Effects And Mechanisms Of Ghrelin On Brain Injury After Cardiac Arrest And Cardiopulmonary Resuscitation

[Objective] Postresuscitation brain injury continues to be the leading cause of morbidity and mortality in patients after CA and CPR. Recently, neuroprotection by ghrelin, a brain-gut peptide, has been widely proven in a series of diseases. This study was performed to assess the effects of ghrelin on post-resuscitation brain injury in a rat model of cardiac arrest and to explore the underlying mechanisms.[Methods] Section A Healthy adult Sprague-Dawley rats were allocated to (1)sham group:rats were subjected to sham operation; (2)vehicle group:rats were subjected to 6 min cardiac arrest induced by transcutaneous electrical epicardium stimulation and resuscitated successfully. Vehicle (saline) was injected immediately after return of spontaneous circulation (ROSC); (3)ghrelin group:rats were subjected to 6 min cardiac arrest induced by transcutaneous electrical epicardium stimulation and resuscitated successfully. Ghrelin (80?g/kg) was injected blindly immediately after ROSC. A tape removal test was performed to evaluate neurological function at 24 h,48 h and 72 h after ROSC. Then brain tissues were harvested and coronal brain sections were analyzed by HE staining for neuronal viability and TUNEL staining for apoptosis in hippocampal CA1 sectors. Section B Healthy adult Sprague-Dawley rats were allocated to(1)sham group; (2)vehicle group; (3)ghrelin group. Rats were killed and brain tissues were harvested 6 h after ROSC for (1)western blot and immunohistochemistry staining assay to evaluate hippocampal levels of apoptosis related factors including cleaved caspase-3, bax and bcl-2. (2)ELISA to evaluate hippocampal levels of inflammation related factors including IL-1? and TNF-?. (3)biochemical assay to evaluate hippocampal levels of oxidative stress indictors including MDA and SOD, and western blot and immunohistochemistry staining assay to evaluate hippocampal uncoupling protein-2 level. (4)functional analysis on mitochondrial ATP and ROS synthesis, and biochemical assay to evaluate cortex levels of MDA, SOD activity and ATP, and western blot to evaluate cortex uncoupling protein-2 level. A JC-1 assay was performed to assess the mitochondrial membrane potential level. Transmission electron microscopy was performed to evaluate mitochondrial morphology.[Result] (1) Animals treated with ghrelin had improved neurological performances, reduced neuronal injury and inhibited neuronal apoptosis compared with the vehicle group. (2) ghrelin treatment was associated with a decrease in caspase-3 up-regulation and an increased Bcl-2/Bax ratio in hippocampus. (3) ghrelin treatment was associated with a reduction in hippocampal TNF-? and IL-1? levels. (4)ghrelin treatment was associated with a reduction in hippocampal MDA content and an up-regulation in SOD activity. (5) ghrelin treatment was associated with an increase in hippocampal uncoupling protein 2 (UCP-2) expression. (6)ghrelin treatment was associated with improved mitochondrial ATP synthesis and reduced the ROS leakage by mitochondria isolated from cortex. (7)ghrelin treatment was associated with a reduction in MDA content and an up-regulation in SOD activity and ATP content in the cortex. (8) ghrelin treatment was associated with an increase UCP-2 expression in the cortex. (9)ghrelin caused an increase in mitochondrial membrane potential. (10)ghrelin markedly improved mitochondrial morphology compared with the vehicle animals.[Conclusion] Our results suggest that ghrelin treatment attenuated post-resuscitation brain injury in rats, possibly via regulation of apoptosis, inflammation, oxidative stress and mitochondrial function.