| Cardia carrest(CA)is the most fatal emergencies,which has become one of the major issues of public health worldwide.According to statistics updated by the American Heart Association in 2019,the incidence of out-of-hospital cardiac arrest(OHCA)were an average of 140.7 adults per 100,000 people,the survival to discharge were 10.4%,and only 8.4 % of survivors had good neurological function.At present,although people have a better understanding of the pathogenesis and progression of cardiac arrest,and many treatment measures such as mild hypothermia treatment,neuroprotective drugs,removal of active oxygen,inhibition of white blood cell activation,and Anti-inflammatory therapy,and so on.However,the treatment effect of heart and brain dysfunction after cardiac arrest are unsatisfactory.Therefore,it is important to further clarify the mechanism of heart and brain dysfunction after CA,and to develop more effective methods for organ protection after resuscitation.Up to now,there have been more and more researches on the protection of ischemia-reperfusion injury after cardiopulmonary resuscitation.A large number of studies have tried to find new drugs that can reduce or prevent ischemia-reperfusion,and some of them have made progress in animal models.In recent years,studies have shown that sodium octanoate(SO),mesenchymal stem cells(MSCs)and aldehyde dehydrogenase activator-1(Alda-1)may have an improved effect on organ function protection after resuscitation.In this study,we established a CA ventricular fibrillation pig model to explore the efficiency and safety of sodium octanoate,mesenchymal stem cells,and Alda-1.The successful implementation of the above-mentioned studies will likely provide powerful new methods for clinical resuscitation and treatment,which has far-reaching scientific significance and social value.Part 1 Study on the Effect and Mechanism of Sodium Octanoate in Improving Myocardial and Cerebral Dysfunction after Cardiopulmonary ResuscitationObjective:To establish a porcine model of CA and resuscitation,and then investigated the effects of sodium octanoate on cardiopulmonary resuscitation efficacy and post-resuscitation multiple organ dysfunction in a porcine model.Methods:Twenty-four male domestic pigs weighing 39.0±2.3 kg were utilized.The animals were then randomized with the Sealed Envelope Method into 1 of the 3 groups: sham(n=6),cardiopulmonary resuscitation group(CPR,n=10),CPR and sodium octanoate group(CPR+SO,n=8).The animals in the Sham group only finished the animal preparation without experiencing the procedure of CA and resuscitation.In the CPR and CPR+SO groups,ventricular fibrillation was induced by 1-m A alternating current via the pacing catheter and untreated for 8 min,and then CPR was performed for 8 min;both of which was used to establish the pig model of CA and resuscitation.At the beginning of CPR,the CPR+SO group was given sodium octanoate of 28.5mg/kg dissolved in normal saline,and the other groups were given the same amount of normal saline.At 8 min after successful resuscitation,The following measurements were carried out in the experiment:(1)Baseline heart rate(HR),mean arterial pressure(MAP),regional cerebral oxygen saturation(r SO2),pressure of end-tidal carbon dioxide(PETCO2),arterial blood gas and lactate were measured prior to the induction of CA.(2)The changes of CPP,ETCO2,r SO2 and CBF were recorded at 1,2,3,4,5,6,7 and 8 min during CPR,and the outcomes of CPR including rate of resuscitation success,duration of CPR,number of defibrillations and dosage of epinephrine were recorded during CPR.(3)After ROSC,the MAP,HR,r SO2,PETCO2 were evaluated at 1,2,4h post-resuscitation(4)Myocardial function including stroke volume(SV)and global ejection fraction(GEF)were evaluated with the Pi CCO system at 1,2,4h post-resuscitation.Arterial and venous blood samples were collected at1,2,4,24 h post-resuscitation.The pyroptosis related inflammatory cytokine including Interleukin1?(IL-1?)and interleukin-18(IL-6),the injury biomarkers of heart and brain including cardiac troponin I(c Tn I),neuron-specific enolase(NSE)and S100 B protein(S100B).(5)The neurological deficit score(Neurological deficit score,NDS)was scored at 24 h post-resuscitation,and then the animals were euthanized.For the evaluation of pyroptosis and cell apoptosis in the heart and brain,left ventricular myocardium and cerebral cortex were harvested immediately after animal euthanasia at 24 h post resuscitation.Subsequently,the levels of IL-1?,IL-18 and pyroptosis related proteins,apoptosis expression in the heart and brain were measured.Results:1.Before CA,body weight,HR,MAP,PETCO2,lactate and body temperature did not differ among the four groups(all P>0.05).2.During CPR,the value of CBF、PETCO2,r SO2 in CPR+SO group were significantly better compared with the CPR group(all P<0.05),and CPP value was better than CPR group all the time except at 5 min(all P<0.05).Consequently,a 100% of resuscitation success was achieved in the CPR+SO groups(8/8);however,only 50% of resuscitation success was obtained in the CPR group(9/12).The rate of resuscitation success,the duration of CPR,number of defibrillation and dosage of epinephrine were superior to the CPR group(all P?0.05).3.After resuscitation,HR,MAP,r SO2 and CBF were decreased in both CPR group and CPR+SO group.The values of CBF and r SO2 in CPR+SO group were higher than those in CPR group after resuscitation,and the value of r SO2 in CPR group was significantly higher than that in CPR group at 1 h,2 h and 4 h after resuscitation(all P<0.05).The value of HR in CPR group was higher than that in CPR+SO group after 0.5 h,and there was significant difference between the two groups(all P<0.05),but the trend of MAP value in the two groups was consistent,and no significant difference between the two groups(P>0 05).4.After resuscitation,at the 0.5 h the p H value in the Sham group was significantly lower than that in the Sham group and CPR+SO group,while the lactate level in the CPR group was significantly higher than that in the Sham group and the CPR+SO group(all P<0.05).At 1 h after resuscitation,the p H value of the animals in the CPR+SO group and the CPR group gradually returned to the baseline level,while the level of lactic acid decreased gradually,and there was no significant difference between the two groups(all P>0.05).After resuscitation,the Pa O2 value of CPR group was significantly lower than that of Sham group at each time point after resuscitation,and the difference between groups was statistically significant(all P<0.05),but also lower than that of CPR+SO group,no significant difference between groups(all P>0.05).5.After resuscitation,compared with Sham group,CPR group and CPR+SO group,SV and GEF were decreased and the level of c Tn I was increased.However,compared with the CPR group,the GEF value at 1 h,2 h and 4 h after resuscitation and the SV value at 1 h,2 h and 4 h after resuscitation in the CPR+SO group were significantly higher than those in the CPR+SO group(all P<0.05).At 4h and 24 h after resuscitation,the level of c Tn I in CPR+SO group was significantly lower than that in CPR group(all P?0.05).6.After resuscitation,the serum levels of NSE and S100 B were observed in CPR group and CPR+SO group.However,compared with the CPR group,the NSE in CPR+SO group were decreased significantly at 1 h,2 h,4 h and 24 h after resuscitation,(all P < 0.05).Compared with the CPR group,the S100 B value of the CPR+SO group decreased further at 2h and 24 h after resuscitation,and it was significant difference between the two groups(all P<0.05).In addition,the NDS value at 24 hours after resuscitation in the CPR+SO group was significantly lower than that in the CPR group(all P<0.05).7.At 24 h post-resuscitation,the contents of IL-1? and IL-18 in myocardium,cortex and hippocampus of CPR group and CPR+SO group were increased.However,the levels of IL-1? and IL-18 in myocardium,cortex and hippocampus in CPR+SO group were significantly lower than those in CPR group.(all P<0.05).8.At 24 h post-resuscitation,apoptosis was found in the heart and brain tissues of CPR group and CPR+SO group,which was higher than that of Sham group(all P< 0.05).However,the positive rate of apoptotic cells in CPR+SO group was significantly lower than that in CPR group(all P<0.05).9.At 24 h post-resuscitation,the results of Western Blot in myocardium,cortex and hippocampus of CPR group and CPR+SO group showed that the pyroptosis related proteins ASC,C-CAS-1,GSDMD and NLRP3 were higher than those of Sham group.However,compared with the CPR group,all the pyroptosis related proteins were decreased in the CPR+SO group,and there were significant differences between the two groups(all P<0.05).Conclusion:In a porcine model of CA and resuscitation,the use of sodium octanoate can increase the level of cardio-cerebral perfusion during resuscitation.After resuscitation,it can improve the cardio-cerebral dysfunction and improve the outcome.Among them,sodium octanoate produces cardio-cerebral protective effect after resuscitation by inhibiting pyroptosis reaction and cell death.Part 2 Study on mechanism and effects of mesenchymal stem cells in Improving Myocardial and Cerebral Dysfunction During Cardiopulmonary ResuscitationObjective:To establish a porcine model of CA and resuscitation,and then investigate the effects of mesenchymal stem cells(MSC)on post-resuscitation cardiac and cerebral injuries and its potential mechanisms.Methods:Twenty-four male domestic pigs weighing 39±1.5 kg were used.The animals were then randomized with the Sealed Envelope Method into 1 of the 3 groups: sham(n=6),CPR group,(n=10),MSC group(n=8).The animals in the Sham group only finished the animal preparation without experiencing the procedure of CA and resuscitation.MSCs were pretreated in MSC group.In the CPR and MSC groups,ventricular fibrillation was induced by 1-m A alternating current via the pacing catheter and untreated for 8 min,and then CPR was performed for 8 min;both of which was used to establish the pig model of CA and resuscitation.MSC pretreatment scheme: each experimental pig took 5000000/kg unit MSCs as the total amount,and 250 ml of it were injected at the three days before the experiment and at the one and a half days before the experiment.At 8 min after successful resuscitation,The following measurements were carried out in the experiment:(1)Baseline HR,MAP,r SO2,PETCO2,arterial blood gas and lactate were measured prior to the induction of CA.(2)The changes of CPP,ETCO2,r SO2 and CBF were recorded at 1,2,3,4,5,6,7 and 8 min during CPR,and the outcomes of CPR including rate of resuscitation success,duration of CPR,number of defibrillations and dosage of epinephrine were recorded during CPR.(3)After ROSC,the MAP,HR,r SO2,PETCO2 were evaluated at 1,2,4h post-resuscitation(4)Myocardial function including SV and GEF were evaluated with the Pi CCO system at 1,2,4h post-resuscitation.Arterial and venous blood samples were collected at1,2,4,24 h post-resuscitation.The pyroptosis related inflammatory cytokine including IL-1? and IL-6,the injury c Tn I,NSE and S100 B.(5)The NDS was scored at 24 h post-resuscitation,and then the animals were euthanized.For the evaluation of pyroptosis and cell apoptosis in the heart and brain,left ventricular myocardium and cerebral cortex were harvested immediately after animal euthanasia at 24 h post resuscitation.Subsequently,the levels of IL-1?,IL-18 and pyroptosis related proteins,apoptosis expression in the heart and brain were measured.Results:1.Before CA,body weight,HR,MAP,PETCO2,lactate and body temperature did not differ among the four groups(all P>0.05).2.During CPR,CPP in CPR group and MSC group increased gradually in the first 4 minutes,then decreased and remained stable in the last 4 minutes,r SO2 in both groups increased steadily within 8 minutes,while CBF decreased gradually in both groups,however,the indexes of CPP,PETCO2 and CBF in MSC group were significantly better than those in CPR group at most time points during resuscitation(all P<0.05).In addition,compared with CPR group,the cardiopulmonary resuscitation time,defibrillation times,epinephrine dosage were less,but there was no statistical difference.However,the rate of return of spontaneous circulationsuccess,4h survival rate and 24 h survival rate were increased in MSC group,and there were significant differences between in the two groups(all P?0.05).3.After resuscitation,HR,MAP,rSO2 and CBF were decreased in both CPR group and CPR+SO group.The values of CBF and r SO2 in CPR+SO group were higher than those in CPR group after resuscitation,and the value of r SO2 in CPR group was significantly higher than that in CPR group at 1 h,2 h and 4 h after resuscitation(all P<0.05).The value of HR in CPR group was higher than that in CPR+SO group after 0.5 h,and there was significant difference between the two groups(all P<0.05),but the trend of MAP value in the two groups was consistent,and no significant difference between the two groups(all P>0 05).3.After resuscitation,compared with Sham group,HR in CPR group and MSC group were increased,while MAP,r SO2 and CBF were decreased.However,compared with CPR group,HR gradually slowed down and r SO2 increased continuously in MSC group.Among them,HR in MSC group was significantly better than that in CPR group at 2 h and 3 h post-resuscitation,and there were significant differences between the two groups(all P<0.05).There was no significant difference in the levels of MAP and CBF between the two groups after resuscitation(all P>0 05).4.After resuscitation,compared with the Sham group,the level of p H decreased and the level of lactic acid increased in both groups.At 0.5 h post-resuscitation,the p H value of CPR group was significantly lower than that of Sham group and MSC group,and the level of lactic acid was significantly higher than that of Sham group and MSC group(all P<0.05).After 1 hour of resuscitation,the p H value of animals in CPR group and MSC group gradually returned to the baseline level,while the level of lactic acid decreased gradually,and no significant difference between the two groups(all P>0.05).Pa O2 decreased after resuscitation in both groups.Among them,the Pa O2 value of CPR group was significantly lower than that of Sham group all the time after resuscitation,,but there was no significant difference between the CPR group and MSC group(all P> 0.05).5.After resuscitation,the values of SV and GEF decreased and the level of serum c Tn I increased both in the CPR group and MSC group.However,compared with the CPR group,the GEF value at 1 h,2 h and 4 h post-resuscitation and the SV value at 1 h and 2 h post-resuscitation in the MSC group were significantly higher than those in the MSC group(all P< 0.05).At 24 hours after resuscitation,the level of c Tn I in the MSC group was significantly lower than that in the CPR group,and there were significant differences between the two groups(all P<0.05).6.Compared with Sham group,the serum levels of NSE and S100 B in CPR group and MSC group increased after resuscitation.However,compared with CPR group,NSE values at 1 h,2 h,4 h and 24 h post-resuscitation and S100 B at 1 h,2 h and 24 h post-resuscitation in MSC group were significantly lower than those in MSC group(all P<0.05).Compared with the CPR group,the NDS score of the MSC group was significantly better than that of the CPR group,and the difference between the two groups was statistically significant(P<0.05).7.Compared with Sham group,the expression of inflammatory factors related to pyroptosis in myocardium,hippocampus and cortex was increased in CPR group and MSC group at 24 hours post-resuscitation.Compared with CPR group,the levels of IL-1? in myocardium,IL-18 in cortex,and IL-1? and IL-18 in hippocampus were statistically significant different between the two groups(all P<0.05).8.At 24 hours post-resuscitation,apoptosis was found in the heart and brain tissues of CPR group and MSC group,which was higher than that of Sham group(all P<0.05).However,the positive rate of apoptotic cells in MSC group was significantly lower than that in CPR group(all P<0.05).9.The results of Western Blot showed that the expressions of NLRP3,ASC,C-CAS-1 and GSDMD in myocardium,cortex and hippocampus in CPR group and MSC group were higher than those in Sham group.However,the protein expressions of NLRP3,ASC,C-CAS-1 and GSDMD in myocardium,cortex and hippocampus in MSC group were significantly lower than those in CPR group(all P<0.05).Conclusion:In a porcine model of CA and resuscitation,the pretreatment of MSCs can increase the level of cardio-cerebral perfusion,reduce the injury of heart and brain organs,and then improve the outcome of resuscitation.MSCs protect heart and brain after resuscitation by improving cell death and inhibiting pyroptosis.Part 3 The effects of different durations of ABO on the outcomes of resuscitation after TCAObjective:To establish a porcine model of CA and resuscitation,and then investigate the effects of Alda-1 on post-resuscitation cardiac and cerebral injuries and its potential mechanisms.Methods: Twenty-four male domestic pigs weighing 38.4±2.6 kg were utilized.The animals were then randomized with the Sealed Envelope Method into 1 of the 3 groups: sham(n=6),CPR group,(n=10),Alda-1 group(n=8).The animals in the Sham group only finished the animal preparation without experiencing the procedure of CA and resuscitation.MSCs were pretreated in MSC group.In the CPR and Alda-1 groups,ventricular fibrillation was induced by 1-m A alternating current via the pacing catheter and untreated for 8 min,and then CPR was performed for 8 min;both of which was used to establish the pig model of CA and resuscitation.At 8 min after successful resuscitation,The following measurements were carried out in the experiment:(1)Baseline HR,MAP,r SO2,PETCO2,arterial blood gas and lactate were measured prior to the induction of CA.(2)After ROSC,the MAP,HR were evaluated at 1,2,4h post-resuscitation(4)Myocardial function including SV and GEF were evaluated with the Pi CCO system at 1,2,4h post-resuscitation.Arterial and venous blood samples were collected at1,2,4,24 h post-resuscitation.The pyroptosis related inflammatory cytokine including IL-1? and IL-6,the injury c Tn I,NSE and S100 B.(5)The NDS was scored at 24 h post-resuscitation,and then the animals were euthanized.For the evaluation of pyroptosis and cell apoptosis in the heart and brain,left ventricular myocardium and cerebral cortex were harvested immediately after animal euthanasia at 24 h post resuscitation.Subsequently,pyroptosis related proteins,pro-inflammatory cytokines,oxidative stress were assessed in samples of heart and brain.Results:1.Before CA,body weight,HR,MAP,PETCO2 and lactate did not differ among the three groups(all P>0.05).2.During CPR,the rate of resuscitation success,the duration of CPR,number of defibrillation and dosage of epinephrine were superior to the CPR group(all P?0.05).3.After resuscitation,the value of HR in the CPR group treated with Alda-1 was significantly lower than that in the CPR group(all P?0.05).Compared with CPR group,the MAP index of Alda-1 group were higher than that of CPR group,and gradually approached to Sham group,and there was no significant difference between these two groups(all P>0.05).4.After resuscitation,the p H decreased and the lactic acid increased,and the values of PO2 and Pa CO2 decreased gradually.At 0.5 h after resuscitation,the p H value of CPR group was significantly lower than that of Sham group and alda-1 group,and the level of lactic acid was significantly higher than that of Sham group and Alda-1 group(all P?0.05).From 1 h to 4 h post-resuscitation,the p H value of the animals in the Alda-1 group and CPR group were gradually returned to the baseline level,while the level of lactic acid decreased gradually,and there was no significant difference between the two groups(all P>0.05).Among them,there was no significant difference between the value of Pa O2 and Pa CO2 among the three groups(all P>0.05).5.Post-resuscitation myocardial dysfunction and damage,which were indicated by decreased GEF and increased c Tn I,were observed in the CPR and Alda-1 groups compared to the Sham group.However,compared with the CPR group,the values of SV and GEF in the Alda-1 group were significantly higher than those in the Alda-1 group 1 hour after resuscitation,and there were significant differences between the two groups(all P < 0.05).In the c Tn I index,the overall trend of the Alda-1 group was lower than that of the CPR group.At 4 h post-resuscitation,the serum c Tn I level of the CPR group was significantly lower than that of the CPR group(all P<0.05).6.Post-resuscitation brain damage,which were indicated by significantly increased NSE and S100 B,were observed in the CPR and Alda-1 groups compared to the Sham group(all P<0.05).However,compared with the CPR group,the NSE values of the Alda-1 group decreased significantly at 2 h and 4 h after resuscitation(all P<0.05).In addition,the NDS value at 24 h after resuscitation in the Alda-1 group was significantly lower than that in the CPR group(P<0.05).7.At 24 hours after resuscitation,the increase of inflammatory factors related to pyroptosis and the rate of apoptotic positive cells in myocardium,cortex and hippocampus in CPR group and Alda-1 group were higher than those in Sham group.Compared with CPR group,the levels of IL-1b and IL-18 in myocardium and hippocampus in Alda-1 group were decreased significantly(all P<0.05).In addition,compared with the CPR group,the positive rate of apoptotic cells were significantly decreased in the Alda-1 group(all P>0.05).8.At 24 h post-resuscitation,the pyroptosis related proteins ASC,C-CAS-1,GSDMD and NLRP3 in hippocampus,cortex and myocardium in CPR group and Alda-1 group were higher than those in Sham group.However,compared with CPR group,the expression of pyroptosis related protein were significantly decreased in Alda-1 group,and there were significant differences between the two groups(all P>0.05).9.At 24 h post-resuscitation,the levels of 4-hydroxynonenal and malondialdehyde in the hippocampus of the Alda-1 group and the CPR group were higher than those of the Sham group.However,the levels of both in Alda-1 group were significantly lower than those in CPR group(all P<0.05).10.At 24 h after resuscitation,the level of ALDH2 protein was highest in Sham group,and Alda-1 group followed it.Compared with CPR group,the expression of ALDH2 in Alda-1 group were significantly decreased(all P<0.05).Accroding to the result of ALDH2 activity,the activity of Alda-1 group was the highest,Sham was the second,and CPR was the lowest,and Alda-1 group were significantly higher than CPR group(all P<0 05).Conclusion:Our results demonstrate that Alda-1 improves cardiac and neurological dysfunction post-resuscitation in a swine model of CA,at least partly via inhibiting ROS mediated NLRP3 inflammasome activation and pyroptosis... |
PDF Full Text Request