A Retrospective Study Of Acute Respiratory Distress Syndrome And The Protective Effect Of MTOR-siRNA On Acute Lung Injury In Mice

Part 1 A retrospective study of Acute Respiratory Distress SyndromeObjective:Our study aims to investigate the clinical characteristics and possible prognostic risk factors in patients with acute respiratory distress syndrome(ARDS).And to determine whether clinical characteristics and prognosis differ between pulmonary and extrapulmonary ARDS.Methods:This retrospective study included patients with ARDS who were admitted to the Intensive Care Units in our hospital between January 2014 and December 2018.Demographic,comorbidities,laboratory data,severity,organ failure and survival outcomes were collected and analyzed.The patients were classified into 2 groups(survival and non-survival)based on the hospital survival.The prognostic factors were identified using univariate logistic regression and the independent predictors associated with mortality were identified by multivariate logistic regression analysis.Moreover,patients were classified into pulmonary(direct)and extrapulmonary(indirect)ARDS and the differences in clinical characteristics and prognosis were analyzed.Results:85 patients with ARDS were analyzed.The overall hospital mortality was 55.3%.The median age was 60 years old,and the elderly(≥60 yrs)accounted for 51.8%.Comorbidities occurred in 65.9%of patients and the overall Charlson Comorbidity Index(CCI)was 1.0(0.0-3.0).The mean PaO2/FiO2 was 141.0 ± 45.9 mmHg.Acute Physiology and Chronic Health Evaluation II(APACHE II)score was 20.3±9.2.More than 80%of patients combined with extrapulmonary organ failure,Sequential Organ Failure Assessment(SOFA)score was 9.1 ± 3.9 and the number of extrapulmonary organ failure was 2.0(1.0-3.0).Multivariate logistic regression analysis showed that PaO2/FiO2(OR=0.969,95%CI=(0.952-0.985),p<0.001)was independent risk factors for reduced hospital mortality,while CCI(OR=1.685,95%CI=(1.140-2.492),p=0.009)and non-pulmonary SOFA score(OR=1.511,95%Cl=(1.196-1.908),p=0.001)were independent risk factors for increased hospital mortality.Age,BMI,APACHE II score and the number of extrapulmonary organ failure were associated with mortality as well.There was no difference in hospital mortality between pulmonary and extrapulmonary ARDS(56.8%vs 53.7%,p=0.470),as well as CCI score,PaO2/FiO2 and APACHE II score.Compared with pulmonary ARDS,the patients with extrapulmonary ARDS had higher non-pulmonary SOFA score(p<0.05)and more extrapulmonary organ failure(p<0.05).Conclusion:PaO2/FiO2 was independent risk factors for reduced hospital mortality,while CCI score and non-pulmonary SOFA score were independent risk factors for increased hospital mortality.Compared with pulmonary ARDS,there was no difference in mortality while extrapulmonary ARDS tended to have organ failure.Part 2 The protective effect of mTOR-siRNA on Acute Lung Injury in miceObjective:Establishing the animal model of Lipopolysacchride(LPS)induced Acute Lung Injury(ALI)and the administration method of giving mTOR-siRNA intratracheally.Then to investigate the protective effect of mTOR-siRNA in LPS induced ALI.Methods:Acute Lung Injury in mice was induced by giving LPS intratracheally(100mg/kg).The cell number in Bronchoalveolar Lavage Fluid(BALF)and the cytokines in the lung tissue were evaluated.The pulmonary edema was evaluated by detecting the total protein in BALF and the wet to dry ratio.The histopathological changes were evaluated as well.The expression of mTOR and p-S6 induced by LPS was evaluated using Western Blot,immunohistochemistry and immunofluorescence.We then gave different doses of mTOR-siRNA intratracheally to suppress the expression of mTOR.Pre-treated the mice with mTOR-siRNA(5mg/kg)intratracheally before LPS exposure,and evaluated the changes of inflammation,pulmonary edema,histopathological injury and survival to investigate the protective effect.Results:Compared to control group,mice given with LPS intratracheally(100 mg/kg)had higher total cell number and neutrophil number as well as higher percentage of neutrophil(p<0.01)in BALF.The level of IL-1β,IL-6,TNF-a and MIP-2 in lung tissue elevated significantly(p<0.05).The total protein in BALF and wet to dry ration increased significantly(p<0.05),suggesting pulmonary edema.The histopathology showed alveolar destruction and thickening of alveolar wall with neutrophils aggregation and alveolar hemorrhage,suggesting obvious lung injury.Consistently,the lung injury score,thickness of alveolar wall and crowed score increased while the number of alveoli decreased significantly(p<0.001).The expression of p-mTOR and p-S6 were elevated in the lung tissue after LPS exposure.Given mTOR-siRNA(5mg/kg,in a manner of 25 μg siRNA per day for 4 continuous days)intratracheally could downregulate the expression of p-S6,suggesting the suppression of mTOR.Compared with con-siRNA+LPS group,mTOR-siRNA could reduce the total cell numbers and neutrophil(p<0.05)in BALF,as well as the expression of IL-1β,IL-6,TNF-α and MIP-2(p<0.01)in lung tissue。The elevation of total protein in BALF and wet to dry ration induced by LPS were decreased significantly by mTOR-siRNA(p<0.05).The histopathology showed an attenuated pattern of lung injury.Consistently,the elevated lung injury score,thickness of alveolar wall and crowed score were also reduced while the number of alveoli increased significantly(p<0.05).However,the survival of mice treated with mTOR-siRNA did not get improved significantly(p=0.406).Conclusion:LPS induced the activation of mTOR and downstream protein p-S6 in lung tissue in ALI model.Intratracheal administration of mTOR-siRNA could decrease the mTOR activation induced by LPS,and significantly attenuated the inflammation,pulmonary edema and histopathological lung injury.Though the survival did not get improved significantly,mTOR-siRNA has a protective effect on LPS-induced acute lung injury in mice.