HMGB-1及其基因多态性与严重胸部钝性伤患者预后关联性的临床研究

发布时间：2018-04-24 05:22

  本文选题：胸部创伤 + 高迁移率族蛋白-1　； 参考：《重庆医科大学》2013年硕士论文

【摘要】：研究背景 创伤已成为全球性健康问题，是45岁以下人群死亡的主要原因，而胸部创伤是严重威胁生命的急症，约占创伤相关入院率的30%-40%，占创伤相关死亡的25%。机体在遭受严重创伤后，常诱发全身炎症反应综合征（systemic inflammatory response syndrome, SIRS），并继发脓毒症（Sepsis）和多器官功能障碍综合征（multiple organ dysfunctionsyndrome, MODS），是严重创伤患者住院死亡的主要原因。胸部创伤并发Sepsis和MODS时，病情复杂、变化快，救治难度大，如果不及时有效的进行处理，可造成严重后果。虽然目前Sepsis和MODS的治疗方法众多，但是治疗水平尚未达到理想目标，其原因主要是现阶段的Sepsis和MODS的治疗都是在创伤并发症发生以后。由于缺乏早期判别创伤后严重并发症风险性的预警诊断方法，因而难以在包括Sepsis和MODS以内的严重并发症发生前或发生即刻进行预防和早期治疗。如何早期识别和干预创伤后并发症，以提高救治预后是临床一直以来期待解决的医学课题。 机体在遭受创伤时，组织细胞会主动分泌或被动释放损伤相关分子模式（damage-associated molecular patterns, DAMPs），引起炎症反应，促进坏死组织和细胞碎片的清除及损伤修复。DAMPs主要包括高迁移率族蛋白1(High-mobility group box1, HMGB1)、S100蛋白家族、热休克蛋白等。HMGB1是HMGB家族中唯一可分泌到细胞外并具有细胞因子活性的蛋白，多项研究已证实HMGB1是一种重要的晚期炎症介质，可从细胞内释放至体内调控炎症反应，在感染和损伤引起的炎症性疾病中具有重要的作用。 现已研究表明，基因组序列上的变异，即基因多态性（genepolymorphism）是决定人体对疾病易感性与抵抗性、疾病临床表型多样性以及人体对药物治疗反应差异性的重要因素。因此，，个体基因表达上的差异，很可能是部分患者易发生创伤后严重并发症或/和预后较差的重要因素之一。基于以上事实，对于严重胸部钝性伤临床进程、创伤后炎症反应程度及严重并发症发生风险等创伤后预后的相关性研究可以从基因多态性的角度去分析。 第一部分HMGB1在严重胸部钝性伤患者中的动态变化和临床意义目的： HMGB1是一种重要的晚期炎症介质，在创伤后的炎症反应中发挥着重要的调控作用。本研究为了探讨HMGB1和TNF-α在严重创伤患者血液中的动态变化及临床意义，并评估血浆HMGB1在严重胸部钝性伤患者伤后并发症发生风险及临床预后中的作用。 材料与方法： 1.样本：本研究样本来自于重庆医科大学附属第一医院胸外科和重庆市急救医疗中心收集的创伤患者血液样本。患者纳入标准为：①创伤后24小时内；②I SS评分≥16分；③年龄≥18岁且≤65岁，性别不限；④存活时间超过1周。 2.创伤后的第1、3、5、7d各取外周静脉血，冷冻保存，标本收齐后同时应用ELISA法检测HMGB1和TNF-α水平的测定。 3.分析HMGB1和TNF-α动态变化与创伤严重程度、伤情特点及致伤特征、近远期预后的关系。 4.比较胸部创伤患者伤后不同时间点血清HMGB1和TNF-α对预后判断的ROC曲线结果。 结果： 1.严重胸部钝性伤患者血浆HMGB1水平在伤后第1天、第3天逐渐升高，第5天达高峰，第7天仍呈持续上升趋势。而TNF-а水平在创伤后第1天至第7天呈逐渐降低的趋势。 2.严重胸钝性伤患者伤后第1、3、5、7天血浆HMGB1与ISS评分呈正相关，其中第3天、第5天及第7天相关性具有统计学差异（P均0.01）；TNF-а水平与ISS评分也呈正相关（P均0.05）。 3.严重胸钝性伤患者伤后第3、5、7天血浆HMGB-1与伤后感染、Sepsis、MODS、创伤后应激障碍及近期生活质量（SF-36）相关（P均0.05）。 4.两者在创伤后并发症预测方面，伤后第1天，HMGB1和TNF-а水平的ROC曲线下面积差异无统计学意义（P0.05）；而在第3、5、7天血浆HMGB1水平对预测伤后并发症的ROC曲线下面积均大于TNF-а的ROC曲线下面积（P均0.05）。 结论： 我们的研究发现严重胸部钝性伤患者血浆HMGB1水平在伤后进行性增高，且与创伤的严重程度相关，相关性强于TNF-а，其预测并发症发生情况，敏感性要优于血浆中TNF-а的变化，尤其是在创伤后晚期；创伤后血浆HMGB1水平升高提示患者预后不佳，心理健康及生存质量较差。 第二部分HMGB1基因标签单核苷酸多态性与创伤并发症风险性的关联研究 目的： 本实验旨在探讨HMGB1基因标签单核苷酸多态性与严重胸部钝性伤后并发Sepsis和MODS风险性的关联性。 材料和方法： 一、主要信息资源及分析工具： 1.主要查询数据库：NCBI数据库：www.ncbi.nlm.nih.gov；Hapmap数据库：www.hapmap.org。 2.分析软件：Haploview(4.0版)； 二、实验方法： 1.标签单核苷酸多态性的筛选。HMGB1基因的转录起始位点上游3-10kb至其终止子下游3-10kb范围及基因所有内含子和外显子范围内进行标签单核苷酸多态性的筛选，用Haploview软件构建基因的单倍域，利用Tagger软件在单倍域内进行标签单核苷酸多态性的筛选。 2.基因分型。采用焦磷酸测序法对HMGB1基因进行标签SNPs的分型。按照仪器说明进行分型反应，根据每个碱基的峰高进行结果的自动判读。 3.临床关联研究，Sepsis和MODS诊断根据诊断标准，创伤ISS评分采用简明损伤定级标准（2005版）。 4.统计分析：各SNPs的等位基因型频率由基因计数获得。H-W平衡采用卡方检验计算；多态性与MODS评分的关系采用单因素方差方法统计；等位计量效应采用多元Logister回归分析统计，并用年龄、性别及ISS评分校正；各基因型与Sepsis、MODS发生率之间的关系采用卡方检验方法统计。 结果： 1.人HMGB1基因位于13号染色体29930877至29938081之间，长度为7.204kb。HMGB1基因共有7个SNPs，都是常见SNP，构建了2个单倍域。HMGB1挑选了3个标签SNPs，分别是rs1412125/rs2249825/rs1045411。 2.在HMGB1的3个标签SNPs中，只有rs2249825的C→G突变与创伤后Sepsis发生和MODS评分密切相关，而另外2个标签SNPs不影响患者创伤后并发症的发生率。 结论： HMGB1基因的rs2249825在中国汉族人群中与创伤后脓毒症和MODS发生密切的多态性位点。HMGB1基因rs2249825的C→G突变可能是增加创伤后Sepsis和MODS发生风险性的遗传因素。
[Abstract]:Research background
Trauma has become a global health problem, the main cause of death of people under 45 years of age, and chest trauma is a serious threat to life, accounting for 30%-40% of trauma related admission rate, and the 25%. body of trauma related death often induces systemic inflammatory response syndrome (systemic inflammatory response syndrome). SIRS), secondary sepsis (Sepsis) and multiple organ dysfunction syndrome (multiple organ dysfunctionsyndrome, MODS) are the main causes of death in patients with severe trauma. When thoracic trauma is complicated with Sepsis and MODS, the disease is complicated, fast and difficult to treat. If not treated in time and effectively, it can cause serious consequences. At present, there are many methods of treatment for Sepsis and MODS, but the level of treatment has not reached the ideal goal. The main reason is that the treatment of Sepsis and MODS at the present stage is after the occurrence of traumatic complications. Due to the lack of early warning diagnosis of the risk of severe complications after the early discrimination, it is difficult to include the strictness within Sepsis and MODS. Prevention and early treatment immediately before or before the occurrence of heavy complications. How to identify and intervene early posttraumatic complications and improve the prognosis of treatment is a medical subject that has been expected to be solved in clinical.
When the body is traumatized, tissue cells release or passively release the damage-associated molecular patterns (DAMPs), causing inflammatory reactions and promoting the removal of necrotic tissue and cell debris and the repair of.DAMPs mainly including high mobility group protein 1 (High-mobility group box1, HMGB1), S100 protein .HMGB1 is the only protein in the family, such as heat shock protein, which can be secreted into the HMGB family and has the activity of cytokines. A number of studies have confirmed that HMGB1 is an important late inflammatory mediator. It can be released from the cell to the body to regulate the inflammatory response and plays an important role in the inflammatory diseases caused by infection and injury.
It has been shown that the variation in the genome sequence, that is, gene polymorphism (genepolymorphism), is an important factor in determining the susceptibility and resistance of the human body to disease, the diversity of the clinical phenotype and the difference in the response of the human body to the drug treatment. One of the important factors for severe complications or / and poor prognosis. Based on the above facts, the correlation of post-traumatic prognosis in the clinical process of severe blunt trauma, the degree of post traumatic inflammation and the risk of severe complications can be analyzed from the perspective of genetic polymorphism.
Part one: dynamic changes and clinical significance of HMGB1 in patients with severe blunt chest trauma.
HMGB1 is an important late inflammatory mediator and plays an important role in the post traumatic inflammatory response. This study aims to explore the dynamic changes and clinical significance of HMGB1 and TNF- alpha in the blood of patients with severe trauma, and to evaluate the risk and clinical prognosis of plasma HMGB1 in patients with severe blunt chest injury. Effect.
Materials and methods:
1. sample: the samples were collected from the blood samples collected from the Department of thoracic surgery in First Affiliated Hospital of Chongqing Medical University and the Chongqing emergency medical center. The patients were included in the criteria: (1) 24 hours after the trauma; (2) I SS score was more than 16; (3) the age of 18 years old and less than 65 years, and the sex was not limited; (4) the survival time was over 1 weeks.
2. after the trauma, peripheral venous blood was collected from each 1,3,5,7d. After cryopreservation, the levels of HMGB1 and TNF- TNF- were detected by ELISA.
3. to analyze the relationship between dynamic changes of HMGB1 and TNF- alpha and severity of trauma, characteristics of injury and characteristics of injury, and prognosis.
4. to compare the ROC curves of serum HMGB1 and TNF- alpha at different time points after trauma in patients with thoracic trauma.
Result锛

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