimental group(41.46±0.01)%,no load group(30.51±0.32)%,the control group(27.07±0.6)%,P<0.05],and has no obvious effect on cell apoptosis[experimental group(7.63±0.12)%,no load group(7.98±0.21)%,the control group(8.33±0.01)%,P>0.05].Conclusion CLOCK,C-myc and CyclinD1 play a role as tumor promoting factors.Bmal1,Per1,Per2,Per3,Cry1 and Wee1 play a role as tumor suppressor factors.Moreover,these rhythmic genes interact with each other to regulate the circadian rhythm of the human body,thereby affecting the proliferation of tumor cells.Among them,Per1,Per2 and Per3 play an important role in the occurrence and development of leukemia.Overexpression of Per2 can inhibit the growth and proliferation of Chronic granulocyte KCL22 cells,induce cell cycle arrest in G1 phase,and have no obvious effect on cell apoptosis.