The Influence Of Extracorporeal Membrane Oxygenation And With Inline Continuous Renal Replancement Therapy On Intestinal Mucosa Barrier In Traumatic ARDS Piglet Model

Because of replace effect to lung completely and heart function partly,extracorporeal membrane oxygenation technology(ECMO)is now considered one of most effective supportive therapy for ARDS patient.However ECMO is an invasive treatment of non-biological material.mountains of evidence show that ECMO can lead to a series complications,such as systemic and local immune inflammatory response,oxidative stress,coagulation disorders and renal,pulmonary and cardiac functional damage.So ECMO treatment was extremely restricted,and only use treatment of late ARDS patients.So how to avoid the ECMO-related complications is an key point to improve its efficacy.With the advances in materials technology,the compications such as hemolytic reaction,coagulation disorders and other complications are decreasing during ECMO treatment.But systemic immune inflammatory response and catheter infection rate is still high.However in critically ill patients,the gastrointestinal tract is the main source of infection and inflammation occur.Our recent study also found ECMO treatment can lead to oxidative stress in systemic and local(gut,brain).Our recent study also found ECMO treatment can lead to intestinal mucosal barrier damage.These results showed that ECMO treatment itself can lead to the occurrence of the mucosal barrier.However,these studies were only in the normal state or normal oxygen and blood supply states.In severe ARDS state,because the immune inflammatory response and hypoxia can also lead to intestinal mucosal injury,ECMO therapy may increase the oxygen supply to protect intestinal barrier theoretically.Thus,there is both contradictory issues for ECMO treatment,the first aspect:ECMO therapy can correct organization hypoxia to protect the gastrointestinal mucosa barrier under severe ARDS;on the other hand:ECMO treatment itself can also lead to gastrointestinal mucosal barrier damage.How early intervention needs further discussion once injury the intestinal mucosa.About 80%of patients require continuous renal Continuous Renal Replacement Therapy(CRRT)during ECMO therapy.CRRT had been used to manage the fluid overload and it also proved to remove the inflammatory cytokines.Furthermore,CRRT therapy can increased intestinal tight junction protein,maintain the integrity of the intestinal tight junctions,and reduce the incidence of bacterial translocation.Therefore,we hypothesized that CRRT therapy combined with ECMO treatment can improve the intestinal mucosal barrier injury under ARDS condition.The means of the effect of ECMO and CRRT treatment on mucosal barrier under ARDS condition are still to further exploreMitochondria are organelles for oxygen metabolism.its damage can cause cell necrosis and apoptosis.In the previous study,we found that,ECMO treatment can lead to injury of myocardial mitochondrial ultrastructure and function.In addition,we also found,ECMO therapy can cause oxidative stress injury in intestine.The evidence shows that ECMO treatment can affect mitochondrial function.Another study shows that ECMO combined CRRT therapy can increases oxygen utilization than single ECMO treatment.Our previous study found that compared with CRRT combined ECMO treatment can relieve the destruction of myocardial mitochondrial structure,mitochondrial function and oxidative stress significantly resulting from ECMO treatment.This suggests that CRRT can improve mitochondrial dysfunction,increasing its functionality and reduce oxidative stress caused by ECMO.Therefore,we hypothesized that,ECMO may impair mitochondrial function through the intestinal mucosal barrier,but CRRT therapy can relieve mitochondrial damage caused thereby protecting the intestinal mucosa barrier resulting from ECMO treatment.Therefore,this study established a stable and suitable animal model of ARDS longer extracorporeal life support systems research by ARDS model by pulmonary contusion combined with ischemia-reperfusion injury.On the basis,to investigate the impact of ECMO treatment on ARDS porcine intestinal mucosa barrier the first time.We establish a integration of connection for CRRT and ECMO innovatively.On the basis,we investigate the influence of Continuous Renal Replacement Therapy(CRRT)combined with Extracorporeal membrane oxygenation treatment on intestinal mucosal barrier in ARDS piglet model.Finally,we further investigate the mechanism of the effects of ECMO therapy and ECMO combined with CRRT therapy on intestinal mucosa injury,namely its impact on mitochondrial structure and function.The results of this study will improve the clinical effectiveness of ECMO treatment,reduce the incidence of complications,and provide a theoretical basis for ECMO combined with CRRT treatment.Part I:The influence of extracorporeal membrane oxygenation therapy on intestinal mucosal barrier in a porcine model for traumatic acute respiratory distress syndromeStudy I:Establishing ARDS model by pulmonary contusion and ischemia-reperfusion injuryObjective:We explored a variety of early animal model of ARDS ways including lung lavage(saline or gastric juice),intravenous injection of oleic acid,gunshot wounds,ischemia-reperfusion combined injection of endotoxin.However,these models do not apply to in vitro studies long life-support systems.we aimed to establish a stable and suitable animal model of ARDS longer extracorporeal life support systems research by pulmonary contusion combined with ischemia-reperfusion injury.Methods:12 pigs were randomly divided into three groups.In each group,we used weighty fell on the chest of the pig(group A=0.2kg/(kg weight);group B=0.4kg/(kg weight);group C= 0.6kg/(kg weight)).Then the pig underwent the systemic ischemia-reperfusion and observed for 24hours.Hemodynamic and ventilation parameters were recorded.We assessed CT score after trauma,and the histopathological changes of lungs.Results:there was no remarkably difference of survival time between groups.The morbility of Group C was 75%.The animals in Group A all survived.And in Group A,the light changes of oxygenation index,CT score and the pathohistology were observed.In Group B,the oxygenation index was between 200 to 300.The CT score indicated that the half lung was contused.And there was mild to severe inflammatory infiltration and exudates in the lung parenchyma.Conclusion:This study established a stable and suitable animal model of ARDS longer extracorporeal life support systems research by pulmonary contusion combined with ischemia-reperfusion injury.Study ?:The influence of extracorporeal membrane oxygenation therapy on intestinal mucosal barrier in a porcine model for traumatic acute respiratory distress syndromeObjective:Venovenous Extracorporeal membrane oxygenation(vv-ECMO)treatment can significantly increase the immune inflammatory response,intestinal oxidative stress,as well as mucosal barrier injury.However,these studies were performed under non-ARDS conditions and in a short time.Whether ECMO treatment will injure the intestinal muscosal barrier under ARDS condition in a longer time has not been reported.Methods:Pulmonary contusion combined with ischemia-reperfusion injury was induced in 18 piglets.The animals were randomly divided into control,model,and ECMO groups.Blood samples were collected at-1,0,2,6,12,and 24 h during vv-ECMO therapy in order to measure the levels of I-FABP,D-lactate,and endotoxin in serum.The jejunum and colon were collected post-mortem and evaluated histopathologically.The tissue was also examined using electron microscopy,and intestinal tight junction proteins(ZO-1 and occludin)were measured after 24 h of ECMO therapy.Results:The serum markers evaluating the intestinal mucosal barrier deteriorated in the model group compared to the control group(p<0.05).At 2 h,serum I-FABP,D-lactate,and endotoxin were significantly increased in the ECMO group compared to the model group(p<0.05).At 12 h,I-FABP and D-lactate in the ECMO group dropped to model group levels.Serum D-lactate was slightly lower in the ECMO group(p>0.05)and serum I-FABP was significantly lower than in the model group(p<0.05)at 24 h.Similarly,serum endotoxin was slightly lower in the ECMO group than in the model group(p>0.05)at 24 h.After 24 h of ECMO therapy,the occludin and ZO-1 protein concentrations in jejunum and colon mucosa increased moderately compared to that in the model group(p<0.05).Morphologic structure of the jejunum and colon did not improved significantly after ECMO therapy.In conclusion,intestinal mucosal barrier continued to deteriorate in the model group.ECMO therapy aggravates intestinal mucosal injury at early(2 hours).The results show that ECMO therapy has adamage effect on the intestinal mucosal barrier in.Part ?:Continuous Renal Replacement Therapy(CRRT)treatment attenuates the damage of intestinal mucosal barrier induced by Extracorporeal membrane oxygenation(ECMO)treatment in traumatic ARDS piglet model Study I:Integrated Connection ways of Continuous renal replacement therapy combined venovenous extracorporeal membrane oxygenation circuit in vitro and in vivoObjective:ECMO and CRRT treatment at present are commonly used catheters respectively,and there is no integrated connection ways.This must increase a deep venous catheter,and will lead to an increase in the chance of infection.Therefore,this study established an integration connections for ECMO and CRRT innovatively and provide a theoretical basis for further research and clinical applications for the CRRT combined with ECMO treatment.Methods:we designed 4 methods to connecting CRRT to ECMO.The two devices was tested in a closed circuit primed using colloids.We varied the ECMO blood flow and the CRRT location,recording the pressures in the two devices.The systems were then used in the experimental surgery laboratory on a pig,and further evaluated the advantages and disadvantages of the connecting mannaers.Results:Connecting CRRT to ECMO before centrifugal pump,the CRRT did not operate.Connecting CRRT to ECMO between centrifugal pump and oxygenator,the ECMO circuit need to be cut.Connecting CRRT to ECMO before and after oxygenator,it would reduce the blood flow that passing through the oxygenator.Connecting CRRT to ECMO after oxygenator,we used the arterial cannula to replace the venous cannula,and there was no influence on the pressure and oxygenation.Conclusions:Connecting CRRT to ECMO,was the most practicable and economic method,and could used in the clinic practice.Study ?:Continuous Renal Replacement Therapy(CRRT)treatment attenuates the damage of intestinal mucosal barrier induced by Extracorporeal membrane oxygenation(ECMO)treatment in traumatic ARDS piglet modelObjective:ECMO treatment can significantly increase the immune inflammatory response,intestinal oxidative stress,as well as mucosal barrier injury.Continuous renal replacement therapy(CRRT)can improve intestinal mucosal barrier injury for MODS patients.Therefore,the study speculated,CRRT treatment can attenuates the damage of intestinal mucosal barrier caused by ECMO treatment.Methods:Pulmonary contusion combined with ischemia-reperfusion injury was induced in 18 piglets.The animals were randomly divided into Model,ECMO and CRRT+ECMO groups.Blood samples were collected at-1,0,2,6,12,and 24 h during vv-ECMO therapy in order to measure the levels of I-FABP,D-lactate,and endotoxin in serum.The jejunum and colon were collected post-mortem and evaluated histopathologically.The tissue was also examined using electron microscopy,and intestinal tight junction proteins(ZO-1 and occludin)were measured after 24 h of ECMO therapy.Results:At 2 h,serum I-FABP,D-lactate,and endotoxin were significantly increased in the ECMO group compared to the model group(p<0.05).since than,I-FABP and D-lactate in the ECMO group dropped to model group levels at 12 h.serum endotoxin was slightly lower in the ECMO group than in the model group(p>0.05)at 24 h.After 24 h of ECMO therapy,the occludin and ZO-1 protein concentrations in jejunum and colon mucosa increased moderately compared to that in the model group(p<0.05).Morphologic structure of the jejunum and colon did not improved significantly after ECMO therapy.ECMO combined CRRT treatment,the I-FABP,D-Lacate and Endoxtin at 2,6 h and 12h were significantly lower than that ECMO group respectively(p<0.05).Jejunum and colon pathology and electron microscopic examination showed:ECMO+CRRT group was significantly better than that model group(p<0.05),in which the jejunum pathology scores showed:ECMO joint CRRT group was significantly lower than that ECMO group(p<0.05).Although tight junction protein Occludin was slightly higher than the model and ECMO group in the ECMO+CRRT treatment group(p>0.05),ZO-1 in the jejunum and colon were significantly higher than the model group(p<0.05).In conclusion,CRRT treatment can attenuates the damage of intestinal mucosal barrier induced by ECMO treatment in early for ARDS piglet model.In the late ECMO combined CRRT therapy can continue to improve the role of mucosal injury.Part III:Effects of Continuous Renal Replacement Therapy(CRRT)on intestinal mitochondrial function during Extracorporeal membrane oxygenation(ECMO)treatment in traumatic ARDS piglet model Objective:ECMO treatment can damage the intestinal mucosal barrier,but the mechanism is unknown.Some research shows Continuous Renal Replacement Therapy(CRRT)can improve the mitochondrial function and oxidative stress injury.Mitochondrion is important organelles to maintain normal intestinal structure and function.Therefore,this study is to explore the effect of CRRT combined with ECMO treatment on intestinal mitochondrial function and oxidative stress injury by ARDS animal model.Methods:Pulmonary contusion combined with ischemia-reperfusion injury was induced in 18 piglets.The animals were randomly divided into Model,ECMO and CRRT+ECMO groups.Blood samples were collected at-1,0,2,6,12,and 24 h during vv-ECMO therapy in order to measure the levels MDA,SOD,GSH and T-AOC in serum.The jejunum and colon were collected post-mortem to analyse the MDA,SOD,GSH and T-AOC;mitochondrial function was assessed by activities of mitochondrial complexes I-V,mitochondrial membrane potential and mitochondrial ultrastructure by electron microscopy in jejunum and colon after 24 h of ECMO therapy.Results:The results show MDA in serum were increased persistently,conversely SOD,GSH and T-AOC were decreased persistently.After two hour's ECMO treatment,MDA concentration in serum was significantly higher than that in the model group(p<0.05),and reached the peak at 6 hours(p<0.05),since then began to decline.SOD decreased significantly after ECMO treatment and was significantly lower than the Model group in the 6 hours(p<0.05),but then began to rise,and is slightly higher than the model group level in the 24 hours(p>0.05).GSH was significantly lower than model group after two hours ECMO treatment(p<0.05),and to a minimum(p<0.05)at 6 hours,then gradually increased,and is slightly higher than the level of the Model group at 24 hours(p>0.05).MDA,SOD,GSH and T-AOC had no significant difference in the jejunum and colon between ECMO and Model group(p>0.05).After combined with CRRT treatment,the MDA in serum is decreased significantly than that of ECMO group,which in the first 12 hours of the most significant(p<0.05),and was significantly lower than that of the model group at 24 hours(p<0.05).Inversely SOD,GSH and T-AOC were higher than ECMO treatment group at different time points in different degrees,and gradually higher than the model group.And SOD,GSH and T-AOC were significantly higher than the level of model group at 24 hours(p<0.05).After CRRT joint ECMO treatment,MDA were significantly lower than the model group and the ECMO group;SOD,GSH and T-AOC were varying degrees higher than the model group and the ECMO group in jejunum and colon tissues.Mitochondrial electron microscope examination showed mitochondrial was swelling into vacuoles,mitochondria ridge dissolve or disappear between ECMO and Model group;mitochondrial complexes I-V and the mitochondrial membrane potential had no significant difference between ECMO and Model group(p>0.05).The swelling and vacuoles in mitochondrial were attenuated and mitochondrial ridge was more clearly than that of ECMO and the Model group after ECMO combined CRRT treatment.And increasing mitochondrial complex activity in jejunum and colon at varying degrees,which enzyme I,IV the most significant and V(vs model group,p<0.05).Mitochondrial membrane potential is also higher than the Model group,and was most significant in jejunum(vs Model group,p<0.05).Conclusion:ECMO treatment at early can be significantly increased oxidative stress in the blood for ARDS animal models,but the ECMO therapy at late,the damage relieved gradually.Oxidative stress had no significant effect in jejunum and colon.CRRT joint ECMO treatment can significantly reduce the oxidative stress induced by ECMO treatment in early,and has continued to improve the injury and ultimately significantly improve intestinal oxidative stress for ARDS animal models.ECMO treatment had no significant effect on mitochondrial function destroyed in ARDS condition.ECMO combined with CRRT treatment can significantly improve the mitochondrial function.