| Backgrouds and objective Brain is the most oxygen hungry organ and the most vulnerabl organ to hypoxia. The incidence of perioperative stroke is 0.08～2% in general surgery but up to 4～13% in cardiopulmonary bypass operation and carotid endarterectomy surgery, which may caused by hemodynamic instability, hypercoagulable state and organ transplantation during perioperative period. The secondary injury after cerebral ischemia is the major cause of death and neurological dysfunction perioperative period, and gradually taken seriously. Dexmedetomidine (Dex), a highly selective α2-adrenoceptor agonist, is widely used for sympatholysis, sedation, analgesia and anxiolysis during general anesthesia as anaesthetic-adjuncts and in intensive care unit (ICU). Its protective effect on vita organs such as heart and brain during ischemia reperfusion (I/R) has been studied extensively recent years. Phosphocreatine (PCr), an important substances involved in cellular energy metabolism, compensate for ATP consumption in vital movements. Exogenous phosphocreatine, as a safe and high-efficiency conpound, is widely used in card iopulmonary bypass surgery and neonatal hypoxic-ischemic encephalopathy, can improve the prognosis of patients. Cerebral protective effects of dexmedetomidine and phosphocreatine alone pretreated on cerebral I/R injury have been reported. However, the individual effect of dexmedetomidine and phosphocreatine posttreated at the onset of reperfusion is still unclear. Moreover, whether the neuroprotective efficacy of dexmedetomidine and phosphocreatine in combination is better than either alone needs to be elucidated. Therefore, the present study was designed to clarify the above questions and investigate potential mechanisms in a rat model of focal cerebral I/R injury.Materials and methods Adult male Sprague-Dawley rats (SPF grade), weighing 220-260g, were used in the study. I/R was induced by intraluminal middle cerebral artery occlusion of 90 minutes followed by a 24 hours reperfusion. Ninety rats were randomly allocated to sham group (Group Sham) and I/R group. I/R group was further divided into four subgroups according to the corresponding treatments:Group I/R, Group Dex, Group PCr and Group Dex+PCr. The dose was dexmedetomidine 9μg.kg-1, phosphocreatine 180mg.kg-1 and the combination, respectively. Desired dose was calculated and dissolved in 2ml normal saline. The intravenous infusion was started immediately at onset of reperfusion at a contant speed of 8 ml.h-1. All treatments were given intravenously at the onset of reperfusion in 15 minutes. Group Sham and Group I/R received an intravenous infusion of equal amounts normal saline at the same time. Twenty-four hours after the reperfusion, neurological deficit score and magnetic resonance scan were performed to confirm successful modeling. Blood samples were taken to measure serum concentrations of malonaldehyde and 4-hydroxynonenal. Brains were harvested to estimate infarct volume and cerebral pathology by optical and electron microscopy. Expressions of NeuN and cleaved caspase-3 were detected by immunohisto chemistry. Neuronal apoptosis was further assessed by the expression of cleaved caspase-3 and co-localization of cleaved caspase-3 and NeuN by immunofluorescence, apoptotic index was calculated.Results There was no neurological deficiency and cerebral infarction in MRI and TTC staining detected in Group sham. Rats in other groups showed different degrees of neurological deficiency and cerebral infarction (P<0.05), which confirmed cerebral I/R model was made successfully. Neurological deficit score and infarct volume: Compared with Group I/R, dexmedetomidine and phosphocreatine significantly reduced neurological deficit score (P<0.05), infarct volume (P<0.05). Compared with Group Dex and Group PCr, neurological deficit score and infarct volume was futher decreased in Group Dex+PCr (P<0.05). Serum MDA and 4-HNE concentrations:I/R significantly increased the contents of the two specific products of oxidative stress. Compared with Group I/R, serum MDA and 4-HNE concentrations were decreased in Group Dex and Group PCr (P<0.05). Compared with Group Dex and Group PCr, the concentrations were futher decreased in Group Dex+PCr (P<0.05). Cerebral pathological changes:Under the light microscope, neurons of cortex and hippocampus are arranged in neat rows in Group Sham, the hemispheric structure was sparse because so many cells fosing caused by I/R. The surviving cells were shrinking or edematous in Group I/R. In treated groups, more cells survived with complete membrane and clear nuclei. Under electron microscope, the structure of blood brain barrier, neurons and abiomembrane was complete in Group Sham, I/R induced uncomplete blood brain barrier, tissue edema and apoptotic neurons; In dexmedetomidine-and phosphocreatine-treated groups, the damages were alleviated obviously, in the co-treated group, the structure was more complete. The expression of NeuN and caspase-3 I/R caused the neuronal apoptosis as the increase of caspase-3 and decrease of NeuN (P<0.05). Compared with Group I/R. While the expression of caspase-3 was greatly reduced, the expression of NeuN was increased in Group Dex and Group PCr (P<0.05). Compared with in Group Dex and Group PCr, the expression of caspase-3 was further reduced and the expression of caspase-3 was further increased in Group Dex+PCr (P<0.05). Neuronal apoptosis The apoptotic index(AI, defined as percent of caspase-3 positive cells in NeuN positive cells) was calculated for comparision. Compared with Sham, AI in Group I/R was greatly increased (P<0.05). Compared with Group I/R, AI was decreased in Group Dex and Group PCr (P<0.05). Compared with Group Dex and Group PCr, AI was further decreased in Group Dex+PCr (P<0.05).Conclusion The results show significant decreases in neurological deficits, ischemic infarction and cerebral damages of dexmedetomidine or phosphocreatine as post-ischemia treatment, and both in combination has greater protective effects than each alone. However, suppressions of oxdative stress and neuronal apoptosis are likely to account for the neuroprotective effects. |
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