右美托咪定经海马CA1区PPARγ磷酸化在大鼠体外循环脑保护中的作用

The role of PPARγphosphorylation in the neuroprotective effect of dexmedetomidine through the hippocampal CA1 region during rat extracorporeal circulation

目的探讨PPARγ磷酸化在右美托咪定对大鼠体外循环脑保护中的作用。方法成年健康雄性SPF级SD大鼠36只,体重350~450 g,随机分为3组,每组12只。假手术组(S组):在戊巴比妥钠麻醉下行气管插管,右颈内静脉及双侧股动脉穿刺置管,不进行体外循环。体外循环组(C组):同S组麻醉穿刺置管,复制大鼠体外循环模型,进行体外循环2 h。右美托咪定组(D组):在体外循环前15 min至体外循环2 h期间经右颈内静脉以5μg/(kg∙h)的速度泵注右美托咪定注射液,其余同C组。S组置管期间及C组在转流期间均恒速泵注等容积的生理盐水。置管或转流2 h后采集标本,采用苏木精-伊红染色观察海马组织病理变化,酶联免疫吸附试验测定大鼠血浆中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)及中枢神经特异蛋白(S100β)水平,Western blotting检测海马组织中Bcl-2、Bax、Cleaved Caspase-3、PPARγ、p-PPARγ蛋白表达,免疫组织化学染色检测海马组织p-PPARγ蛋白的阳性细胞数。结果S组海马CA1区细胞形态完整;C组海马CA1区细胞受损情况较重,核固缩,间隙增宽,可见细胞死亡;D组受损较轻,海马CA1区仅见少量细胞死亡。C组和D组TNF-α、IL-6及S100β水平均高于S组(P<0.05)。与C组比较,D组TNF-α、IL-6和S100β水平降低(P<0.05)。与S组比较,C组和D组海马组织中Bcl-2蛋白相对表达量降低(P<0.05),Bax、Cleaved Caspase-3、p-PPARγ/PPARγ蛋白相对表达量升高(P<0.05);与C组比较,D组海马组织中Bcl-2蛋白相对表达升高(P<0.05),Bax、Cleaved Caspase-3、p-PPARγ/PPARγ蛋白相对表达量降低(P<0.05)。结论右美托咪定对大鼠体外循环脑保护的作用机制可能与抑制PPARγ磷酸化有关。

Objective To investigate the role of PPARγphosphorylation in the neuroprotective effect of dexmedetomidine during rat extracorporeal circulation.Methods Thirty-six adult healthy male SPF-grade SD rats weighing 350 to 450 g were randomly divided into three groups(n=12 each):Sham group(S group),Extracorporeal Circulation group(C group),and Dexmedetomidine group(D group).The S group underwent tracheal intubation under pentobarbital sodium anesthesia,with catheterization of the right internal jugular vein and bilateral femoral arteries without extracorporeal circulation.The C group underwent the same procedures as the S group but with the establishment of a rat extracorporeal circulation model for 2 hours.The D group received a continuous intravenous infusion of dexmedetomidine injection at a rate of 5μg/(kg·h)15 minutes before and during the 2-hour extracorporeal circulation via the right internal jugular vein.Saline was infused at a constant volume during catheterization or perfusion in the S and C groups.After 2 hours of catheterization or perfusion,specimens were collected.Hematoxylin and eosin staining were used to observe pathological changes in the hippocampal tissue.Enzyme-linked immunosorbent assay was employed to measure plasma concentrations of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),and central nervous system-specific protein(S100β).Western blotting was conducted to assess the protein expression of Bcl-2,Bax,Cleaved Caspase-3,PPARγ,and p-PPARγin the hippocampal tissue.Immunohistochemistry staining was performed to determine the number of positive cells expressing p-PPARγprotein in the hippocampal tissue.Results In the S group,the hippocampal CA1 region cells showed intact morphology.In the C group,the hippocampal CA1 region cells were severely damaged,exhibiting nuclear condensation,widened gaps,and visible cell death.In the D group,the damage was less severe,with only a small amount of cell death observed in the hippocampal CA1 region.Levels of TNF-α,IL-6,and S100βin the C and D groups were higher than those in the S group(P<0.05).Compared with the C group,levels of TNF-α,IL-6,and S100βin the D group were reduced(P<0.05).Relative expression levels of Bcl-2 protein in the hippocampal tissue were lower in the C and D groups than in the S group(P<0.05),while expression levels of Bax,Cleaved Caspase-3,and p-PPARγ/PPARγproteins were higher(P<0.05).Compared with the C group,the relative expression of Bcl-2 protein in the hippocampal tissue increased in the D group(P<0.05),while the expression levels of Bax,Cleaved Caspase-3,and p-PPARγ/PPARγproteins decreased(P<0.05).Conclusion The neuroprotective effect of dexmedetomidine during rat extracorporeal circulation may be associated with the inhibition of PPARγphosphorylation.