LBP基因多态性、APACHEⅡ评分和降钙素原对ARDS预警作用的初步研究

发布时间：2017-12-31 18:02

本文关键词：LBP基因多态性、APACHEⅡ评分和降钙素原对ARDS预警作用的初步研究　出处：《第三军医大学》2014年硕士论文　论文类型：学位论文

【摘要】：背景：急性呼吸窘迫征（Acute respiratory distress syndrome，ARDS）是一种由肺部感染、创伤及急性胰腺炎等多种肺内、外致病因素导致的肺部炎症反应，，以肺顺应性降低、顽固性低氧血症及非心源性肺水肿为主要临床表现，预后差。尽管近年对ARDS的发病机制有了深入的理解，临床治疗ARDS的策略也有一定的改进，但其死亡率仍没有明显下降。因此，如能早期甄别ARDS高危患者，及时进行ARDS的防治，可望降低ARDS发病率、改善其预后。然而，由于ARDS的发病因素复杂，影响疾病进展的因素多，目前尚无理想的ARDS预警模型。目前的研究表明，仅依靠患者临床资料或一些肺部炎症反应、细胞损伤相关的生物标志物并不能取得满意的预测效果。因此，本研究拟分别从与ARDS发病相关的基因及临床资料联合生物标志物两方面着手，探讨LBP基因多态性、APACHEⅡ评分和降钙素原对急性呼吸窘迫综合征的预警作用。研究目的：本研究拟探讨LBP基因多态性、APACHEⅡ评分和降钙素原对急性呼吸窘迫综合征的预警作用，以期得到较为理想的ARDS预警模型。研究内容和方法： 1.实验设计：本研究分为以下两个部分：(1)以222例内毒素血症患者为研究对象，通过snapshot方法检测LBP15个SNP位点的基因多态性，结合患者是否发展为ARDS，寻找LBP与ARDS易感性相关的SNP位点及基因型。(2)以78例肺部感染患者为研究对象，结合患者是否发展为ARDS，绘制受试者工作特征曲线，探讨APACHEⅡ评分、降钙素原对肺部感染患者发展为ARDS的预测价值。 2.外周血的处理和DNA提取：在患者入住ICU24小时内采集5ml EDTA抗凝静脉血，分离血浆并进行内毒素检测。提取DNA，于-80℃冰箱保存备用。 3. snapshot方法检测患者LBP基因15个SNP位点信息。 4.记录患者的临床基线资料，包括APACHEⅡ评分，年龄、性别、病人来源，慢性合并症，入院诊断等基本人口学资料和采集血液标本时患者的细菌培养结果。其中肺部感染患者还需记录动脉血气、血沉、C-反应蛋白、乳酸、降钙素原及1周内的痰培养结果。 5.统计方法：每个SNP的等位基因及基因型的频率根据基因检测结果计算得到，基因型分布的检验通过HWE（Hardy Weinberg Equilibrium,哈迪-温伯格平衡定律）检验得到，P0.05表示符合哈迪-温伯格平衡定律。LD图的构建通过Haploview软件制作得到。rs2232618与ARDS易感性的分析通过卡方检验获得。年龄、性别、内毒素浓度等混杂因素的影响通过logistic回归方法排除。正态分布资料的组间比较采用t检验或方差分析，非正态分布资料的组间比较采用秩和检验。预测作用采用受试者工作特征曲线（ROC曲线）方法，两预测指标的联合采用二元Logistic回归方法。研究结果： 1.基因预警实验中，总共纳入内毒素血症患者222例，其中男性154例，女性58例，年龄61.0±17.3（17～93）岁。7天内发展为ARDS的患者49例，ARDS发生率为22.1%。革兰阴性菌感染66例，占29.7%；其他病原菌及混合感染46例，占21.1%；未得到明显致病菌110例，占49.2%。 2.222例内毒素血症患者15个SNP位点的MAF与HapMap数据库中相似。其中13个SNP位点基因型的分布符合哈迪-温伯格平衡定律(P0.05)，提示其等位基因及基因型频率在人群中的遗传是恒定的。APACHEII评分在rs2232618位点不同的基因型中无差异（P0.05）。 3.仅rs2232618与ARDS的易感性有关。携带有等位基因C（基因型CC/CT）的患者其发生ARDS的概率明显高于不携带等位基因C（基因型TT）的患者（P=0.001）。通过logistic回归分析发现，在排除年龄、性别、内毒素浓度等影响后，rs2232618与ARDS的易感性仍密切相关(P=0.002)。 4.临床资料及生物标志物预警实验中，总共纳入肺部感染患者78例，其中男性患者60例，女性患者18例，年龄65.2±17.6(25～93)岁。7天内发展为ARDS的患者有33例，ARDS发生率42.3%。ARDS组APACHEⅡ评分和降钙素原显著高于非ARDS组，分别为[(19.9±8.3) vs (15.6±6.0), P=0.013]和(12.6±11.4)ng/L vs (7.5±10.5)ng/L, P=0.046]，氧合指数低于非ARDS组，为[(109.9±82.6) vs (252.1±92.4), P=0.003]。 5. APACHEⅡ评分和降钙素原对肺部感染诱发ARDS的预测作用相当（AUROC:0.651vs0.657，P=0.929），且两者联合后，对ARDS的预测作用较两者单独的预测作用无显著提高。结论： 1. LBP基因rs2232618位点携带等位基因Ｃ（基因型CC+TC）的患者更易发生内毒素性ARDS。该位点T→C的突变可能是预测内毒素性ARDS的一个较为理想的预警指标。 2. APACHEⅡ评分和血浆中降钙素原的表达与肺部感染患者的ARDS发生率密切相关。两者对肺部感染诱发的ARDS均有一定的预测价值，但两者联合后的预测价值并无明显提高。
[Abstract]:Background: acute respiratory distress syndrome (Acute respiratory distress syndrome, ARDS) is a lung infection, trauma and acute pancreatitis such as lung, lung inflammation leads to external pathogenic factors, to reduce lung compliance, refractory hypoxemia and non cardiogenic pulmonary edema were the main clinical manifestations, despite the depth of poor prognosis. The understanding of the pathogenesis of ARDS in recent years, the clinical treatment of ARDS strategy also has some improvement, but the mortality rate is still not decreased significantly. Therefore, if early screening high-risk patients with ARDS, timely prevention and treatment of ARDS, is expected to reduce the incidence of ARDS, improve the prognosis. However, due to the incidence of ARDS complex which factors influence the progress of the disease, there is no ideal ARDS early warning model. The present study shows that only rely on the clinical data of patients or some lung inflammation, related biomarkers of cell injury It can not achieve satisfactory prediction effect. Therefore, this study is going to start from two aspects of gene and clinical data combined with biomarkers associated with the pathogenesis of ARDS, and explore the early warning effect of LBP gene polymorphism, APACHE II score and procalcitonin on acute respiratory distress syndrome.
Research purposes: the purpose of this study is to explore the early warning effect of LBP gene polymorphism, APACHE II score and procalcitonin on acute respiratory distress syndrome, in order to get an ideal ARDS early warning model.
Research contents and methods:
1. experimental design: This study is divided into the following two parts: (1) in 222 cases of patients with endotoxemia as the research object, through the snapshot method for detection of LBP15 gene polymorphism SNP loci, according to whether patients developed ARDS, LBP associated with ARDS susceptibility loci and SNP genotype (2) in. 78 cases of pulmonary infection patients as the research object, according to whether patients developed ARDS, receiveroperating characteristic curve of APACHE score, procalcitonin in patients with pulmonary infection for the development of the predictive value of ARDS.
2. peripheral blood treatment and DNA extraction: 5ml EDTA anticoagulation venous blood was collected in patients ICU24 hours, plasma was separated and endotoxin test was carried out. DNA was extracted and stored in the refrigerator at -80 degrees.
The 3. snapshot method was used to detect the 15 SNP loci information of the patient's LBP gene.
The baseline clinical data of 4. patients were recorded, including APACHE score, age, gender, source of patients, chronic complications, admission diagnosis and other basic demographic data and blood samples were collected in bacterial culture also recorded. Results of the arterial blood gas of patients with pulmonary infection, blood sedimentation, C- reactive protein, lactate and procalcitonin and within 1 weeks the results of sputum culture.
5. statistical methods: allele and genotype frequencies of each SNP was calculated according to the results of gene detection, test the genotype distribution by HWE (Hardy Weinberg Equilibrium, the Hardy Weinberg equilibrium test), P0.05 Hardy - Weinberg said in accordance with building balance law.LD map by Haploview software and.Rs2232618 analysis the susceptibility of ARDS obtained by chi square test. The age, gender, effects of endotoxin concentration of such confounding factors by logistic regression method to eliminate. Normal distribution data was compared using the t test or analysis of variance, compared with non normal distribution data between groups using the Wilcoxon rank sum test and prediction function. The receiver operating characteristic curve (ROC curve) method, combined with two yuan Logistic two prediction index regression method.
The results of the study:
1. gene warning experiment, included a total of 222 cases of patients with endotoxemia, including 154 cases of male, female 58 cases, age 61 + 17.3 (17 ~ 93) at the age of.7 days for the development of the 49 cases of ARDS patients, the incidence rate of ARDS was 66 22.1%. leather cases, gram-negative bacteria infection accounted for 29.7%; and other pathogens 46 cases of mixed infection, accounting for 21.1%; has not been significantly pathogens in 110 cases, accounting for 49.2%.
Similar to 2.222 cases of 15 SNP loci in endotoxemia patients with MAF and HapMap database. The distribution of 13 SNP loci genotype with Hardy - Weinberg equilibrium (P0.05), suggesting that the genetic allele and genotype frequencies in the population is constant.APACHEII score no difference in genotype rs2232618 of different sites in (P0.05).
Only about 3. rs2232618 and the susceptibility to ARDS. Carrying the C allele (CC/CT genotype) in patients with ARDS was significantly higher than the probability of not carrying the C allele (TT genotype) patients (P=0.001). The logistic regression analysis showed that gender, age are excluded, effects of endotoxin concentration etc. and susceptibility to rs2232618 and the ARDS are still closely related (P=0.002).
4. clinical data and biomarkers of early warning experiment, included a total of 78 patients with pulmonary infection, including 60 cases of male patients, 18 female patients, age 65.2 + 17.6 (25 ~ 93) at the age of.7 days for the development of ARDS in 33 patients, the incidence of ARDS in group 42.3%.ARDS, APACHE score and calcitonin the original is significantly higher than that in group ARDS, respectively [(19.9 + 8.3) vs (15.6 + 6), and P=0.013] (12.6 + 11.4) ng/L vs (7.5 + 10.5) ng/L, P=0.046], oxygenation index lower than non ARDS group, for [(109.9 + 82.6) vs (252.1 + 92.4), P=0.003].
5., APACHE II score and procalcitonin played a similar role in predicting ARDS induced by pulmonary infection (AUROC:0.651vs0.657, P=0.929), and the prediction effect of ARDS on them was not significantly higher than that of either group.
Conclusion:
1. LBP rs2232618 gene carrying allele C (genotype CC+TC) mutations were more susceptible to endotoxin induced ARDS. T, the site of C may be an ideal index to predict early warning of endotoxin induced ARDS.
2., the APACHE II score and the expression of procalcitonin in serum are closely related to the incidence of ARDS in patients with pulmonary infection. Both of them have certain predictive value for ARDS induced by pulmonary infection, but the predictive value of combination of them is not significantly improved.

【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R563.8

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