【作者】 曾林祥；

【导师】 郭光华；

【作者基本信息】 南昌大学 ， 外科学， 2008， 博士

【摘要】 吸入性损伤的发病率及死亡率一直很高,重度烧伤合并吸入性损伤病人是烧伤临床治疗上的重点和难点,病死率仍然高达50-60%。机械通气是治疗重度烧伤合并吸入性损伤不可缺少的手段之一,但常规机械通气可能产生和加重肺损伤。作为一种“保护性肺通气策略,高频振荡通气和部分液体通气日益受到重视。本研究将新西兰大白兔制成蒸气吸入性损伤并急性呼吸衰竭模型,随机分组后采用四种机械通气模式进行治疗(CMV,CMV+PLV,HFOV及HFOV+PLV),分别在治疗后0.5,1.5,2.5及3.5h取动脉血行血气分析、血流动力学监测。通气治疗3.5h后,为评估四种机械通气模式对通气肺组织的炎症反应及肺损伤程度的影响,作了大体标本、组织病理学观察及肺组织损伤评分;使用酶联免疫吸附法(ELISA)检测肺组织匀浆中和血清中可溶性E-选择素、P-选择素、ICAM-1和ET-1的含量,用RT- PCR法测肺组织中E-选择素、P-选择素、ICAM-1和ET-1 mRNA表达。为评价四种机械通气模式对通气肺组织细胞凋亡的影响,应用TUNEL DNA片段末端标记方式(FragELTM)法检测肺组织细胞凋亡,采用RT- PCR法测肺组织匀浆中caspsase-3含量,应用逆转录聚合酶链反应(RT-PCR)技术检测肺组织caspsase-3 mRNA表达水平,用凝胶电泳条带成像系统照相分析结果。收集肺泡灌洗液细胞后分离出中性粒细胞,使用流式细胞仪Annexin V-FITC法检测中性粒细胞凋亡及坏死;肺泡灌洗上清液则进行各种炎性因子(E-选择素、P-选择素、ICAM-1和ET-1)总蛋白、丙二醛检测。结果:1.血气分析: 1)伤后各组动物PaO2均降至60mm Hg以下,各组致伤前与致伤后比较均有显著性差异(P<0.01),各组致伤前、致伤后(治疗前)血气组间比较均无显著性差异(P >0.05)。2).四个治疗组的PaO2在通气治疗0.5h后均明显上升,通气治疗3.5h内一直有良好的PaO2。3).HFOV组和HFOV+PLV组的PaO2、PaO2/FiO2在治疗治疗各时间点均分别高于相应时间点的CMV组和CMV+PLV组(P<0.05和P<0.01);CMV+PLV组在治疗1.5h、2.5h PaO2优于CMV组(P<0.05);HFOV+PLV组在治疗3.5h PaO2优于HFOV组(P<0.01)。4)肺泡气-动脉血氧分压差[P(A-a)O2]在各治疗组无统计学差异(P >0.05)。5)四个治疗组伤后各时相点的PaCO2均无明显变化(P >0.05)。2.肺组织病理学检查:HFOV组的损伤程度较CMV组明显减轻,HFOV+PLV组损伤最轻,CMV+PLV组次之,CMV组损伤最重。3.炎症反应:E-选择素、P-选择素和ICAM-1在肺组织和血清中的含量及其在肺组织中mRNA表达,在HFOV组及HFOV+PLV组分别低于CMV组及CMV+PLV组,组间比较统计学上有显著差异(p<0.01)。E-选择素、P-选择素和ICAM-1在肺组织和血清中的含量及其在肺组织中mRNA表达在CMV组和HFOV组分别高于CMV+PLV组及HFOV+PLV组.ET-1在肺组织和血清中的含量及其在肺组织中mRNA表达在CMV、HFOV、HFOV+PLV,及CMV+PLV四组间比较统计学上无显著性差异(P>0.05)。4.细胞凋亡:肺组织细胞凋亡及评分指数,HFOV组及HFOV+PLV组分别较CMV、CMV+PLV明显减少,两组比较有显著差异(p<0.01)。而CMV+PLV组和HFOV+PLV组又较CMV组和HFOV组减少,两组比较也有显著性差异(p<0.05)。caspsase-3活性及caspsase-3 mRNA表达,在HFOV及HFOV+PLV组,分别较CMV,CMV+PLV组明显减少,组间比较有显著差异(p<0.01)。而HFOV+PLV及CMV+PLV组又分别较HFOV和CMV组减少,两组比较也有显著性差异(p<0.05)。5.肺泡灌洗液: HFOV+PLV组中性粒细胞凋亡率和早期凋亡率较其它三组高,同时坏死细胞数量也最少。HFOV组及HFOV+PLV组中中性粒细胞早期凋亡率较CMV组和CMV+PLV组明显升高,差别有显著性(P<0.01);CMV组和CMV+PLV组中性粒细胞晚期凋亡率较HFOV组及HFOV+PLV组高,差异明显(P<0.01)。四个治疗组中CMV组中性粒细胞凋亡率最低,但与CMV+PLV组比较无明显差异(P>0.05)。总蛋白、丙二醛浓度各治疗组间的差异无统计学意义(P>0.05)。肺泡灌洗液E-选择素、P-选择素、ICAM-1和ET-1浓度各治疗组间的差异无统计学意义(P>0.05)。结论:与其它三种通气模式相比,HFOV联合PLV能提高蒸汽吸入性损伤兔的氧合水平,降低肺组织内粘附分子浓度及mRNA表达,减少肺组织调亡,减少肺组织内caspsase-3活性及caspsase-3 mRNA表达,提高肺泡中性粒细胞凋亡,减少肺组织内炎性细胞浸润,减轻肺损伤。是治疗吸入性损伤并呼吸衰竭较好的方法。更多还原

【Abstract】 Inhalation injury with respiratory failure or acute lung injury has high morality, need to be supported with ventilator .Conventional mandatory ventilation could cause ventilator-induced lung injury and could deteriorate pulmonary injury. As the protective lung ventilation strategy, high frequency oscillatory ventilation (HFOV) and partial liquid ventilation have important significance. Methods Thirty-two New Zealand rabbits were randomly divided into four groups after acute respiratory failure models caused by severe steam inhalation injury were established. Then they were ventilated by CMV,HFOV,CMV+PLV or HFOV+PLV,respectively. Femoral arterial blood samples were obtained respectively at 0.5,1.5,2.5,3.5h treatment timepoints for blood gas examination. respiratory mechanics and hemodynamic parameters were recorded. After the experimental period,the lowest lobe of the right lung was weighed for calculating weight ratio of lung water and lung tissue [(W-D)/W] , the middle lobe in the right lung were excised for histological sections. The right middle lobe of rabbit was excised for the examination of mRNA expression of E-selectin, P-selectin,ICAM-1 and ET-1 and their contents by the method of RT-PCR ( reverse transcription polymerase chain reaction ) and enzyme linked immunosorbent assay (ELISA) respectively. Partial excised lung tissue was used for pathological assessment and histological sections. The middle lobe in the right lung were excised for examination of mRNA expression of caspase-3 by the method of Real Time PCR (real time polymerase chain reaction ). Caspase-3 contents in lung tissue homogenates was examined by the method of colorimetric assay. Apoptosis indexes in the lung tissue were assessed with TUNEL FragELTM(Fragment End Labeling). Each one underwent whole-lung lavage. Cells were removed from bronchoalveolar lavage fluid (BALF) by low-speed centrifugation, and the resulting pellets were analyzed for total cell﹑differential counts and apoptosis analysis . Total white-cell numbers were determined by manual counting. Cell-free lavage fluid was processed for total protein (TP) and malondialdehyde (MDA) determinations. Results 1.(A) PaO2 in the four groups all decreased below 60mmHg and PaO2 of postinjury had statistical significance compared with pre-injury at the same group(P<0.01). PaO2 of postinjury and pre-injury between-group comparison showed no significance(P>0.05). (B) PaO2 in the four groups had increased since 0.5h treatment timepont,and values of PaO2 sustained continuously within 3.5 h treatment time. PaO2、PaO2/FiO2 in the HFOV and HFOV+PLV groups were higher than the corresponding data in CMV and CMV+PLV groups respectively (P<0.05, P<0.01). PaO2 in the CMV+PLV group at 1.5h and 2.5h treatment timepoint and PaO2 in HFOV+PLV group at 3.5h treatment timepoints showed significant advantages over the data in the corresponding non-PLV groups at the same timepoints (P<0.05, P<0.01). (C) Histological injury score found the HFOV groups were more alleviated than the CMV group,the lowest in the HFOV+PLV group and the highest in the CMV group. 2. (A) The contents of E-selectin,P-selectin, ICAM-1 and their mRNA expression in lung tissue homogenates in HFOV group, HFOV+PLV group were lower than them in CMV group ,CMV+PLV group respectively, and their comparison showed statistical significance(p<0.01). The contents of E-selectin, P-selectin, ICAM-1 and their mRNA expression in lung tissue homogenates in HFOV group, CMV group were higher than them in HFOV+PLV group , CMV+PLV group respectively , and their comparison showed statistical significance (p<0.01). The contents of ET-1 and mRNA expression in lung tissue homogenates in HFOV, HFOV+PLV, CMV, CMV+PLV showed no statistical significance (P>0.05). (B) Histological injury found the HFOV groups were more alleviated than the CMV group,the lowest in the HFOV+PLV group and the highest in the CMV group. 3. (A) Lung tissue apoptosis indexes in HFOV group and HFOV+PLV group were lower than them in CMV group and CMV+PLV group , between-group comparison had significant difference(P<0.01). HFOV+PLV group showed lowest apoptosis indexes.(B) caspase-3 and caspase-3 mRNA expression in the lung tissue in HFOV group and HFOV+PLV group were lower than them in CMV group and CMV+PLV group (P<0.01). caspase-3 and caspase-3 mRNA expression in the lung tissue in CMV+PLV group and HFOV+PLV group were lower than them in CMV group and HFOV group (P<0.05). 4. (A)in the CMV group ,the number of total and polymorphonuclear cell was obviously increased compared with other group( P<0.01) , HFOV+PLV have the lowest total and polymorphonuclear cell number .polymorphonuclear cell number in the CMV+PLV and HFOV+PLV groups comparison showed significance( P>0.05). (B) polymorphonuclear cell apoptosis in HFOV+PLV group were higher than other group, There was significant difference when compared with CMV,HFOV or CMV+PLV group. CMV group have the lowest polymorphonuclear cell apoptosis, There was no significant difference between CMV and CMV+PLV group. Conclusion high frequency oscillatory ventilation ; combined with partial liquid ventilation can improve arterial oxygenation; decreased the concentration and mRNA expression of E-selectin, P-selectin, ICAM-1 ;suppress lung tissue apoptosis induced by inhalation injury ,reduced the caspase-3 concentration and mRNA expression ;HFOV combined with PLV can increase polymorphonuclear cell apoptosis,alleviate polymorphonuclear cell infiltration in inhalation lung injury , reduce lung injury.high frequency oscillatory ventilation combined with partial liquid ventilation may be a optimal method for the treatment of acute respiratory failure caused by inhalation injury .更多还原