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 PPARγ phosphorylation in the neuroprotective effect of dexmedetomidine through the hippocampal CA1 region during rat extracorporeal circulation

摘要:

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5 μg/（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-PPAR γ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-PPAR γ/PPAR γ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。

Abstract:

Objective To investigate the role of PPARγ phosphorylation 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 5 μg/(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-PPARγ in the hippocampal tissue. Immunohistochemistry staining was performed to determine the number of positive cells expressing p-PPARγ protein 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 S100β in 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 S100β in 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 / PPARγ proteins 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 / PPARγ proteins decreased (P < 0.05).Conclusion The neuroprotective effect of dexmedetomidine during rat extracorporeal circulation may be associated with the inhibition of PPARγ phosphorylation.