自噬在机械通气所致大鼠呼吸机相关性肺损伤中的作用及意义

发布时间：2018-07-01 20:00

本文选题：呼吸机相关肺损伤 + 大鼠　；参考：《郑州大学》2016年博士论文

【摘要】：目的建立大鼠机械通气所致相关性肺损伤的动物模型,研究不同潮气量机械通气对大鼠肺组织的影响情况。方法健康雄性SPF级SD大鼠30只,周龄7周,体重(220±20g),实施麻醉和气管切开术后分别接受不同潮气量(VT)的通气(通气时间均为4h)。随机分为以下3组:对照组(A组,仅行气管插管并保留自主呼吸);常规通气组(B组,VT=10ml/kg);损伤通气组(C组,VT=40ml/kg)。A组在气管插管后即刻,其他组在机械通气结束后放血处死动物,计算肺湿/干比重比值(W/D比值),HE染色观察肺组织病理学变化。结果在大潮气量机械通气4h后,光镜下病理学检查结果显示A、B组病理形态大致正常,C组肺组织中可见肺泡间隔增宽,肺泡腔渗液明显增加,其内可见炎性细胞浸润并有大量散在出血点;与A、B相比,C组肺组织W/D比值明显增加(P0.05)。结论大潮气量机械通气能够诱导大鼠呼吸机相关性肺损伤的发生,通过肺组织湿干比重和肺组织病理学检查提示复制大鼠呼吸机相关性肺损伤动物模型成功。目的评价机械通气相关性所致肺损伤(VILI)过程中自噬相关蛋白Beclin1、LC3和炎症相关因子NF-κB表达的意义。方法健康雄性SPF级SD大鼠30只,周龄7周,体重(200-250 g),实施麻醉和气管插管术后分别接受不同潮气量(VT)的通气(通气时间均为4 h)。随机分为以下3组:对照组(A组,仅行气管插管并保留自主呼吸);常规通气组(B组,VT=10 ml/Kg);损伤通气组(C组,VT=40 ml/Kg)。A组在气管插管并自主呼吸4h后,其他组在机械通气结束后放血处死动物,计算BALF中白细胞计数(WBC)和总蛋白含量,ELISA试剂盒检测BALF中炎症因子TNF-α和IL-1β的含量,使用MPO、SOD及MDA试剂盒检测肺组织匀浆中相应指标的表达水平,Western Blot法检测肺组织中自噬相关基因Beclin1、LC3及NF-κB的表达水平。结果在大潮气量机械通气4h后,肺组织中总蛋白含量、白细胞计数和MPO、MDA等指标均显著增加(P0.05),SOD含量明显降低(P0.05);肺组织中Beclin1、LC3、NF-κB表达水平均显著增加(P0.05);而这一作用能够被自噬特异性抑制剂3-MA所抑制。结论在大潮气量通气所致的大鼠呼吸机相关性肺损伤过程中,自噬对于VILI过程中的炎症反应起着十分重要的作用,其机制可能与NF-κB信号通路有关,这一作用可以被自噬特异性抑制剂3-MA所抑制,说明在VILI导致炎症形成的过程中自噬可能是一种保护机制。
[Abstract]:Objective to establish an animal model of lung injury induced by mechanical ventilation in rats and to study the effects of different tidal volume mechanical ventilation on lung tissue in rats. Methods Thirty healthy male SPF SD rats, aged 7 weeks and weighing (220 卤20 g), were ventilated with different tidal volume (VT) after anesthesia and tracheotomy respectively (ventilation time was 4 h). They were randomly divided into the following three groups: control group (group A, with only tracheal intubation and spontaneous respiration), routine ventilation group (group B) (VT 10 ml / kg), injury ventilation group (group C, VT 40 ml / kg). Group A, immediately after tracheal intubation, and other groups, were bled and killed after mechanical ventilation. Lung wet / dry specific gravity ratio (W / D ratio) was calculated to observe the histopathological changes of lung by HE staining. Results after 4 hours of mechanical ventilation, pathological examination under light microscope showed that alveolar septum widened and alveolar effusion increased obviously in group C with normal pathological morphology. The infiltration of inflammatory cells and a large number of scattered bleeding spots were observed in the lung tissue. Compared with AMB group, the ratio of W / D in lung tissue in group C was significantly increased (P0.05). Conclusion Mechanical ventilation can induce ventilator-associated lung injury in rats. The wet and dry specific gravity of lung tissue and pathological examination of lung tissue indicate that the rat model of ventilator associated lung injury is successful. Objective to evaluate the significance of the expression of autophagy associated protein Beclin1 (LC3) and inflammatory related factor (NF- 魏 B) in ventilation-induced lung injury (VILI). Methods 30 healthy male SPF SD rats were given ventilation with different tidal volume (VT) after anesthesia and tracheal intubation for 7 weeks (200-250 g),) (ventilation time was 4 h). They were randomly divided into the following three groups: control group (group A, only tracheal intubation with spontaneous respiration), routine ventilation group (group B, 10 ml / kg), injury ventilation group (group C, 40 ml / kg), group A, 4 h after tracheal intubation and spontaneous respiration. In other groups, the animals were bled out after mechanical ventilation. WBC and total protein contents in BALF were calculated. The levels of inflammatory factors TNF- 伪 and IL-1 尾 in BALF were detected by Elisa kit. The expression level of corresponding indexes in lung homogenate was detected by MPOO SOD and MDA kit. Western blot was used to detect the expression of autophagy related gene Beclin1, LC3 and NF- 魏 B in lung tissue. Results after 4 hours of mechanical ventilation, the total protein content in lung tissue was determined. WBC count and MPOO MDA were significantly increased (P0.05) and the expression level of NF- 魏 B in Beclin1 + LC3 was significantly increased (P0.05), which could be inhibited by autophagy specific inhibitor 3-MA. Conclusion autophagy plays an important role in the inflammatory response of Vili in the process of ventilator associated lung injury induced by spring tide ventilation in rats. The mechanism of autophagy may be related to NF- 魏 B signaling pathway. This effect can be inhibited by 3-MA, a specific inhibitor of autophagy, suggesting that autophagy may be a protective mechanism in the process of inflammation induced by VILI.
【学位授予单位】：郑州大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R563.8

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