深低温停循环后再灌注肺损伤的临床和基础研究

发布时间：2018-01-13 20:29

本文关键词：深低温停循环后再灌注肺损伤的临床和基础研究　出处：《福建医科大学》2016年博士论文　论文类型：学位论文

【摘要】：目的评估急性呼吸窘迫综合征(acute respiratory distress syndrome,ARDS)对Stanford A型主动脉夹层(aorta dissection,AD)手术患者预后的影响,寻找Stanford A型AD术后发生ARDS的危险因素,为相关疾病的临床防治提供参考依据。材料与方法选取329例于2013年1月至2015年7月在我科接受常规开胸手术的Stanford A型AD患者作为研究对象,以回顾性巢式病例对照作为研究手段,对其中的126例患者(非ARDS组和ARDS组各63例)一般情况、病因、术前并发症、术式、术中情况、围术期输血情况、血清TNF-α水平变化进行统计学分析。在初步筛选Stanford A型AD术后发生ARDS的可疑危险因素后,通过多元logistics回归分析找出独立危险因素。通过比较患者死亡率差别、术后并发症和预后、住院总费用、绘制生存曲线等方法评估ARDS对Stanford A型AD手术患者预后的影响。结果Stanford A型AD术后ARDS总的发生率20.7%。两组间年龄、性别等一般临床资料无显著性差别;主动脉弓部处理方式、手术操作涉及的范围亦无显著性差异。ARDS组患者的CPB时间和DHCA时间显著长于非ARDS组,而主动脉阻断时间两组间无显著性差异。相比非ARDS组,ARDS组患者围术期输注了更多的红细胞、血小板及新鲜冰冻血浆。肺部感染发生率、MODS发生率及死亡率显著增加,入住ICU时间、呼吸机辅助时间显著延长,及住院总费用显著增加。两组围术期的OI值和ApacheⅡ分值变化有显著性差异,ARDS组术后OI值总体水平低于非ARDS组,ApacheⅡ分值总体水平高于非ARDS组。ARDS组患者的血清TNF-α作用强度水平及变化趋势显著高于非ARDS组。多元logistic回归分析提示DHCA时间、血清TNF-α作用强度和围术期输血量(不论输注类型)是Stanford A型AD术后ARDS的独立危险因素。生存曲线分析显示ARDS组患者术后的总体存活率明显低于非ARDS组。结论Stanford A型AD术后ARDS严重影响Stanford A型AD患者的预后,其发生与DHCA时间、围术期输血及血清TNF-α水平密切相关。通过改进手术技术,减少DHCA时间和围术期输血,降低术后血清TNF水平可能成为改善Stanford A型AD术后肺功能的有效手段。目的寻找建立深低温停循环(deep hypothermic circulatoryarrest,DHCA)下大鼠肺缺血再灌注损伤(ischemia-reperfusion injury,IRI)在体模型的有效方法,为动物体内干预试验提供实验基础。材料与方法以体重相近的健康成年雄性SD大鼠为实验对象,分成缺血再灌注组(I/R组,n=15)和假手术组(S组,n=15),以气管插管全麻和直视开胸手术作为操作方法,以恒温水浴箱为控温设备,制作深低温停循环下的大鼠左下肺叶缺血再灌注的在体模型。通过观察和比较两组间的血气分析结果、气道阻力、湿/干重比率、BALF细胞数和成分、组织切片下的病理学改变,来评估I/R后ARDS。结果本实验共消耗大鼠32只,成功建立大鼠在体I/R肺模型15例,匹配假手术组15例,1例死与低氧血症,来自实验组;1例术后未观察到典型的ARDS表现,建模失败。I/R建模成功率88.2%(15/17),术后12h存活率100%。麻醉及术前准备耗时25.5±6.4min,手术过程耗时41.2±9.2min,复苏过程耗时23.6±3.2min,总手术时间耗时96.5±12.3min。结论该模型停循环范围局限于左下肺叶,对大鼠的呼吸功能损伤较小,死亡率低,手术操作简便,单人肉眼即可完成。模型可复制性好,成功率高,易于批量建模,为进行后续的大规模干预实验研究提供了坚实的基础。麻醉药量控制、气管插管、颈动脉插管和停循环时间的把握是建模成功的关键。目的以大鼠深低温停循环下肺缺血再灌注损伤模型为基础,探讨TNF-α对Stanford A型AD术后ARDS的影响及机制,为Stanford A型AD术后ARDS寻找新的临床治疗靶点。材料与方法以72只雄性SD大鼠为研究对象,均分为假手术组(S组)、对照组(C组)和实验组(Ab组),每组再按肺组织标本留取时间均分为术后4h亚组、术后12h亚组和术后24h亚组。实验模拟主动脉夹层手术中的深低温停循环过程,造成肺组织缺血再灌注损伤,实验组术前常规应用抗TNF-α单克隆抗体拮抗TNF-α的作用。术后通过肉眼和HE染色镜检观察肺组织大体和微观结构改变;血气分析测定动脉血Pa O2和Pa CO2评估大鼠围术期肺氧合功能和肺泡通气功能;称重法检测肺组织干湿重比率评估肺水肿程度;呼吸机参数读取气道峰压评估围术期气道阻力变化;TUNEL法检测肺组织细胞凋亡指数;ELISA法检测术后血清TNF-α、IL-6/10水平和肺组织MPO、MDA、SOD含量;RT-PCR法检测肺组织术后caspase3/8和NF-κB表达水平;生存曲线分析评估大鼠存活率差异。结果C组大鼠肺组织水肿、气道分泌物增加,镜检见肺间质水肿、肺泡结构破坏、炎症细胞和红细胞浸润。S组未发现上述病理改变,Ab组病变程度居中;与S组相比,C组和Ab组术后动脉血Pa O2明显降低,Pa CO2明显升高;术后肺组织干湿重比率、气道阻力、肺组织凋亡指数、肺组织TNF-α表达强度、血清TNF-α、IL-6水平、肺组织MPO、MDA含量、肺组织TNF-α、caspase3/8和NF-κB表达水平均明显增加,C组增加程度显著高于Ab组;与S组相比,C组和Ab组术后血清IL-10水平明显增高,Ab组增高更明显;三组大鼠术后肺组织SOD含量变化无显著性差异。共计85只SD大鼠进行了上述实验,死亡13只(除外麻醉意外和大出血所致的死亡),其中S组2只,C组7只,Ab组4只,总死亡率15.3%,三组大鼠术后24h内的存活率有显著性差异。结论TNF-α可通过诱导肺水肿损害肺泡氧合和通气功能、募集和激活中性粒细胞、促进炎症反应和氧化应激、诱导肺泡上皮细胞凋亡等机制促进深低温体循环术后肺损伤的发生。通过术前预防性的使用TNF-α拮抗剂可在一定程度上减轻上述机制造成的肺损伤。TNF-α有可能成为防治Stanford A型AD术后ARDS的新靶点。
[Abstract]:Objective to evaluate the acute respiratory distress syndrome (acute respiratory distress syndrome, ARDS) of Stanford type A aortic dissection (aorta dissection AD) affect the prognosis of patients, the risk factors for ARDS Stanford A after AD, to provide reference for clinical prevention and treatment of related diseases. Materials and methods: 329 cases in January 2013 to July 2015 in our department to accept the Stanford A type AD patients with conventional thoracic surgery as the research object, in a retrospective nested case-control study as a means of one of the 126 patients (non ARDS group and ARDS group with 63 cases in each group) general condition, etiology, preoperative complications, operation, intraoperative, peri perioperative blood transfusion, changes of serum levels of TNF- were analyzed. The suspicious risk factors of ARDS in the preliminary screening of Stanford A type AD postoperatively, by multivariate logistics regression analysis to identify independent risk factors By comparing the difference. The mortality of patients, postoperative complications and prognosis, the total hospitalization expenses, draw survival curves were used to analyze effects of ARDS on the prognosis of patients with Stanford type A AD surgery. Results the incidence rate of ARDS 20.7%. between the two groups of age total Stanford type A after AD, there is no significant difference between gender and other general clinical data; aortic arch approach, scope of operation involved and there were no significant difference in.ARDS group were CPB time and DHCA time was significantly longer than non ARDS group, and aortic clamping time had no significant difference between two groups. Compared with non ARDS group, infusion of more red blood cells of ARDS patients during the perioperative period, platelet and fresh frozen plasma. The incidence of pulmonary infection, the incidence of MODS and mortality increased significantly in the ICU time, ventilation time was significantly prolonged, and the total cost of hospitalization was significantly increased. The changes of Apache and II scores of two groups of perioperative OI value There was significant difference in OI value, the overall level of ARDS group were lower than non ARDS group, Apache II score overall level higher than the non ARDS group.ARDS serum TNF- alpha intensity levels and trends were significantly higher than non ARDS group. Multivariate logistic regression analysis showed that the DHCA time, serum TNF- intensity and perioperative blood transfusion the amount (whether infusion type) is the independent risk factors of ARDS Stanford A AD after operation. The survival curve analysis showed that the patients of the ARDS group after the overall survival rate was significantly lower than that in non ARDS group. Severe ARDS influence the prognosis of patients with Stanford type A AD type A AD conclusion Stanford after operation, the occurrence and duration of DHCA, Wai blood transfusion and serum levels of TNF- patients are closely related. Through the improvement of surgical technique, reduce the DHCA time and perioperative blood transfusion, reduce the postoperative serum TNF level may be an effective means of improving pulmonary function of Stanford A after AD. To build Vertical deep hypothermic circulatory arrest (deep hypothermic, circulatoryarrest, DHCA) lung ischemia reperfusion injury in rats (ischemia-reperfusion injury, IRI) in the method body model, to provide the experimental basis for animal test in vivo intervention. Materials and methods with similar weight healthy adult male SD rats were divided into ischemia reperfusion group (group I/R, n=15) and sham operation group (group S, n=15), with tracheal intubation anesthesia and open thoracotomy as the operation method, with constant temperature water bath box for temperature control equipment, production of deep hypothermic circulatory arrest in rats with left lower lobe ischemia / reperfusion in vivo. Through the observation and comparison of two blood gas between group analysis, airway resistance, wet / dry weight ratio, the number and composition of BALF cells, the pathological changes of the tissue sections, to assess the I/R ARDS. after the results of this experiment consumed a total of 32 rats were successfully established in vivo in rat lung model 15 I/R Cases, the sham operation group 15 cases, 1 cases died of hypoxemia, from the experimental group; 1 cases were not observed in the typical ARDS findings,.I/R modeling modeling failed success rate of 88.2% (15/17), the postoperative survival rate of 12h 100%. anesthesia and preoperative preparation time was 25.5 + 6.4min, 41.2 + operation time 9.2min, the recovery process takes 23.6 + 3.2min, the total operation time consuming 96.5 + 12.3min. conclusion the model of circulatory arrest is confined to the left lower lobe, respiratory function injury in rats is small, the mortality rate is low, the operation is simple, single eye can be completed. Model can be replicated, high success rate, easy batch modeling, provide a solid foundation for the study of large-scale intervention in subsequent experiments. The amount of anesthetic drug control, tracheal intubation, carotid artery intubation and circulatory arrest time is the key to successful modeling. With the purpose of deep hypothermic injury of rat model of lung ischemia-reperfusion cycles Based on the mechanism of ARDS on TNF- alpha and Stanford type A after AD, to find a new target for clinical treatment of Stanford type A AD ARDS after the operation. Materials and methods: 72 male SD rats as the research object, divided into sham operation group (S group) and control group (C group) and experimental group (Ab group), each group according to the lung tissue specimens divided into postoperative 4H subgroups, postoperative 12h subgroup and 24h subgroup. After experimental simulation of deep hypothermic circulatory arrest in aortic dissection, causing pulmonary ischemia reperfusion injury in experimental group, the preoperative routine the application of anti TNF- alpha monoclonal antibody against TNF- alpha. After operation by changing the gross and microscopic observation of HE staining of lung tissue specimens and microstructure; blood gas analysis of arterial blood Pa O2 and Pa CO2 in rats were evaluated perioperative pulmonary oxygenation and pulmonary ventilation function; weighing method for detection of lung tissue wet / dry weight ratio the evaluation process of pulmonary edema The degree of ventilator parameters; read peak airway pressure to evaluate peri operative changes of airway resistance; apoptosis of the lung cells was detected by TUNEL ELISA method after surgery; detection of serum TNF- level and lung tissue IL-6/10, MPO, MDA, SOD content; RT-PCR lung tissues were detected after caspase3/8 and NF- K B expression; survival curve analysis and evaluation of the survival rate of rats. The differences in lung tissue of rats in C group were edema, airway secretions increase, the histological examination showed pulmonary interstitial edema, alveolar structural damage, inflammatory cells and red blood cell infiltration.S group found no pathologic changes in the Ab group, the severity of the middle; compared with S group, C group and Ab group. After arterial blood Pa O2 were decreased, Pa CO2 increased significantly; postoperative lung wet / dry weight ratio, airway resistance, apoptosis index of lung tissue, lung tissue expression of TNF- alpha, alpha IL-6 levels, serum TNF-, lung MPO, MDA content in lung tissue of TNF- alpha, caspase3/8 and NF- K B expression The C group was significantly increased, increased significantly higher than group Ab; compared with S group, serum IL-10 levels were significantly increased in C group and Ab group after operation, Ab group was more obvious; there was no significant difference in SOD content in lung tissue of rats in three groups after operation. A total of 85 SD rats of the experiment. Only 13 deaths (except the anesthetic accident and death caused by bleeding), 2 rats in S group, 7 rats in group C, 4 rats in group Ab, the total mortality rate was 15.3%, three groups of rats at the survival rate of 24h had significant difference. Conclusion TNF- can be induced by alpha damage of oxygenation and pulmonary edema ventilation function, recruitment and activation of neutrophils, promote inflammation and oxidative stress induced apoptosis of alveolar epithelial cells and other mechanisms to promote the circulation of deep hypothermia after lung injury. The preoperative prophylactic use of TNF- antagonists in some extent caused by reducing the mechanism of lung injury is likely to become the alpha.TNF- A new target for the prevention and treatment of ARDS after Stanford A AD.

【学位授予单位】：福建医科大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R654.2

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