Dissertation > Excellent graduate degree dissertation topics show

DEK Expression and clinical significance in chronic lymphocytic leukemia

Author: WangDongMei
Tutor: XuWei
School: Nanjing Medical University
Course: Internal Medicine
Keywords: Chronic lymphocytic leukemia DEK Real - time quantitative RT-PCR Apoptosis p53 Fludarabine Nutlin-3
CLC: R733.7
Type: Master's thesis
Year: 2011
Downloads: 34
Quote: 0
Read: Download Dissertation

Abstract

The purpose of chronic lymphocytic leukemia (chronic lymphocytic leukemia, CLL) is the most common type of leukemia in Western countries, accounting for 1/3 of the adult leukemia incidence in China is lower than in Western countries, but with the aging of the population, CLL incidence increased year by year trend. The DEK gene located on chromosome 6p22.3, expressed in the proliferation of cells and tumor cells, the earliest in acute myeloid leukemia (acute myeloid leukemia, AML) found in t (6; 9), along DEK may be involved in tumor formation mainly involved in interfering with cell division, DNA repair, inhibition of cell differentiation, senescence and apoptosis. DEK gene expression found in a variety of tumors in recent years, have also been reported in CLL and related carcinogenicity may affect p53 function, the present study was to investigate the CLL the DEK gene mRNA expression levels and its clinical significance. Using real-time quantitative reverse transcription PCR (quantitative reverse transcription-polymerase chain reaction qRT-PCR) technology, SYBR Green fluorescent dye was detected 65 cases of CLL patient peripheral blood lymphocyte the DEK gene expression level, using the threshold cycle (cycle threshold, Ct ) Comparison Method for relative quantitative analysis of mRNA expression, the relative expression levels of these genes are expressed in formula 2 (- △ Ct), wherein △ Ct = Ct target gene - Ct control gene. The application of statistical software SPSS17.0 statistical analysis of the data, the differences in gene expression levels between the different packet using the Mann-Whitney U test, P lt; 0.05 difference was statistically significant. ① 65 CLL patients peripheral blood lymphocytes DEK mRNA expression median 6.792 × 10-2 (1.438 × 10-2 to 3.201 × 10-1). The ② DEK expression levels of ≥ 60 years of age and lt; 60 years of age (P = 0.773), clinical stage earlier and later clinical stage (Binet A) (expression of Binet B phase, C phase) (P = 0.612), serum increased levels of lactate dehydrogenase (lactate dehydrogenase, LDH) (P = 0.115) and β2-microglobulin (β2-microglobulin, β2-MG) (P = 0.330) and normal patients, as well as the expression of ZAP-70 positive and negative ( P = 0.552) no significant difference between patients; CD38 expression in patients with positive DEK expression levels were significantly higher than negative patients (P = 0.047); fluorescence in situ hybridization (fluorescence in situ hybridization, FISH) to detect the existence del ( 13q14), 12, immunoglobulin heavy chain (immunoglobulin heavy chain IgH) translocation and del (11q22.3) abnormalities in patients with no such patients with abnormal was no significant difference (P values ??were 0.159,0.546 0.883,0.498), del (17p13) patients with abnormal DEK level significantly higher than the non-del (17p13) patients with abnormal (P = 0.006); DEK expression level of p53 gene mutations in patients with p53 gene mutations in patients phase no significant difference (P = 0.655); immunoglobulin heavy chain variable region (immunoglobulin heavy chain variable region IgVH) mutations in patients with DEK expression levels were significantly higher than IgVH mutation patients (P = 0.025). Conclusion DEK expression level of p53 gene deletion and IgVH mutation status. DEK expression levels may be associated with the prognosis of CLL patients. The purpose of chronic lymphocytic leukemia (chronic lymphocytic leukemia, CLL) is a heterogeneous hematologic malignancies, chemotherapy effect to some extent determines the prognosis of patients with fludarabine for the treatment of CLL commonly used drugs by antagonizing purine metabolic cause DNA damage to activate the p53 gene, so as to play its role in anti-tumor CLL cells into the mitochondrial apoptotic pathway. Nutlin-3 is a potent selective MDM2 2 (murine double mimute 2, MDM2) antagonist, wild-type p53 pathway activation by antagonizing MDM2. In this study, the active substance with fludarabine (F-ara-A) and Nutlin-3 in vitro stimulation of CLL cells, to explore the role and status of DEK in p53-dependent apoptosis pathway. Method sterile separation of the 26 cases in the peripheral blood of patients with CLL CLL cells cultured in vitro, set up to contain the the 3.5μM fludarabine Nutlin-3 10 μM of the experimental group and drug-free control group, the cells were collected after 24h culture. Flow cytometry of annexin V-FITC (annexin V) / propidium iodide (propidium iodide, PI) to clarify the degree of apoptosis; SYBR Green fluorescent dye method of real-time quantitative RT-PCR (quantitative reverse transcription-polymerase chain reaction, qRT-PCR) was used to detect the expression level of DEK; disease mutation of interphase fluorescence in situ hybridization (fluorescence in situ hybridization, FISH) to detect ataxia telangiectasia (ataxia-telangiectasia mutated, ATM) gene and p53 gene missing; PCR Combined DNA sequencing to detect immunoglobulin heavy chain variable region (immunoglobulin heavychain variable region, IgVH) gene mutation and p53 gene mutations; p53/p21 protein level by flow cytometry CLL cells to clear the p53 gene function status. Application of statistical software SPSS17.0 statistical analysis of the data, the level of apoptosis in CLL cells between the experimental and control groups, and the level of gene expression differences using a paired t-test, P lt; 0.05 difference was statistically significant. Results ① CLL cells separated median apoptotic rate was 8.62% (1.83% to 36.28%), simple in vitro culture 24h apoptosis rate was 19.23% (7.96% to 55.94%) and pure cultured in vitro for 24h CLL cells spontaneously apoptosis increased significantly (P = 0.004); fludarabine and Nutlin-3 stimulate 24h apoptosis rate were 40.85% (4.59% to 75.31%) and 42.82% (18.11% to 79.71%), fludarabine the foreshore and Nutlin-3-induced apoptosis compared with the control group, a significant increase (P lt; 0.001 and P lt; 0.001); ② 8 cases of CLL cells were treated with fludarabine before and after stimulation of p21 protein levels did not change, suggesting that p53 dysfunction. Three cases of CLL cell after fludarabine stimulation, the presence of the ATM gene loss of heterozygosity, two cases of cytoplasmic p21 protein expression was significantly increased, suggesting that the normal p53 function; (3) the experimental group CLL cells were treated with fludarabine and Nutlin- 3 stimulate 24h the DEK water of normal p53 function CLL cells were significantly lower than those in the control group (P = 0.042 and P = 0.038); abnormal p53 function CLL cells were treated with fludarabine and Nutlin-3 stimulation 24h after CLL cells compared with the control group, DEK expression levels had no significant change (P = 0.834 and P = 0.477); ④ experimental group CLL cells after fludarabine and Nutlin-3 stimulation after 24h, p53 gene deletion the DEK significantly were lower than the control group (P = 0.017 and P = 0.029); CLL cells with p53 gene deletion after fludarabine and Nutlin-3 stimulus 24h, compared with the control group, DEK expression level There was no significant change (P = 0.111 and 0.378); ⑤ experimental group CLL cells without p53 gene mutations in CLL cells after fludarabine and Nutlin-3 stimulation after 24h, DEK was significantly lower than the control group (P = 0.037 CLL cells with p53 gene mutation fludarabine and Nutlin-3 stimulation 24 hours, compared with the control group, the DEK expression levels had no significant change (P = 0.263 and P = 0.378) and P = 0.017); . Conclusion fludarabine and Nutlin-3 can be effective in inducing apoptosis in CLL cells. lacking p53 and / or p53 mutations can lead to abnormal p53 function, but the ATM gene loss of heterozygosity of CLL cells with normal p53 function. Lowered DEK level of expression is dependent on the normal p53 gene.

Related Dissertations

CLC: > Medicine, health > Oncology > Hematopoietic and lymphoid neoplasms > Leukemia
© 2012 www.DissertationTopic.Net Mobile