急性白血病患者凝血因子XⅢ表达和活性变化的研究

发布时间：2018-01-15 06:18

本文关键词：急性白血病患者凝血因子XⅢ表达和活性变化的研究　出处：《北京协和医学院》2017年博士论文　论文类型：学位论文

【摘要】：背景和目的凝血因子ⅩⅢ(FⅩⅢ)是一种重要的凝血因子,可促进纤维蛋白单体交联形成稳定的纤维蛋白凝块。FⅩⅢ本质是一种谷氨酰胺转移酶,除纤维蛋白外,其底物可多达140余种,因此参与到多种生理和病理过程中。FⅩⅢ由A亚单位二聚体(FⅩⅢ-A2)和B亚单位二聚体(FⅩⅢ-B2)结合而成,其中FⅩⅢ-A2在骨髓细胞中合成,主要存在于单核/巨噬细胞和巨核细胞中,FⅩⅢ-B2由肝细胞合成。研究发现,急性髓系白血病粒-单核细胞型(AML-M4)和单核细胞型(AML-M5)患者肿瘤细胞内FⅩⅢ-A2含量升高。随后发现急性早幼粒细胞白血病(APL)患者肿瘤细胞内FⅩⅢ-A2含量也升高。正常淋巴细胞内不表达FⅩⅢ-A2,但在部分急性淋巴细胞白血病(ALL)患者的肿瘤细胞内FⅩⅢ-A2的含量升高。本研究拟探讨急性白血病患者外周血FⅩⅢ浓度和活性的变化,以及该变化是否与骨髓或外周血肿瘤细胞内FⅩⅢ-A2的合成有关。此外,FⅩⅢ作为一种重要的凝血因子,目前仍无标准的活性测定方法,本研究拟建立FⅩⅢ活性测定方法,并分析急性白血病患者体内FⅩⅢ活性和浓度的变化与患者临床出血表现的相关性,同时测定血友病患者FⅩⅢ的浓度和活性,观察该类出血性疾病患者体内FⅩⅢ的变化。方法收集2016年11月至2017年5月于北京协和医院住院的初治急性白血病患者外周血和骨髓标本,统计患者的临床资料。分离外周血血浆,并提取外周血和骨髓单个核细胞的RNA。分别利用ELISA方法和凝血仪测定外周血血浆FⅩⅢ浓度和活性,利用实时定量PCR(qRT-PCR)方法测定单个核细胞内FⅩⅢmRNA的含量。分析患者经过治疗后外周血FⅩⅢ浓度和活性的变化,以及外周血和骨髓单个核细胞内mRNA表达量的变化。根据测定的外周血FⅩⅢ浓度和活性,分析其与急性白血病患者临床出血表现的相关性。利用ELISA方法和凝血仪测定血友病患者外周血FⅩⅢ浓度和活性的,分析血友病患者FⅩⅢ的变化。结果研究共纳入31例初治的急性白血病患者,其中AML24例,ALL7例。AML和ALL患者经过治疗后外周血FⅩⅢ浓度和活性的变化相反。AML和ALL患者治疗前外周血FⅩⅢ浓度分别为24.09(20.91)μg/mL和32.08±10.60μg/mL,治疗后FⅩⅢ浓度分别降低为18.43±4.74μg/mL和19.21±8.69μg/mL。AML患者治疗前外周血FⅩⅢ活性为72.75±28.98%,治疗后升高为94.74± 17.72%。ALL患者外周血FⅩⅢ活性在治疗后也有明显升高趋势。AML和ALL患者治疗前外周血和骨髓单个核细胞内FⅩⅢ-AmRNA的含量与相应对照组无明显差异。AML患者治疗前外周血单个核细胞内,FⅩⅢ A亚单位(FⅩⅢ-A)外显子 3 和外显子 14-15 的 △ Ct 值(△ Ct exon3 和 △ Ct exon14-15)分别为 0.93±2.36,1.53±2.21,骨髓相应的 △Ctexon3 和 △Ctexon14-15 分别为 2.21±2.45,1.70±2.40;均与相应对照组无明显差异。ALL患者治疗前外周血单个核细胞内 FⅩⅢ-A A Ct exon3 和 △ Ct exon14-15 分别为 1.49±1.43 和 1.66±1.48,骨髓相应的 △Ctexon3 和 △Ctexon14-15 分别为 1.30±1.58,1.85±2.94;均与相应对照组无明显差异。急性白血病患者外周血FⅩⅢ活性与患者临床出血表现无明显相关性。临床表现为关节出血的血友病A患者外周血FⅩⅢ浓度(10.74±3.15μg/mL)低于正常对照(16.55±3.29μg/mL),患者FⅩⅢ活性(80.63±20.52%)也低于正常对照(103.49±13.06%)。结论AML和ALL患者治疗后FⅩⅢ浓度低于治疗前,但FⅩⅢ活性高于治疗前,经过治疗后外周血FⅩⅢ浓度和活性的变化相反。急性白血病患者治疗前外周血和骨髓单个核细胞内FⅩⅢ-AmRNA的含量与对照组无明显差异,推测外周血中升高的FⅩⅢ浓度与急性白血病肿瘤细胞无明显相关,其来源尚需进一步的探究。急性白血病患者临床出血表现与外周血FⅩⅢ的活性降低无明显相关性。临床表现为关节出血的血友病A患者除FⅧ缺乏外,可能也存在外周血FⅩⅢ浓度和活性的相对降低。
[Abstract]:Background and objective: the blood coagulation factor XIII (F XIII) is an important factor that can promote fibrin monomer cross-linking to form a stable fibrin clot.F XIII in essence is a kind of transglutaminase, in addition to fibrin, the substrate can be up to more than 140, so to participate in a variety of physiological and pathological in the process of.F by A XIII subunit two dimers (F -A2 XIII) and B subunit two dimers (F XIII -B2) together, which F XIII -A2 synthesis in bone marrow cells, mainly in monocytes / macrophages and giant cells, F XIII by -B2 the liver cells formed. The study found that acute myeloid leukemia myelomonocytic type (AML-M4) and mononuclear cell type (AML-M5) increased tumor cells in patients with F XIII -A2 content. Then found in acute promyelocytic leukemia (APL) tumor cells in patients with F XIII -A2 content also increased. The expression of F in normal lymph cell x Part III -A2, but in acute lymphoblastic leukemia (ALL) patients were increased in tumor cell F XIII -A2. This study aims to investigate the changes in peripheral blood of patients with acute leukemia F XIII concentration and activity, and whether the change and bone marrow or peripheral blood tumor cells in the synthesis of -A2 F XIII XIII. In addition, F as an important factor, there is no standard method for the determination of the activity, intends to establish a method for determination of F activity of the XIII, and analyze the changes in patients with acute leukemia F XIII activity and concentration in patients with clinical manifestations of hemorrhage correlation, the simultaneous determination of the concentration and activity of patients with hemophilia F XIII, to observe the bleeding in patients with F disease XIII changes. Methods from November 2016 to May 2017 in Peking Union Medical College Hospital inpatients with acute leukemia bone marrow and peripheral blood specimens of patients with statistics Clinical data. Peripheral blood plasma separation, and extraction of peripheral blood and bone marrow mononuclear cells RNA. respectively using ELISA method and coagulation analyzer peripheral blood plasma F concentration and XIII activity, using real time quantitative PCR (qRT-PCR) to determine the content of mononuclear cell F XIII mRNA method analysis of patients after. After treatment, peripheral blood F XIII concentration and the changes of the activity, and the bone marrow and peripheral blood mononuclear cells mRNA expression changes. According to the assay of peripheral blood F XIII concentration and activity, and to analyze the clinical bleeding manifestations in patients with acute leukemia. The relationship between the ELISA method and the blood coagulation tester of hemophilia patients peripheral blood F XIII concentration and activity, analysis of changes in patients with hemophilia F XIII. Results a total of 31 cases of acute leukemia patients, which AML24 cases, ALL7 cases of.AML and ALL patients after treatment of peripheral blood F XIII concentrations and live The changes of.AML and ALL in patients before treatment, peripheral blood F XIII concentrations were 24.09 (20.91) g/mL and 32.08 + 10.60 g/mL, F after treatment were reduced to 18.43 XIII + 4.74 u g/mL and 19.21 + 8.69 g/mL.AML patients before treatment of peripheral blood F XIII activity was 72.75 + 28.98%, 94.74 + 17.72%.ALL increased after treatment for patients with peripheral blood F XIII activity in treated patients significantly increased trend of.AML content and ALL in peripheral blood and bone marrow mononuclear cells in F -AmRNA XIII had no obvious difference with the corresponding control group before treatment of.AML patients with peripheral blood mononuclear fine intracellular F XIII subunit A (F XIII -A) exon 3 and exon Ct 14-15 value (delta Ct exon3 and delta Ct exon14-15) were 0.93 + 2.36,1.53 + 2.21, bone marrow corresponding Delta Ctexon3 and delta Ctexon14-15 were 2.21 + 2.45,1.70 + 2.40; and the corresponding the control group There was no significant difference in.ALL patients before treatment in peripheral blood mononuclear cells of F -A A Ct exon3 XIII and Ct exon14-15 were 1.49 + 1.43 and 1.66 + 1.48, bone marrow corresponding Delta Ctexon3 and delta Ctexon14-15 were 1.30 + 1.58,1.85 + 2.94; there were no obvious differences with the corresponding control group. Patients with acute leukemia. Disease of peripheral blood F XIII activity and clinical bleeding showed no obvious correlation. The clinical manifestations of joint bleeding for patients with hemophilia A peripheral blood F XIII concentrations (10.74 + 3.15 g/mL) is lower than that of normal control (16.55 + 3.29 g/mL), with F activity XIII (80.63 + 20.52%) was lower than the normal the control (103.49 + 13.06%) is lower than F. Conclusion the concentration of AML and ALL XIII patients before treatment, but higher than that of the F XIII activity before treatment, after treatment of peripheral blood F XIII concentration and activity on the contrary. Peripheral blood before treatment in patients with acute leukemia and bone marrow In eukaryotic cells F XIII -AmRNA were not significantly different from the control group, that increased in the peripheral blood of F XIII concentration and acute leukemia tumor cells had no significant correlation, the source needs to be further explored. Acute leukemia patients with bleeding symptoms and peripheral blood F XIII activity decreased without obvious correlation. The clinical manifestations of joint bleeding in patients with hemophilia A except F VIII deficiency, relative reduction may also exist in peripheral blood F XIII concentration and activity.

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R733.71

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