| Tourniquets are often used clinically to reduce intraoperative blood loss and create bloodless operating fields.However,tourniquet leads to a significant acute limb ischemia reperfusion?I/R?injury,which not only damages the primary ischemic part of the limb,but also causes remote multiple organs injury.In addition to I/R injury,tourniquet inflation can induce a hyperdynamic response.Although tourniquet-induced I/R injury and hyperdynamic response have been recognized for several years,the complex pathophysiologic mechanism and therapeutic interventions are not completely understood.Lots of research has shown that skeletal muscle injury,oxygen free radical lipid oxidation and neutrophils mediated inflammatory reaction plays an important role in the limb ischemia reperfusion induce local and remote organ dysfunction.Perioperative neurocognitive disorders?PNDs?are a common postoperative complication in geriatric patients,which refers to the changes in cognitive functions such as attention deficit and memory loss that occur after surgery in patients with no preoperative mental disorder.PNDs has been shown to be associated with higher healthcare costs,long-term disability,and even increase mortality.Although the pathogenic mechanisms underlying PNDs remain elusive,systemic acute inflammation and pro-inflammatory cytokines released after surgery are considered as a critical factor to its development.Dexmedetomidine?Dex?,a new type of highly selective a2adrenergic receptor agonist.As an agent that provides sedative,anxiolytic,and analgesic effect,which has been appling on surgical patients as adjuvant anesthetic.Several researches have shown that Dex has a protective effect against I/R injury in multiple organs.Currently,protection of Dex mainly focus on brain,heart,liver and other organs,however,whether Dex influences tourniquet-induced I/R injury remains unclear.Oxycodone?Oxy?,an?and?opioid receptor agonist,is increasingly used worldwide to treat variety of painful disorder.Recently,several studies suggested that Oxy not only provided potent analgesia but also had property of relieving inflammatory response.To date,no study has investigated the anti-inflammatory effects of Oxy in limb ischemia-reperfusion.Perioperative medicine is the development direction of anesthesiology.It is not enough for anesthesiologists to only pay attention to the safety of patients during surgery.Anesthesiologist should optimize the anesthesia program,try to prevent perioperative complications and improve the outcome of patients.In summary,the purpose of the study was to evaluate the effects of dexmedetomidine and oxycodone on local and brain injury caused by lower limb ischemia reperfusion and to explore the relevant mechanisms.Objective:In the present study,first,we compared the effects of dexmedetomidine and oxycodone in patients undergoing limb ischemia-reperfusion.Second,to investigate the tourniquet induced local and brain injury through the animal model of lower limb ischemia-reperfusion and evaluate the preventive effect of dexmedetomidine and oxycodone preconditioning on the pathophysiological process.Methods:Firstly,patients undergoing unilateral lower extremity surgery were randomly assigned to control?ischemia-reperfusion,I/R?group,dexmedetomidine?Dex?group,andoxycodone?Oxy?group.Tourniquet-induced hemodynamic parameters changes between groups were compared.The serum concentration of malondialdehyde?MDA?,superoxide dismutase?SOD?,tumor necrosis factor-a?TNF-a?,interleukin-6?IL-6?,fatty acid binding protein 3?FABP3?,endothelin-1?ET-1?,and brain derived neurotrophic factor?BDNF?were measured.Secondly,animal of tourniquet-induced acute hind limb I/R injury was established in C57BL6mice.For experiments,mice were randomized assigned to sham group,I/R group,Dex group and Oxy group,real-time microcirculation imaging system was used to assess limb perfusion,and morphological changes of gastrocnemius muscle were observed by hematoxylin-eosin?HE?staining,Masson trichrome staining and electron microscopy.Serum TNF-a level,the gastrocnemius muscle contractile force and ATP concentration were examined.In addition,TLR4,NF-k B,SIRT1 and PGC-1a expression within the skeletal muscles were detected by western blot.Finally,behavioral alterations were assessed with Morris water maze?MWM?test.The NF-k B expression in the hippocampus in different groups was observed by immunofluorescence.In addition,TLR4,NF-k B,CD68,TNF-a,CD206,IL-10 and NR2B in hippocampus were detected by western blot.Besides,the effects of dexmedetomidine and oxycodone on the s EPSC of hippocampal CA1 neurons in young mice were observed by electrophysiological hippocampal slice technique.Through the above experimental results,we observed whether dexmedetomidine and oxycodone pretreatment would attenuate tourniquet-induced local skeletal muscle damage and brain injury,and explored the underlining molecular mechanisms.Results:1 Compare the protective effects of dexmedetomidine and oxycodone in patients undergoing limb ischemia-reperfusion.In the control group,tourniquet caused significant increases in systolic arterial pressure?SAP?,mean arterial pressure?MAP?,diastolic arterial pressure?DAP?,and rate pressure product.Compared with the Oxy,Dex significantly decreased heart rate?HR?.Both Dex and Oxy lower SAP compared with the control group.No significant difference was observed in DAP between Dex and Oxy.The levels of MDA,TNF-a,IL-6,FABP3,and ET-1 were significantly higher,while the SOD and BDNF significantly lower compared to baseline in the I/R group,but the variation range of those agents was significantly smaller in Dex and Oxy groups,furthermore,the measured values were comparable between the two group.2 Protective effect of dexmedetomidine and oxycodone on local limb damages following hindlimb ischemia reperfusion.2.1 Effects of dexmedetomidine and oxycodone on blood flow recovery following hindlimb ischemia reperfusion.After placing the tourniquet,the blood perfusion of the ischemia limb was significantly reduced,and the ratio of the average blood flow of the left and right limbs?L/R ratio?was significantly decreased.After reperfusion,compared with the control group,the blood flow of ischemia-reperfusion group increased slightly,and no difference was found between the two groups.Compared with the ischemia-reperfusion group,the blood flow after dexmedetomidine and oxycodone pretreatment showed an increasing trend,and no difference was found between the three groups.2.2 To investigate the mechanism of dexmedetomidine and oxycodone alleviate inflammatory response following hindlimb ischemia reperfusion.Compared to the normal muscle morphology,gastrocnemius muscle showed obvious damage after limb ischemia reperfusion.Compared to the I/R group,dexmedetomidine and oxycodone pretreatment significantly reduced the degree of gastrocnemius muscle ischemia-reperfusion injury and inhibited the expression of TLR4/NF-k B in the gastrocnemius muscle,furthermore,the measured values were comparable between the two group.2.3 Dexmedetomidine and oxycodone improve skeletal muscle mitochondrial function following hindlimb ischemia reperfusion.Compared to the normal skeletal muscle,the ATP level and the contractility of the skeletal muscles showed obvious decreased after limb ischemia reperfusion.Compared to the I/R group,dexmedetomidine and oxycodone pretreatment significantly increased the ATP level and the contractility of the skeletal muscles.Besides,SIRT1 and PGC-1a level were significantly reduced in gastrocnemius muscle after I/R,while dexmedetomidine and oxycodone pretreatment could observably increase these proteins expression.3 Effects of dexmedetomidine and oxycodone on cerebral injury following hindlimb ischemia reperfusion.3.1 Dexmedetomidine and oxycodone on the function of learning and memory function following hindlimb ischemia reperfusion.The study showed that the cognitive ability of mice can be impaired by tourniquet induced acute limb ischemia reperfusion,and these changes were significantly prevented by dexmedetomidine preconditioning while oxycodone pretreatment had no effect.Compared to the I/R group,dexmedetomidine and oxycodone pretreatment could decreased levels of TNF-a proteins in serum.Besides,dexmedetomidine and oxycodone preconditioning significantly decreased the expression of NF-k B immunoreactive cells in hippocampus.In addition,dexmedetomidine and oxycodone pretreatment could observably decreased TLR4,NF-k B,CD68 and TNF-a overexpression in hippocampus compared to that in I/R group.For CD206 and IL-10,there was no significant difference by each factor.3.2 Effects of dexmedetomidine and oxycodone on hippocampal neuronal s EPSC and expression of NR2B following hindlimb ischemia reperfusion.Compared with the baseline,the frequency and amplitude of s EPSC in hippocampal CAl neurons were significantly decreased after dexmedetomidine treatment,and the electrophysiological characteristics recovered after wash out.However,oxycodone had no effect.Compared with normal mice,the expression of NR2B in hippocampus significantly increased after limb ischemia-reperfusion.Dexmedetomidine preconditioning significantly reduced NR2B expression,while oxycodone had no effect.Conclusions:1 The present study indicated that patients undergoing lower-extremity surgery with tourniquet use showed a hyperdynamic response and elevated proinflammatory cytokines,and even suffered from remote multiple-organ injury.Compared with Dex,Oxy was no inferior to mitigate tourniquet-induced hyperdynamic response,ameliorate the inflammatory reaction,and protect remote multiple organs in lower extremity surgery patients.2 Tourniquet induced acute limb I/R results in morphological and functional impairment in mouse skeletal muscle.Pretreatment with dexmedetomidine or oxycodone,partly through inactivating of the TLR4/NF-k B pathway inhibits inflammatory response and mediating of the SIRT1/PGC-1a pathway protects mitochondrial function,attenuates the skeletal muscle from acute I/R injuries.3 The learning and memory ability of mice can be impaired by tourniquet induced acute limb ischemia reperfusion.Pretreatment with Dex,partly through inhibiting of microglia transform to the pro-inflammatory M1polarization state and inactivating of the TLR4/NF-k B pathway reduces inflammatory response,furthermore,reducing the hippocampal excitatory neurotransmitter glutamate release in presynaptic membrane and inhibition of glutamate receptor?NR2B?expression,attenuates the cognitive disorders in mice following tourniquet use.Oxycodone pretreatment can also inhibit the transformation of microglia into proinflammatory microglia?M1?and reduce the inflammatory response through the TLR4/nf-kb pathway,but it has no obvious improvement in learning and memory,which may be due to it is has no effect on the expression of hippocampal glutamate receptor?NR2B?. |
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