The current study revealed that administration of Na2S at the time of CPR markedly improves myocardial and neurological function and survival after CA/CPR in mice. The neuroprotective effects of Na2S were associated with attenuated CA/CPR-induced caspase 3 activation in the brain. Global myocardial IR in isolated-perfused heart model confirmed the protective effects of Na2S. Administration of Na2S also prevented systemic oxidative stress and acceleration of mitochondrial permeability transition in the heart. Enhanced endogenous H2S production by cardiomyocyte-specific overexpression of CGL improved outcomes after CA/CPR. The salutary impact of Na2S treatment on the outcome of CA/CPR were associated with preserved serum nitrite and nitrate levels and enhanced phosphorylation of Akt and NOS3 in the brain and heart and AMPKα and GSK-3β in the brain. Finally, NOS3 deficiency abrogated the protective effects of Na2S on the outcome of CA/CPR. Taken together, these observations suggest that H2S confer organ protection and improve survival after CA/CPR at least in part via an NOS3-dependent mechanism.
The majority of research on cardiac arrest over the past half-century has focused on improving the rate of ROSC, and significant progress has been made. However, many patients succumb to the post-cardiac arrest syndrome after successful resuscitation. The current study was designed to investigate whether H2S could improve the outcome after “successful” resuscitation. While there was no difference in the rate of ROSC or CPR time to ROSC, mice that received Na2S immediately before CPR showed a markedly higher survival rate at 24h than did mice that received vehicle before CPR or Na2S 10 min after CPR. These observations suggest that administration of Na2S at the time of CPR improved outcomes at 24h after cardiac arrest by preventing the development of post-cardiac arrest organ injury and dysfunction after ROSC.
The current results suggest that Na2
S can protect against the development of post-cardiac arrest neurological dysfunction and injury when administered at the time of CPR (see ). Cardiac arrest and CPR activated caspase 3 and induced cell death in the brain of vehicle-treated mice. In contrast, the improved neurological function at 24h after CPR in mice treated with Na2
S was associated with marked attenuation of caspase 3 activation and cell death in the brain after CPR. Previously, Qu and colleagues showed that administration of NaHS increased brain infarct volume after permanent middle cerebral artery occlusion 11
. Differences in the models and the higher dose of NaHS (180 µmol/kg of NaHS, equivalent to ~90 µmol/kg of Na2
) used in the Qu study compared to the current study (7 µmol/kg of Na2
S) may explain the conflicting results of these studies.16
Pro-apoptotic effects of relatively high concentrations of H2
S have also been reported in pancreas.25
Nonetheless, our results suggest that physiological level of H2
S can have important protective effects on ischemic brain injury.
To examine the molecular mechanisms responsible for the anti-apoptotic effects of Na2
S, we tested whether or not Na2
S promotes Akt- or AMPK-dependent pro-survival signals in the brain. We found that Na2
S increased phosphorylation of Akt and GSK-3β in the brain cortex. Akt-dependent phosphorylation of GSK-3β inhibits GSK-3β activity and thereby decreases apoptosis 26, 27
. Similarly, administration of Na2
S 1 min before CPR prevented the CA/CPR-induced dephosphorylation of AMPKα and NOS3. It has been reported that activation of cGMP-dependent protein kinase (PKG) may confer cardioprotection via GSK-3β.27
It is conceivable that Akt- and/or AMPKα-dependent NOS3 activation activates the PKG-dependent neuroprotective mechanism via GSK-3β phosphorylation in our model.
Similar to neurological function, myocardial function at 24h after CA/CPR was also improved by administration of Na2
S (see ). These results are reinforced by our findings that administration of Na2
S starting immediately after reperfusion, but not when administered 40 seconds after reperfusion, markedly improved myocardial function in the isolated-perfused mouse heart subjected to global IR. These results are consistent with a previous report that NaHS was cardioprotective in isolated rat hearts 15
. The relatively narrow temporal window of opportunity for Na2
S to improve the outcome of CA/CPR is consistent with previous studies of ischemic postconditioning. For example, Cohen and colleagues recently reported that delay of the onset of postconditioning by only 1 minute aborts protection in rabbit hearts.28
MPT is a central mechanism of IR-induced death.20
S prevented CA/CPR-induced acceleration of Ca2+
-induced mitochondrial swelling, a measure of MPT, in LV mitochondria. The beneficial effect of Na2
S was associated with attenuated oxidative stress indicated by decreased serum hydrogen peroxide levels (see ). Oxidative stress has been shown to trigger MPT.29
These observations suggest that the cardioprotective effects of Na2
S may be mediated by inhibition of MPT.
Cardioprotective effects of endogenous H2
S after IR have been suggested by studies showing deleterious effects of CGL inhibitors 30
and protective effects of CS-CGLtg 13
in myocardial IR. Our observation that CS-CGLtg mice had a markedly shorter CPR time to ROSC suggests endogenous H2
S promotes the recovery of myocardial function after CA/CPR. Although CS-CGLtg mice also had improved neurological function and survival rate 24h after CA/CPR, these findings may be due to the shortened CPR time to ROSC, as well as possible neuroprotective effects of endogenously-produced H2
S. Nonetheless, these observations support an important salutary effect of endogenous H2
S production on the outcomes of CA/CPR.
Improved myocardial function in Na2
S-treated mice was associated with increased phosphorylation of Akt and NOS3 in LV and elevated serum levels of nitrite plus nitrate combined compared to vehicle-treated mice 15 min after CA/CPR. In contrast, delayed administration of Na2
S at 10 min after CPR did not increase the phosphorylation of Akt and NOS3 in LV and failed to improve the outcome of CA/CPR. Furthermore, NOS3 deficiency abrogated the protective effects of Na2
S on the outcome of CA/CPR. In a previous study, we found that NOS3 deficiency markedly worsened outcomes of CA/CPR, suggesting an salutary role of NOS3 on the outcome of CA/CPR.19
Taken together, these results suggest a critical role of NOS3/NO in the mechanisms responsible for the protective effects of Na2
S after CA/CPR.
In summary, the current study revealed robust protective effects of Na2S on the outcome of CA/CPR in mouse. Administration of Na2S 1 min before the start of CPR markedly improved myocardial and neurological function and survival 24h after normothermic CA/CPR. These observations, if extrapolated to human beings, may be highly clinically relevant because they suggest that Na2S administration can improve the outcome of cardiac arrest at the time of CPR when IV access is obtained. Further studies are warranted to elucidate the impact of exogenous and endogenous H2S on the longer term outcome of CA/CPR complicated by post-cardiac arrest syndrome.
Sudden cardiac arrest is one of the leading causes of death worldwide. Despite advances in cardiopulmonary resuscitation (CPR) methods, including the introduction of the automatic electrical defibrillator and therapeutic hypothermia, 60–80% of these arrests result in immediate death, and of the remaining, only about 5 percent are successfully resuscitated to the extent that they are returned to productive lives. Poor outcome from cardiac arrest is at least partly due to the lack of therapeutic possibilities. In this study, we examined effects of hydrogen sulfide, a gaseous molecule with multi-faceted protective properties, on the outcome of cardiac arrest and CPR in mice. We found that administration of sodium sulfide, a hydrogen sulfide donor, 1 min before CPR markedly improved neurological and myocardial function and survival after 8 min of cardiac arrest in mouse. These observations, if extrapolated to human beings, may be highly clinically relevant because they suggest that sodium sulfide administration can improve the outcome of sudden cardiac arrest at the time of CPR when intravenous access is obtained. Further studies are warranted to elucidate the impact of exogenous and endogenous hydrogen sulfide on the longer term outcome of cardiac arrest and CPR complicated by post-cardiac arrest syndrome.