右美托咪定预先给药对大鼠单肺通气时内质网应激诱导的细胞凋亡及c-Jun氨基末端激酶通路的影响

Effects of dexmedetomidine pretreatment on cell apoptosis induced by endoplasmic reticulum stress and c-Jun N-terminal kinase pathway during one-lung ventilation in rats

目的 评价右美托咪定预先给药对大鼠单肺通气（OLV）时内质网应激（ERS）诱导的细胞凋亡及c-Jun氨基末端激酶（JNK）通路的影响.方法 雄性SD大鼠60只,采用随机数字表法分为6组（n=10）：假手术组（Sham组）、OLV组、OLV＋阿替美唑（α2受体拮抗药）处理组（AD组）、OLV＋阿替美唑＋右美托咪定处理组（DEX＋ AD组）、OLV＋右美托咪定低剂量组（DEX-L组）和OLV＋右美托咪定高剂量组（DEX-H组）.Sham组大鼠行双肺通气2.5h.OLV组行右肺OLV 2.0h、双肺通气0.5h.DEX-L组和DEX-H组分别于OLV前1h静脉输注右美托咪定2.5 μg＊ kg^-1·h^-1、5.0μg·kg^-1·h^-1,1 h完成.AD组于OLV前1h静脉注射阿替美唑250 μg/kg.DEX＋AD组于静脉输注DEX 5.0 μg·kg^-1 h^-1即刻静脉注射阿替美唑250 μg/kg.于双肺通气0.5h时处死大鼠取左肺组织,测定肺组织湿/干重比（W/D）,光镜下观察肺组织病理改变并测定肺泡损伤率（IAR）,电镜下观察肺组织超微结构,TUNEL法检测肺组织细胞凋亡,RT-PCR和Western blot法分别测定葡萄糖调节蛋白78 （GRP78） mRNA及蛋白、JNK mRNA、磷酸化JNK （p-JNK）蛋白表达水平.结果 与Sham组比较,OLV组、AD组和DEX＋AD组W/D、AI和IAR升高（P＜0.01）;与OLV组、AD组和DEX＋AD组比较,DEX-L组和DEX-H组W/D、AI和IAR降低（P＜0.01）.OLV组、AD组和DEX＋AD组肺组织结构均发生明显损伤,而DEX-L组和DEX-H组肺组织结构损伤则明显减轻.OLV组、AD组和DEX＋AD组有大量肺血管内皮细胞和肺泡上皮细胞发生凋亡,而经DEX干预后细胞凋亡则明显减少.与Sham组比较,OLV组、AD组和DEX＋AD组GRP78 mRNA和蛋白、JNK mRNA和p-JNK蛋白表达水平升高（P＜0.01）;与OLV组、AD组和DEX＋AD组比较,DEX-L组和DEX-H组GRP78 mRNA和蛋白、JNK mRNA和p-JNK蛋白表达水平降低（P＜0.01）.结论 右美托咪定预先给药对单肺通气时的肺脏具有保护作用,该作用的可能机制与其抑制JNK信号通路,减轻ERS诱导的细胞凋亡有关.

Objective To evaluate the effects of dexmedetomidine （DEX） on cell apoptosis induced by endoplasmic reticulum stress and c-Jun N-terminal kinase （JNK） pathway during one-lung ventilation （OLV） in rats.Methods Sixty male Sprague-Dawley rats were randomly allocated into 6 groups （n =10 each）：sham operation group （Sham group）,OLV group,OLV ＋ atipamezole （α2 receptor antagonist） group （AD group）,OLV ＋ atipamezole ＋ DEX group （DEX＋AD group）,OLV ＋ low-dose DEX group （DEX-L group） and OLV ＋ high-dose DEX group （DEX-H group）.The animals were anesthetized with 10% chloral hydrate 4.5 ml/kg,tracheostomized and mechanically ventilated.Bilateral lungs were ventilated for 2.5 h in Sham group.The right lung was ventilated for 2.0 h followed by 0.5 h two-lung ventilation in OLV group.In DEX-L and DEX-H groups,DEX was infused intravenously for 1 h at a rate of 2.5 μg ＆#183; kg^-1 ＆#183; h^-1 and 5.0 μg ＆#183; kg^-1 ＆#183; h^-1,respectively,starting from 1 h prior to OLV.Atipamezole 250 μg/kg was injected intravenously at 1 h prior to OLV in AD group.Atipamezole 250 μg/kg was injected intravenously at the onset of DEX infusion （5.0 μg ＆#183; kg^-1 ＆#183; h^-1） in DEX＋AD group.The rats were sacrificed and left lungs were removed for determination of weight to dry lung weight ratio （W/D）,cell apoptosis in lung tissues （by TUNEL）,and expression of glucose-regulated protein 78 （GRP78） mRNA and protein,JNK mRNA and phosphorylated JNK （p-JNK） protein （by RT-PCR and Western blot）.Pathological changes of lungs were examined and the injured alveolus rate （IAR） was counted under light microscope.The changes in ultrastructure of lung tissues were observed under transmission electron microscope.Apoptosis index （AI） was calculated.Results W/D,AI and IAR were significantly higher in OLV,AD and DEX＋AD group than in Sham group,while lower in DEX-L and DEX-H groups than in OLV,AD and DEX＋AD groups.The pathological changes of the structure of lung tissues were observed in OLV,AD and DEX＋AD groups,while the pathological changes were significantly alleviated in DEX-L and DEX-H groups.In OLV,AD and DEX ＋ AD groups,there was apoptosis in lots of pulmonary vascular endothelial cells and alveolar epithelial cells,while cell apoptosis was significantly reduced after administration of DEX.The expression of GRP78 mRNA and protein,JNK mRNA and p-JNK protein was significantly higher in OLV,AD and DEX＋AD groups than in Sham group,and lower in DEX-L and DEX-H groups than in OLV,AD and DEX ＋AD groups.Conclusion DEX pretreatment can protect lungs during OLV,and inhibited JNK signaling pathway and reduced cell apoptosis induced by endoplasmic reticulum stress may be involved in the mechanism.