亚甲蓝病毒灭活血浆与新鲜冰冻血浆比较蛋白质组学研究

发布时间：2018-05-15 18:57

本文选题：输血 + 新鲜冰冻血浆　；参考：《吉林大学》2017年硕士论文

【摘要】：目的:血浆占血液成分的50%,血浆输注是临床治疗的重要组成部分。临床主要应用亚甲蓝病毒灭活血浆和新鲜冰冻血浆,以往涉及两种血浆的对比研究多注重于分析血浆凝血因子、血浆蛋白含量及相应临床生化指标的改变,但对这两种血浆中特异性差异蛋白的研究甚少。本课题应用比较蛋白质组学的研究方法分析亚甲蓝病毒灭活血浆和新鲜冰冻血浆这两种血浆中特异性差异蛋白的改变,为临床医生对需要输血病人的治疗提供实验和理论依据,以减少血浆不合理输注和输注无效,特别是对一些特殊病例提供参考资料。方法:1.对临床62例输注不同血浆患者的输血后凝血酶原时间和纤维蛋白原含量进行统计学分析。2.三氯醋酸(TCA)/丙酮沉淀法去除血浆中高丰度白蛋白。3.硫酸铵沉淀法去除血浆中高丰度白蛋白。4.SDS-PAGE分离鉴定去除血浆中高丰度白蛋白后的血浆蛋白。5.Clean-up去除血浆蛋白中干扰2DE的物质。6.SDS-PAGE和2-DE验证clean-up是否有效去除血浆蛋白中干扰2DE的物质。7.2-DE分离病毒灭活血浆和新鲜冰冻血浆的血浆蛋白。8.采用质谱法分析鉴定两种血浆中显示差异明显的蛋白点。9.重新收集两种不同的血浆18-74例,进行Western blot和ELISA检测验证其中两种差异明显的蛋白质含量。结果:1.经统计分析临床中应用两种不同血浆后患者的化验数据,结果提示两种血浆的应用均具有一定的治疗效果,但新鲜冰冻血浆(FFP)组患者的凝血酶原时间(PT)值优于亚甲蓝病毒灭活血浆(MB-FP)组(P0.05),有统计学意义,而两种的纤维蛋白原含量差异(P0.05),无统计学意义。2.三氯醋酸/丙酮沉淀法去除高丰度白蛋白比饱和硫酸铵沉淀法去除高丰度白蛋白的效果好。3.两种方法去除血浆中高丰度白蛋白后的血浆,再应用clean-up方法对去除血浆中的干扰2DE的物质效果不明显。4.亚甲蓝病毒灭活血浆与新鲜冰冻血浆的2DE图谱分析显示,两种血浆出现一些差异蛋白质的表达。5.经过质谱分析及数据库比对鉴定出8个差异蛋白质点(含6种蛋白质)发生了修饰作用,它们是:Complement C1r subcomponent,Inter-alpha-trypsin i nhibitor heavy chain H4,Keratin(type II cytoskeletal 1),Hemopexin,Fibrinoge n gamma chain,Transthyretin.我们选取了Hemopexin,Keratin(type II cytoskel etal1)进行了ELISA和Western blot验证。6.对18例血浆的Western Blot检测结果表明在新鲜冰冻血浆和病毒灭活血浆中均检测到Keratin,type II cytoskeletal1,但其蛋白条带的表观差异并不明显。7.对48例血浆的ELISA检测结果表明,新鲜冰冻血浆中Hemopexin含量明显高于亚甲蓝光化学法病毒灭活的血浆。结论:临床输注新鲜冰冻血浆后患者的凝血酶原时间(PT)明显优于输注亚甲蓝病毒灭活血浆。比较蛋白组学的研究结果揭示,亚甲蓝病毒灭活血浆的同时使血浆中一些特殊蛋白质如:Complement C1r subcomponent,Inter-alpha-trypsin,Inhibitor heavy chain H4,Keratin,type II cytoskeletal 1,Hemopexin,Fibrinogen gamma chain,Transthyretin发生了明显的修饰作用,而这些修饰作用对蛋白质的功能有何影响、对血浆临床应用的效果有何影响值得我们深入研究。
[Abstract]:Objective: plasma is 50% of the blood components. Plasma infusion is an important part of clinical treatment. Clinical application of methylene blue virus inactivated plasma and fresh frozen plasma. The comparative study of two kinds of plasma in the past mainly focuses on the analysis of plasma coagulation factor, plasma protein content and the changes of corresponding clinical biochemical indexes, but the two kinds of plasma plasma protein content and corresponding clinical biochemical indexes change. There are few studies on specific differential proteins in plasma. This subject uses comparative proteomics to analyze the changes in specific differential proteins in two kinds of plasma, methylene blue virus inactivated plasma and fresh frozen plasma, to provide experimental and theoretical basis for clinicians to treat patients in need of blood transfusion in order to reduce the unreasonable plasma levels. Ineffective infusion and infusion, especially for some special cases. Methods: 1. pairs of clinical 62 patients with different plasma plasma prothrombin time and fibrinogen content after transfusion were statistically analyzed by.2. three chloroacetic acid (TCA) / acetone precipitation method to remove the high abundance albumin.3. ammonium sulfate precipitation in plasma to remove plasma High abundance albumin.4.SDS-PAGE separation and identification of plasma protein.5.Clean-up after removal of high abundance albumin in plasma to remove the substance.6.SDS-PAGE and 2-DE that interfere with 2DE in plasma protein to verify whether clean-up effectively removes the substance that interferes with 2DE in plasma protein,.7.2-DE separation virus inactivating blood plasma and fresh frozen plasma protein.8. extraction Mass spectrometric analysis was used to identify two different plasma levels of protein point.9., which were significantly different in two kinds of plasma, and two different plasma levels were re collected, and Western blot and ELISA tests were used to verify the protein content. Results: 1. the data of two different blood plasma patients were statistically analyzed by statistical analysis. The results suggested two kinds of blood. The application of the plasma has a certain therapeutic effect, but the prothrombin time (PT) value of the fresh frozen plasma (FFP) group is better than the methylene inactivated plasma (MB-FP) group (P0.05), and the difference between the two kinds of fibrinogen (P0.05) is not statistically significant, and there is no statistically significant.2. three chloroacetic acid / acetone precipitation method to remove the high abundance albumin ratio. The effect of the removal of high abundances with saturated ammonium sulfate precipitation method is good.3. two methods to remove the plasma of high abundance albumin in plasma, and then the effect of clean-up method on removing the interference of 2DE in plasma is not obvious, and the analysis of 2DE Atlas of.4. methylene blue virus inactivated plasma and fresh frozen plasma shows that some kinds of plasma appear some of them. The expression of differential protein.5. identified 8 different protein points (including 6 proteins) through mass spectrometric analysis and database comparison. They are: Complement C1r subcomponent, Inter-alpha-trypsin I nhibitor heavy chain H4, Keratin (type 1). We selected Hemopexin, Keratin (type II cytoskel etal1) and ELISA and Western blot verification.6. for Western Blot test results of 18 cases of plasma. The results showed that the content of Hemopexin in fresh frozen plasma was significantly higher than that of methylene blue photochemical inactivated plasma. Conclusion: the prothrombin time (PT) of patients after infusion of fresh frozen plasma was significantly better than that of inactivated methylene blue virus inactivated plasma. Comparison of the results of proteomics revealed that methylene blue virus inactivated plasma. There are some special proteins in plasma such as Complement C1r subcomponent, Inter-alpha-trypsin, Inhibitor heavy chain H4, Keratin, type II cytoskeletal 1. What is the effect of the effect is worth studying in depth.

【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R457.1

【相似文献】