舒芬太尼调节cAMP/PKA/CREB通路减轻HIBD新生大鼠神经元损伤

Sufentanil alleviates neuronal damage in HIBD neonatal rats by regulating cAMP/PKA/CREB pathway

目的探讨舒芬太尼通过调节环磷酸腺苷(cAMP)/蛋白激酶A(PKA)/cAMP反应元件结合蛋白(CREB)信号通路减轻缺氧缺血性脑损伤(HIBD)新生大鼠神经元损伤的机制。方法采用Rice-Vannucci法构建HIBD新生大鼠模型,随机分为:模型组、舒芬太尼低剂量组、舒芬太尼高剂量组、SQ22536组、舒芬太尼高剂量+SQ22536组,每组10只,另取10只新生大鼠设为假手术组,以舒芬太尼和SQ22536分组干预后,以避暗实验检测大鼠认知功能;以伊文思蓝实验检测大鼠血脑屏障功能;以H-E染色检测各组大鼠脑皮质与海马神经元病理形态;以尼氏染色检测各组大鼠脑皮质与海马神经元数量;以试剂盒检测各组大鼠血清炎性因子前列腺素2(PGE2)、诱导型一氧化氮合酶(iNOS)、抗氧化因子总抗氧化能力(TAC)、超氧化物歧化酶(SOD)水平及脑组织cAMP水平;以免疫印迹法检测各组大鼠脑组织cAMP/PKA/CREB通路相关表达。结果与假手术组比较,模型组大鼠脑皮质与海马神经元均病理损伤严重,步入潜伏期、脑皮质与海马神经元数量、血清TAC与SOD水平、脑组织cAMP水平及p-PKA/PKA、p-CREB/CREB显著降低(P<0.05),犯错次数、伊文思蓝含量、血清PGE2与iNOS水平显著升高(P<0.05)。与模型组比较,舒芬太尼低剂量组、舒芬太尼高剂量组大鼠脑皮质与海马神经元病理损伤均减轻,步入潜伏期、脑皮质与海马神经元数量、血清TAC与SOD水平、脑组织cAMP水平及p-PKA/PKA、p-CREB/CREB均升高(P<0.05),犯错次数、伊文思蓝含量、血清PGE2与iNOS水平均降低(P<0.05)。结论舒芬太尼可通过激活cAMP/PKA/CREB信号而抑制炎症,增强抗氧化活性,从而减轻HIBD新生大鼠神经元损伤,改善其认知和血脑屏障功能。

Objective To investigate the mechanism of sufentanil in alleviating neuronal damage in neonatal rats with hypoxic-ischemic brain damage(HIBD)by regulating the cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA)/cAMP responsive element binding protein(CREB)signal pathway.Methods The neonatal rat model of HIBD was established by Rice-Vannucci method and randomly separated into model group,low-dose sufentanil group,high-dose sufentanil group,SQ22536 group,high-dose sufentanil+SQ22536 group,with 10 rats in each group,another 10 neonatal rats were set as sham operation group,after the intervention of sufentanil and SQ22536,the cognitive function of rats was tested by dark avoidance test.Evans blue test was applied to detect the blood-brain barrier function in rats.H-E staining was applied to detect the pathological morphology of neurons in cerebral cortex and hippocampus of rats in each group.The number of neurons in cerebral cortex and hippocampus of rats in each group was detected with Nissl staining.The levels of serum inflammatory factor prostaglandin 2(PGE2),inducible nitric oxide synthase(iNOS),total antioxidant capacity(TAC),superoxide dismutase(SOD)and cAMP in brain tissue were measured with the kits.The expression of cAMP/PKA/CREB pathway was detected with Western blot.Results Compared with the sham operation group,the cerebral cortex and hippocampus neurons in the model group were severely damaged,the step-through latency,number of neurons in cerebral cortex and hippocampus,the levels of serum TAC and SOD,the level of brain cAMP,p-PKA/PKA,and p-CREB/CREB decreased obviously(P<0.05),the number of mistakes,Evans blue content,the levels of serum PGE2 and iNOS increased obviously(P<0.05).Compared with the model group,the pathological damage of cerebral cortex and hippocampus neurons in the low-dose and high-dose sufentanil groups were alleviated,the step-through latency,number of neurons in cerebral cortex and hippocampus,the levels of serum TAC and SOD,the level of brain cAMP,p-PKA/PKA,and p-CREB/CREB all increased(P<0.05),the number of mistakes,Evans blue content,the levels of serum PGE2 and iNOS decreased(P<0.05).Conclusion Sufentanil can inhibit inflammation and enhance antioxidant activity by activating cAMP/PKA/CREB signal,thereby reducing neuronal damage in neonatal HIBD rats and improving their cognitive and blood brain barrier functions.