Expression Of Survivin In Acute Leukemia And Construction Of Human Survivin Eukaryotic Expression Vector And Its Expression In K562 Cells

ObjectiveLeukemia is a clonal disease of hematopoiesis with poor clinical outcome despite recent improvements in chemotherapy and stem cell transplantation regimens. Most of the patients achieve a complete hematological remission by chemotherapeutical regimes. However, the long-term prognosis for all AL patients is rather poor . The malignant cells failure to express accessory molecules and tumor specific antiges that can't induce effective antitumor immunity. Antitumor immunity depends on cellular immunity mainly. With regard to tumor therapy, more and more people play focus on adjusting auto-immunity or exploring operative immune components to eliminate tumor.Survivin is antiapoptotic protein that belongs to the family of inhibitory apoptotic proteins(IAP). It is essential for cancer cell survival because of its role in regulation of the balance between cell proliferation and programmed cell death during cell cycle. Survivin is overexpressed by practically all solid tumors and is poorly expressed by normal adult cells and tissues. The overexpression of survivin in tumors correlates with a more aggressive disease, poor survival of patients, and drug resistance.To investigate the expression rate of survivin in acute leukemia patients and its clinical significance. We cloned human survivin gene from bone marrow in this test,and construct its eukaryotic expression vector,and then express it in K562 cells. These results lay a foundation for the futher research on the function of surviving in the development and therapy of leukemia.MethodsPart 1: Extract mRNA from 64 acute leukemia patients and 14 donors . Detected the concentration and purity coefficient of mRNA .The RNA of survivin was cloned by RT-PCR technique. Accredit the PCR production and then statistical analysis.Part 2: To clone and express human survivin gene with eukaryotic vector. Be sure the accession BC065497 in GeneBank is the survivin mRNA for transfected. The surivin gene was cloned into pIRES2-EGFP vector and the pIRES2-EGFP-Survivin vector was constructed and transfected into K562 cells. Accredit the transfected production by RT-PCR and prepare for explore the significance of this vector in leukemia curing. Results 1. The positive rates of survivin was higher than normal control(NC). The expression positive-rate of primary cured patients and relapse patients was significantly higher than that in NC.2. The expression of survivin in de novo AL was higher than that in NC.1n replased patients the expression of survivin also higher than that in NC.3. Survivin gene has been cloned into pIRES2-EGFP vector.4. pIRES2-EGFP-Survivin vector expressed in K562 cells successfully.4. The results of RT-PCR indicated that pIRES2-EGFP-Survivin coule increase the expression levels of survivin gene mRNA in transfected K562 cells.Collusions1. Survivin may play an important role in the development and prognosis of acute leukemia. Abnormal expression of survivin genes was related to pathogenesis and progression of AL and may be associated with therapeutic efficacy and prognosis in acute leukemia.2. The recombinant plasmid pIRES2-EGFP-Survivin were successfully constructed and has expressed successfully in K562 cells. It can be used for further study of survivin on diagnosis and therapy leukemia.