GM-CSF对严重烫伤大鼠急性肺损伤影响的研究

发布时间：2018-03-21 17:02

  本文选题：烧伤　切入点：肺水肿　出处：《蚌埠医学院》2017年硕士论文　论文类型：学位论文

【摘要】：研究背景及目的在严重烧伤后早期,微血管扩张,血管通透性增加,大量水分及营养物质丢失,炎症介质入侵,严重可引起休克及多脏器损伤,而肺脏是最常损伤的器官。烧伤后引起急性肺损伤(ALI)可能机制如下:1.中性粒细胞作为ALI发病过程中最为重要的细胞,通过释放活性氧、多种细胞因子及蛋白水解酶引起肺损伤;2.氧化与抗氧化反应的严重失衡可能是ALI发生的机制;3.ALI患者凝血因子发生异常,继而导致凝血与抗凝机制作用异常,造成肺泡内纤维蛋白沉积;4.肺毛细血管内皮细胞的损伤是ALI发病过程中的中心环节,肺微血管通透性增加是早期肺损伤的重要原因。粒细胞巨噬细胞集落刺激因子(Granulocyte macrophage colony-stimulating factor,GM-CSF)是一种具有多种功能的细胞因子,其与GM-CSF受体(GMR)结合促进多种细胞的增殖和分化,保护并提高多类细胞的功能,包括内皮细胞、成纤维细胞、中性粒细胞/单核细胞、树突状细胞及角质形成细胞等。在过去,GM-CSF主要应用于放化疗及骨髓移植后的白细胞减少症、再生障碍性贫血等免疫缺陷性疾病的治疗,发挥抗肿瘤及抗感染作用;现在GM-CSF还可以应用到各类创面的修复,通过加速伤口上皮化、参与新生血管的形成及伤口的收缩以促进伤口愈合;近年来,我们的研究还发现GM-CSF可以调节血管周细胞增加Ang-1的表达,降低VEGF的表达,从而调节微血管通透性,促进伤口愈合。综合以上研究基础,我们设想GM-CSF可能对严重烧伤后的急性肺损伤具有保护作用。本课题在严重烧伤急性肺损伤模型的基础上,腹腔注射GM-CSF,观察GM-CSF对烧伤后急性肺损伤治疗效果,并分析其可能的机理。方法SD大鼠30只随机分为A、B、C三组,A组为假烫组,其余制模成30%Ⅲ度烧伤,B为立即复苏用药组,C为立即复苏不用药组,(复苏液为乳酸林格氏液,按照Parkland公式;GMCSF用药方法60ug/kg,腹腔内注射)各组大鼠均于伤后48h后取外周血,用ELIAS方法检测MPO、TNFa、IL-6,VEGF、ANG-1;取肺组织检测肺组织湿干比并做病理检测,用免疫荧光法,检测肺组织MPO、IL-6、VEGF及ANG-1表达情况。上述结果均用t检验做统计学分析。结果1.大鼠烧伤48h后,各组肺组织湿干比均较A组(4.3502±0.0782)明显增加。其中B组湿干比(4.5834±0.1030)低于C组(4.8509±0.1896),比较差异有显著统计学意义(P0.01)。2.烧伤48h后检测各组肺组织MPO浓度含量情况:C组(144.6±6.1325pg/ml)明显高于B(97.7±4.7603)组,差异有统计学意义(P0.01);A组(68.1±5.7227)略低于B组(97.7±4.7603),差异有统计学意义(P0.05)。3.在建立模型48h后取外周血测得TNF-a、IL-6、VEGF浓度含量:C组(237±18.69pg/ml、124.8±10.8pg/ml、255.7±12.48pg/ml)较A组(135.2±13.17pg/ml、74.5±12.5pg/ml、152.3±8.86pg/ml)及B组(170.2±14.96pg/ml、88.6±8.2pg/ml、179.3±11.2pg/ml)显著升高,C组明显高于B组(P0.01),差异有统计学意义。而外周血中B组(3080±218.27pg/ml)ANG-1浓度含量较C组(2045±98.69pg/ml)显著增高,差异有统计学意义(P0.01),而A组与B组无明显差异,无统计学意义。4.烧伤48h后肺组织中检测TNF-a及IL-6的浓度含量C组较A组及B组明显增加,C组(214.8±8.24pg/ml、116.1±6.0pg/ml)明显高于B组(177.4±3.78pg/ml、82.1±5.27pg/ml),比较差异有统计学意义(P0.01)。C组肺组织中VEGF浓度含量(279.3±8.07pg/ml)明显高于及B组(177.4±3.78pg/ml),差异有统计学意义(P0.01);B组ang-1浓度含量(3106±80.2pg/ml)明显高于C组(2177±93.56pg/ml)(P0.01),差异有统计学意义。5.烧伤48h后对肺组织中的血管通透性指标进行荧光分析检测,观察到C组组中IL-6、MPO及VEGF表达含量较A组与B组明显增多,而ang-1荧光表达A组与B组明显强于C组。6.烧伤48h后C组Evans Blue OD值(0.318±0.337)明显高于B组(0.202±0.397),差异有统计学意义(P0.01)。结论1.大鼠严重烧伤后早期,肺组织水肿为早期并发症,通过腹腔注射GM-CSF用药发现可以减轻肺水肿,提示GM-CSF对严重烫伤后急性肺损伤有保护作用。2.GM-CSF通过促进血管外周细胞ANG-1的表达,降低VEGF作用于血管内皮细胞通透性,增强细胞与细胞之间的紧密性,减少水分及大分子物质的渗出,降低肺毛细血管通透性,对肺部具有保护作用。
[Abstract]:Background and objective in after severe burns, microvascular expansion, increased vascular permeability, large amounts of water and nutrient loss, inflammation invasion, serious can cause shock and multiple organ damage, and lung is the most common organ injury. Burns caused by acute lung injury (ALI) mechanism of neutrophils are as follows: 1. as the most important cell in the pathogenesis of ALI, through the release of reactive oxygen species, a variety of cytokines and proteolytic enzymes caused by lung injury; 2. serious imbalance of oxidation and antioxidation reaction may be the mechanism of ALI; 3.ALI patients with abnormal blood coagulation factor, then lead to the effect of coagulation and anticoagulation mechanism caused by abnormal alveolar fibrin deposition; 4. pulmonary capillary endothelial cell injury is the central link in the pathogenesis of ALI, increase the pulmonary microvascular permeability is an important reason for early lung injury induced by granulocyte macrophage colony. Colony stimulating factor (Granulocyte macrophage, colony-stimulating factor, GM-CSF) is a multifunctional cytokine, and GM-CSF receptor (GMR) combined with the proliferation and differentiation of a variety of cells, protect and improve many types of cells, including endothelial cells, fibroblasts, neutrophils / monocytes, dendritic cells and keratinocytes. In the past, GM-CSF is mainly used in chemotherapy and bone marrow transplantation in the treatment of leukopenia, aplastic anemia and other immunodeficiency diseases, anti-tumor and anti infection effect; now GM-CSF can also be used to repair all kinds of wounds, through accelerated wound epithelialization, formation and participate in wound neovascularization contraction to promote wound healing; in recent years, we have found that GM-CSF can regulate the perivascular cells increase the expression of Ang-1 and decreased the expression of VEGF, from The regulation of microvascular permeability, promote wound healing. Based on the above research foundation, we assume that GM-CSF may have a protective effect on acute lung injury after severe burns. Based on the model of severe burn acute lung injury, intraperitoneal injection of GM-CSF, observe the effect of GM-CSF on the effect of the treatment of acute lung injury after burn injury, and to analyze the possible the mechanism. Methods 30 SD rats were randomly divided into A, B, C three groups, A group was the sham group, the mould into 30% third degree burns, B for immediate recovery group, C for immediate recovery without medication group (resuscitation fluid for lactated Ringer's solution, according to the Parkland formula; use of GMCSF method 60ug/kg, intraperitoneal injection) in all rats after injury after 48h from peripheral blood, MPO, ELIAS was detected by TNFa, IL-6, VEGF, ANG-1; the detection of lung wet dry ratio and pathological detection by immunofluorescence, detection of lung tissue MPO, IL-6. VEGF and ANG-1 杈炬儏鍐，

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