Experimental Study On The Combined Therapy Of Exogenous Hydrogen Sulfide And V-V ECMO In Acute Respiratory Distress Syndrome

BackgroundAs the development of science and technology advances,there has seen a continuous hike with the utility of explosive weapons in modern warfare and terrorist attacks.The dreadful blast wave raises the atmospheric pressure,which formulates acts on human bodies so directly that multiple organ damages can be caused,among which the lungs being the most vulnerable,featuring in a gas filled structure.Blast lung injury can be defined as the damage in the lung under the blast wave of explosion,with the primary pathological changes of the rupture and hemorrhage of pulmonary alveoli,followed by pulmonary edema and emphysema.Minor symptoms include chest pain and stuffiness while severe ones are typical of acute respiratory distress syndrome,also known as ARDS.Oxygen therapy and mechanical ventilation are conventional treating strategy of ARDS.However,the survival rate under these management is not promising--mortality in mild and moderate ARDS can be as high as 45%,whereas severe exceeding 60%.Extracorporeal membrane oxygenation(ECMO),a form of advanced life support,can be introduced as an aspiring way of maintaining patients' lives.It plays a role as a substitute of the lung and heart with the function of gas exchange:after draining the oxygen-free venous blood,the oxygenator realizes the process and then oxygenated blood is reinfused back to the venous or arterial systems.Generally,according to the differences of drainage and reinfusion accesses,ECMO can be categorized into two models:the veno-venous ECMO(V-V ECMO)and the veno-arterial ECMO(VA ECMO),with the former being the very remedy of ARDS.V-V ECMO,notwithstanding,replaces the patients'lungs,it cannot serve as the treatment of lung diseases,but rather striving for more time to recover the function of the lungs,to apply drug therapy and to exploit other direct means of lung treatment.Consequently,complications caused by prolonged ECMO support are likely to occur,namely hemorrhage,infection and pipeline failure,which exacerbates the prognosis of the patients.Hydrogen sulfide(H2S)is a kind of gaseous signal molecule in human bodies and carries much weight in multiple organs.Researches on drug therapies of ARDS indicates that application of exogenous H2S not only promotes the functions of pulmonary ventilation and aeration by relaxing the smooth muscle of the vasa publica and dilating the airway,but also inhibits systemic inflammatory responses.For the first time,this study aims at exploring the effects of the introduction of H2S in the treatment of ARDS with V-V ECMO and its potential molecular biological mechanisms.ObjectivesIn this research,we 'are going to delve into the changes of pulmonary ventilation and aeration functions,inflammatory responses in lung tissues,pathological characteristics and moleculer biology in lung cells changes in the combined therapy of H2S and V-V ECMO in ARDS,as well as its superposition for lung protection.Methods15 healthy adult beagles were selected,weighing from 12 to 18 kg.Before the experiments,each dog had been fasting for 24 hours and was tied up on the operation table on its back.To achieve anesthesia,an intravenous indwelling catheter was inserted in the upper limb to apply 3%pentobarbital sodium(30mg/kg).Then each dog was intubated with 6.5#trachea cannula for respiratory assistance,Routine preoperative preparations included catheterization,skin preparation,disinfection and drape.In addition,the ECG monitor and pulse oximeter monitor were connected.The PiCCO duct was inserted into the left femoral artery,and Swan-Ganz catheter and its outer layer into the left jugular vein,The ARDS model was established by slowly pumping an even mixture of 0.2ml/kg oleic acid and 30ml normal saline thorough the outer layer.Conventional ventilation mode was set before ARDS as follows:with the tidal volume 10ml/kg,respiratory rate 20 times per minute and PEEP at 5cmH2O,while lung protective ventilation strategy was applied following a successful modeling,with the tidal volume 6ml/kg,respiratory rate 25 times per minute and PEEP at 10cmH2O.The ECMO outflow cannula was inserted into the left femoral vein and the inflow cannula into the left jugular vein.Each heparinized,both outflow and inflow cannula were connected to the ECMO system,which had been well prepared during the period of cannulation.These beagles were randomly allocated to 5 groups:Control Group(n=3):PiCCO duct,Swan-Ganz catheter,ECMO drainage tube and ECMO pump line were inserted after anaesthesia,while devoid of the ARDS modeling and ECMO running;PV Group(n=3):PiCCO duct,Swan-Ganz catheter,ECMO drainage tube and ECMO pump line were inserted after anaesthesia,followed by the ARDS modeling;only lung protective ventilation strategy was introduced;PV+NaHS Group(n=3):PiCCO duct,Swan-Ganz catheter,ECMO drainage tube and ECMO pump line were inserted after anaesthesia,followed by the ARDS modeling;lung protective ventilation strategy was introduced combined with the application of exogenous H2S;ECMO+PV Group(n=3):PiCCO duct,Swan-Ganz catheter,ECMO drainage tube and ECMO pump line were inserted after anaesthesia,followed by the ARDS modeling;lung protective ventilation strategy was introduced combined with the application of V-V ECMO;ECMO+PV+NaHS Group(n=3):PiCCO duct,Swan-Ganz catheter,ECMO drainage tube and ECMO pump line were inserted after anesthesia,followed by the ARDS modeling;lung protective ventilation strategy and V-V ECMO was introduced combined with the application of exogenous H2S.Proper fluid infusion,measures to stabilize hemodynamics,maintenance of water and electrolytic balances were conducted accordingly.When V-V ECMO was applied,the activated clotting time(ACT)of each subject was preserved for 180?220s.The parameters of the ECMO was as follows:blood flow of 70ml/kg/min,oxygen concentration of 0.6 and sweep flow of 2.01/min.The concentration of NaHS solution was 50mM,which was administrated at a bolous of 0.5mg/kg/h.Data regarding parameters of the respirator,the ECMO system,hemodynamics,pulmonary ventilating and gas exchange functions were recorded in all groups before and after modeling,as well as 5min,30min,1h,2h,4h,8h,12h and 24h after ECMO operation and/or drug therapy.These parameters included Ppeak,Pplat,Compliance,ELWI,PVPI and 01.Meanwhile,peripheral arterial blood was drawn for arterial blood gas analysis to moniter PaO2,PaCO2,A-aDO2 and electrolyte conditions of each dog.Subjects surviving for 24 hours were executed except for those died in the course of experiments.Time of death of each one was recorded.Afterwards,lung tissues were collected to figure out the dry/wet lung weight ratio,and pathological sections were observed for the structural changes in pulmonary alveoli and infiltration of local inflammatory cells under light microscope.The possible mechanisms of injury and repair were explored through the changes of pulmonary inflammatory mediators(IL 6,TNF?),autophagy(Beclin 1,LC 3),apoptosis(c-Caspase 3),et al.Results1.ARDS modeling101±22min after pumping the mixture of oleic acid,there were decreases in peripheral blood oxygen saturation,lung compliance and arterial partial pressure of oxygen in the subjects.Based on the 2011 Berlin Standard,ARDS was diagnosed for the following criteria:(1)lung compliance decreasing by 50%;(2)OI?200mmHg;(3)PCWP ?18mmHg.Therefore,the success rate of modeling was 100%.2.Outcomes of experimental animalsAll dogs in the PV Group died,at 18h,14.5h and 21h after modeling,respectively,with a mortality of 100%;one dog in the PV+NaHS Group died 18h after modeling,with a mortality of 33.3%;all the dogs in other groups were executed 24h after ECMO operation or drug therapy.3.ECMO system conditionsThe ECMO operation was smooth,without signs of thrombus or aeroembolism in ECMO tubes and oxygenator.Besides,severe complications such as refractory hemorrhage,electrolyte imbalances and unstable hemodynamics did not appear in the experiments.4.Changes of pulmonary ventilation and pneumodynamics with combined therapy of H2S and V-V ECMOThe baseline indicators of respiratory mechanics were not significantly different among each group.After successful modeling,the Ppeak and Pplat of the experimental groups were higher than that of the control group while the Compliance much lower,with no differences among the experimental groups.As for Ppeak,the values of those dogs in ECMO+PV Group and ECMO+PV+NaHS Group dropped after the beginning of ECMO operation,reaching at a point notably lower than that of the PV Group and PV+NaHS Group.Specifically,the Ppeak in the ECMO+PV+NaHS Group were not disparate in the first 8 hours after ECMO operation,and started to excel that of the ECMO+PV Group after 12 hours.With regard to Pplat,the recorded values in ECMO+PV Group and ECMO+PV+NaHS Group commence decreasing remarkably 30 min after ECMO operation than those of the PV Group and PV+NaHS Group,lasting to the termination of experiments;meanwhile,there was no obvious gap compared to the Control Group 1 hour after operation.Furthermore,the figures showed a decline at the 4th hour after ECMO operation in the ECMO+PV+NaHS Group when compared with the ECMO+PV Group,yet the differences vanished 8 hours later.In the PV+NaHS Group,Pplat was significantly lower than that in the PV Group through 4th to 8th hour.In lung compliance,the values in the ECMO+PV+NaHS Group showed a significant increase since the 1st hour after ECMO operation than the PV Group,the PV+NaHS Group and the ECMO+PV Group,which was not notably divergent as the Control Group 12 hour after operation;besides,the values in the PV+NaHS Group exceeded prominently than the PV group 8 hours after drug therapy.The indicators of pulmonary diffusion function at baseline were not significantly different.The alveolo-arterial oxygen partial pressure difference(A-aDO2),extravascular lung water index(ELWI)and pulmonary vascular permeability index(PVPI)in each experimental group were significantly higher than the Control Group after modeling,while the arterial partial pressure of oxygen and oxygenation index much lower.Comparing the A-aDO2,the result showed that the values in PV+NaHS Group was significantly lower than PV Group during 30min?12h,while the values in ECMO+PV Group and the ECMO+PV+NaHS Group were exclusively lower than PV Group and PV+NaHS Group from 30min after operation to the end of the experiments.In addition,parameters in ECMO+PV+NaHS Group indicated a significantly decrease compared to that in ECMO+PV Group during 30min?12h.In terms of the ELWI,the values in the ECMO+PV Group and the ECMO+PV+NaHS Group were exclusively lower than PV Group and PV+NaHS Group from the 30th min to the end of the experiments.In addition,the ECMO+PV+NaHS Group indicated a decrease 2 hours after ECMO operation compared to the ECMO+PV Group,and the PV+NaHS Group lower than PV Group during 4?12h.For PVPI,the figures in all the experimental groups were not significantly different but greater than the control group at each recording point,except that the ECMO+PV+NaHS higher than the ECMO+PV Group at the 12th hour.Comparing the arterial partial pressure of oxygen,the data showed an increase in the ECMO+PV Group and the ECMO+PV+NaHS Group than the PV Group and the PV+NaHS Group 30min after ECMO operation,with no difference between the two former groups.In the ECMO+PV Group,values were not disparate from the control group 2 hours after operation.During 1?8 hours after drug therapy,the PV+NaHS Group saw a promising result than the PV Group,while no difference existed 8 hours later.5.Lung histological changes with combined therapy of H2S and V-V ECMOCompared with control group,the volume as well as the weight of experimental groups is significantly larger.And the lungs of control group were pale pink with no extra lung water,while diffuse alveolar hemorrhage was seen in the lungs of experimental groups and the inferior lobes of both lung were hyperemia and edema with lots of lung water.The alveolar septum structure is normal in CON group,with no obvious edema,consolidation or inflammatory cell infiltration,while no normal alveolar structure could be seen in those of experimental groups.And diffuse inflammatory cell infiltration and large number of red cells exudation were found in lung tissues of experimental groups.But the lesions in lung of ECMO+PV+NaHS group were significantly minor.6.Inflammatory responses in lung with combined therapy of H2S and V-V ECMOCompared with control group,inflammatory factor infiltration occored in experimental groups.Either IL-6 or TNF a was significantly higher in PV group than in other groups.The inflammatory factors appeared similar in PV+NaHS group and ECMO+PV group,while reduced in ECMO+PV+NaHS group.7.Autophagy and apoptosis of lung with combined therapy of H2S and V-V ECMOCompared with control group,LC-3 and Beclin-1,autophagy related protein,were significantly suppressed by the administration of exogenous H2S,especially in ECMO+PV+NaHS group.The same phenomenon appeared in c-Caspase 3,which is related to apoptosis.But the expressions of those proteins between PV+NaHS group and ECMO+PV group were similar.ConclusionsV-V ECMO can significantly improve the body oxygenation and reverse the condition of hypoxemia,by effectively reducing the airway resistance,alleviating lung compliance and protecting lung injuries,which provides more "rest" to the injury lung.And the combined therapy with exogenous H2S can attenuate cell autophagy and apoptosis thus protecting lung cells and improving respiratory function and making the effects of V-V ECMO even better.And if there were no ECMO,continuous intravenous administration of exogenous H2S could also improve the body oxygenation in 8 hours with the support of mechanical ventilation,thus winning the precious time for evacuation.