The Neuroprotective Effect Of Sevoflurane Postconditioning And Its Mechanisms Against Spinal Cord Ischemia/reperfusion Injury In Rabbits

Temporary aortic occlusion in surgical repair of thoracic and thoracoabdominal aneurisms can cause different extent of spinal cord ischemia which leads to the nerve functional impairment, and then come to acute or delayed paraplegia. Thereby it calamitously affects the surgical outcomes, results in a great of the burden of psycho-economy and remains a persistent clinical problem. To date, numerous clinical and laboratory studies, including our studies, in attempt to decrease the risk of this devastating complication have been reported. Despite various refinements in surgical techniques and developments in adjunctive measures, including hypothermia, cerebrospinal fluid drainage, ischemic and chemicals preconditioning, have been used for reducing the impact of the ischemia and preserving spinal cord function, spinal cord ischemia is present in approximately 3% to 18% of patients, depending on the type of aneurysm and other combined diseases. Therefore, other novel measures to protect against spinal cord ischemic injury require further investigation. Postconditioning, defined as the process of limiting ischemia reperfusion injury-induced damage by application of repetitive short ischemic windows during early reperfusion, provides a new option and good ideal for neurprotection. Clinically, postconditioning is well suited for vaso-occlusive emergencies and elective surgical settings involving clamping of arteries and subsequent release. Our previous studies demonstrated that ischemic postconditioning could alleviate spinal cord ischemia/reperfusion (I/R) injury in rabbits through induction of heat shock proteins (HSP) expression and inhibition of overdose production of oxygen free radicals (OFR) during reperfusion. Recent evidences have opened exciting research avenues into pharmacotherapeutic agents mimicking postconditioning-induced protection.Sevoflurane, a novel inhalation anesthetic, is wildly applied in clinic anesthesia because of its rapid induction, stable maintainence and complete recovery. Moreover, studies have demonstrated that sevoflurane protected myocardial ischemia/reperfusion (I/R) injury. However, it is unknown whether postconditioning with sevoflurane could alleviate spinal cord I/R injury. Thus, in the present study, using the model of the spinal cord ischemia/reperfusion (I/R) injury in rabbits induced by infrarenal aorta occlusion, we investigated that the protective effect of sevoflurane postconditioning on spinal cord ischemia/reperfusion injury in rabbits and its mechanisms including the role of oxygen free radicals, antioxidase, mitochondrial ATP-sensitive potassium channel.Experiment 1. Sevoflurane postconditioning induces neuroprotection against spinal cord ischemia/reperfusion injury in rabbits Objective To investigate whether postconditoning with sevoflurane could alleviate spinal cord I/R injury in rabbits. Methods Forty-eight male New Zealand white rabbits were randomly assigned to six groups (n=8 each). Animals in the sham group only underwent sham-operation. Animals in the control group underwent spinal cord ischemia for 20 min without postconditioning. Animals in O₂ postconditioning group (Group O₂) inhaled 100% O₂ at 5 min before reperfusion and last for 13 min. Animals in sevoflurane postconditioning groups (Group Sevo 0.5, Sevo 1.0 and Sevo 1.5) inhaled 0.5, 1.0, 1.5 minimum alveolar concentration (MAC) sevoflurane in 100% O₂ at 5 min before reperfusion for 10 min, and then inhaled 100% O₂ for 3 min to wash out the remaining sevoflurane. Forty-eight hours after reperfusion, hind-limb motor function and histopathology of the spinal cord were assessed in a blinded fashion. Results (1) The neurologic and histopathologic outcomes in the sevoflurane postconditioning groups were better than those in the control group (P<0.05). (2) The histopathologic outcomes in Sevo 1.0 group were better than that in Sevo 0.5 and Sevo 1.5 groups (P<0.05). Conclusion Postconditoning with sevoflurane could alleviate spinal cord I/R injury in rabbits. Postconditioning with 1.0 MAC sevoflurane offers better neuroprotective effect.Experiment 2. Effect of 2-mercaptopropionylglycine on the neuroprotection of sevoflurane postconditioning against spinal cord ischemia/reperfusion injuryObjective To investigate whether the benefical effect induced by sevoflurane postconditioning against spinal cord I/R is depend on oxygen free radicals in rabbits. Methods Thirty-six male New Zealand White rabbits were randomly assigned to six groups (n=6 each). Animals in O₂ and Sevo groups received deionized water (0.5 ml·kg^-1·min^-1 intravenously) 10 min before postconditioning with 100% O₂ and 1.0 MAC sevoflurane for 20 min, respectively. In the MPG+Sevo and MPG+O₂ groups, 2% 2-mercaptopropionylglycine (MPG, a potent oxygen free radical scavenger, dissolved in deionized water) was administered (0.5 ml·kg^-1·min^-1 intravenously) at the same time point. In the Sevo+MPG(Sevo+MPG)and O₂+MPG(O₂+MPG)groups, 2% MPG was administered (0.5 ml·kg^-1·min^-1 intravenously) 10 min after postconditioning with 100% O₂ and 1.0 MAC sevoflurane for 20 min, respectively. Spinal cord ischemia was induced by an infrarenal aorta clamping for 20 min. Forty-eight hours after reperfusion, hind-limb motor function and histopathology of the spinal cord were assessed in a blinded fashion. Results (1) The neurologic and histopathologic outcomes in the Sevo and Sevo+MPG groups were better than those in the others groups (P<0.05). (2) There were no significant diffierences in hind-limb motor function scores and the number of normal neurons at the anterior spinal cord of animals in the Sevo group in comparison with the Sevo+MPG group. Conclusion 2-mercaptopropionylglycine, a scavanger of reactive oxygen species, completely abolished the neuroprotection induced by sevoflurane postconditioning, which indicated that sevoflurane postconditioning against spinal cord I/R injury via release of oxygen free radicals in rabbits.Experiment 3. Effect of sevoflurane postconditioning on antioxidant enzymes activies and malondialdehyde content of spinal cord tissue during reperfusion in rabbitsObjective To investigate the effect of sevoflurane postconditioning on antioxidant enzymes activities and malondialdehyde content of spinal cord tissue after spinal cord I/R injury in rabbits. Methods Eighty male New Zealand rabbits were randomly divided into 4 groups (n=20 each). Sham group: the procedure was the same as the I/R group without aortic occlusion; I/R group: rabbits underwent occlusion of the infrarenal abdominal aorta for 20 min, followed by reperfusion; O₂ group (100% O₂ postconditioning) and Sevo group (sevoflurane postconditioning): rabbits were subjected to 100% O₂ and 1.0 MAC sevoflurane postconditioning, respectively, other procedures were the same as in the I/R group. After the neurological outcomes were assessed, the spinal cord tissue (L5⁷) was sampled at 1 h, 6 h, 24 h and 48 h after reperfusion (n=5 at each time point), and frozen at -70℃for the determination of superoxide dismutase (SOD), catalase (CAT) activities and malondialdehyde (MDA) content by spectrophotometric methods. The neurological outcomes were assessed before sampled at 6h, 24h and 48h after reperfusion, respectively. Results (1) The motor function scores of the Sevo group were significantly better than those of I/R group at 6h, 24h and 48h after reperfusion (P<0.05). (2) SOD activities of spinal cord tissue were significantly higher compared with those in I/R group at 1 h ,6 h and 24 h after reperfusion (P<0.05). CAT activities of spinal cord tissue in Sevo group were significantly higher compared with those in I/R group at 1 h and 6 h after reperfusion (P<0.05). No significant difference was observed between Sevo and I/R groups at corresponding time points on the activities of glutathione peroxidase (GSH-px). (3) Compared with the I/R group, Sevo group produced a significant reduction of the MDA content at 6 h , 24 h and 48 h after reperfusion (P<0.05). Conclusion The protective effects of sevoflurane postconditioning against spinal cord ischemic/reperfusion injury is mediated, at least partially, by up-regulating the activities of SOD and CAT of spinal cord tissue during early reperfusion phase.Experiment 4. Effect of 2-mercaptopropionylglycine on antioxidant enzymes activies and malondialdehyde content of spinal cord after sevoflurane postconditioningObjective To investigate the effect of 2-mercaptopropionylglycine (MPG) on the antioxidant enzymes activies and malondialdehyde content of spinal cord tissue after sevoflurane postconditioning. Methods Twenty-four male New Zealand rabbits were randomly divided into 4 groups (n=6 each). All animals underwent spinal cord ischemia for 20 min. Animals in control group received deionized water (0.5 ml·kg^-1·min^-1 intravenously) 15 min before reperfusion for 20 min. Sevo groups received deionized water (0.5 ml·kg^-1·min^-1 intravenously) 10 min before postconditioning with 1.0 MAC sevoflurane for 20 min. In the MPG+Sevo group, 2% MPG was administered (0.5 ml·kg^-1·min^-1 intravenously) 10 min before postconditioning with 1.0 MAC sevoflurane for 20min. In the MPG group, 2% MPG group was administered (0.5 ml·kg^-1·min^-1 intravenously) 15 min before reperfusion. After the neurological outcomes were assessed at 6 h after reperfusion, the spinal cord tissue was sampled at 6 h after reperfusion, and frozen at -70℃for the determination of antioxidant enzyme activities and malondialdehyde (MDA) content by spectrophotometric methods. The neurological outcomes were assessed before sampled at 6h after reperfusion. Results (1) The motor function scores of the Sevo group were significantly better than those of the control group, MPG+Sevo group and MPG group at 6 h after reperfusion (P<0.05). (2) Activities of superoxide dismutase (SOD) and catalase (CAT) of spinal cord tissue in Sevo group were significantly higher compared with those in the control group, MPG+Sevo group and MPG group 6 h after reperfusion.(P<0.05). (3) Compared with the others groups, Sevo group produced a significant reduction of the MDA level at 6h after reperfusion(P<0.05). Conclusion MPG administration before each session of sevoflurane postconditioning completely abolished the increase of CAT and SOD activities and the reduction of the MDA level induced by sevoflurane postconditioning.Experiment 5. Effect of 5-hydroxydecanoate on neuroprotection of sevoflurane postconditioning against spinal cord ischemia/reperfusion injuryObjective To investigate whether the neuroprotective effectS induced by sevoflurane postconditioning against spinal cord I/R is depend on mitochondrial ATP-sensitive potassium (KATP) channels in rabbits. Methods Thirty-two male New Zealand rabbits were randomly divided into 4 groups (n=8 each). Animals in O₂ and Sevo groups received saline (2 ml/kg intravenously) 10 min before postconditioning with 100% O₂ and 1.0 MAC sevoflurane, respectively. In the 5-HD+Sevo and 5-HD+O₂ groups, 10% 5-hydroxydecanoate (5-HD, a specific mitochondrial KATP channel blocker, dissolved in saline) was administered (2 ml/kg intravenously) at the same time point. Forty-eight hours after reperfusion, hind-limb motor function and histopathology of the spinal cord were assessed in a blinded fashion. Results The neurologic and histopathologic outcomes in the Sevo group were better than those in the others groups (P<0.05). There were no significant diffierences in the neurologic and histopathologic outcomes among the O₂ group, 5-HD+Sevo group and 5-HD+O₂ group. Conclusion 5-hydroxydecanoate, a specific inhibitor of mitochondrial ATP-sensitive potassium channels, abolishes sevoflurane postconditioning-induced protective effects against spinal cord ischemia, suggesting postconditoning with sevoflurane alleviates spinal cord ischemia/reperfusion injury via the activation of mitochondrial ATP-sensitive potassium channels in rabbits.Results1. Postconditoning with 0.5,1.0,1.5 MAC sevoflurane could alleviate spinal cord ischemia/reperfusion injury in rabbits. The neuroprotective effects of postconditioning with 1.0 MAC sevoflurane is better.2. The protective effects of sevoflurane postconditioning against spinal cord ischemia injury via release of oxygen free radicals then up-regulating the activities of SOD and CAT in spinal cord during early reperfusion phase in rabbits.3. Postconditoning with sevoflurane could alleviate spinal cord ischemia reperfusion injury via the activation of mitochondrial ATP-sensitive potassium channels in rabbits.