P2X7受体对阻塞性睡眠呼吸暂停低通气综合征大鼠认知功能障碍的机制研究

Mechanism of P2X7 Receptor on Cognitive Dysfunction in Rats with Obstructive Sleep Apnea Hypopnea Syndrome

目的 探讨P2X7受体(P2X7 receptor,P2X7R)在阻塞性睡眠呼吸暂停低通气综合征(obstructive sleep apnea hypopnea syndrome,OSAHS)诱发的大鼠认知功能障碍中的作用。方法 72只健康雄性Wistar大鼠,随机分为对照组+生理盐水组(C组)、50ml/L间歇性低氧+生理盐水组(IH组)和50 ml/L间歇性低氧+酸性亮蓝G(brilliant blue G,BBG)处理组(BBG组),每组24只大鼠。每组又分为间歇性缺氧7天和21天两个时间点,每组12只大鼠。充入氮气和空气使舱内氧浓度降低并维持在50 ml/L,然后恢复氧浓度至210 ml/L,每个循环2 min,每天8h,构建OSAHS大鼠模型。每组每天给予BBG(选择性P2X7R拮抗剂)50 mg/kg或者等量的生理盐水,直至模型建立完成。模型建立后采用Morris水迷宫实验评价大鼠学习记忆能力。行为学测试后每组取6只大鼠进行灌流取脑,免疫荧光法检测海马组织中P2X7R、M1型小胶质细胞标记物CD68和M2型小胶质细胞标记物CD206表达。每组另取6只大鼠麻醉下断头取脑,酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)测定海马组织中肿瘤坏死因子α(tumor necrosis factor alpha,TNF-α)及白细胞介素10(interleukin 10,IL-10)浓度。结果 与C组相比,IH组大鼠水迷宫测试中逃避潜伏期明显增加(P<0.05),穿越平台次数明显减少(P<0.05);海马组织中P2X7R、CD68表达及TNF-α浓度明显增加(P<0.05),IL-10和CD206明显降低(P<0.05)。与IH组相比,BBG组大鼠水迷宫测试中逃避潜伏期明显减少(P<0.05),穿越平台次数明显增加(P<0.05);海马组织中P2X7R、CD68表达及TNF-α浓度明显减少(P<0.05),IL-10和CD206明显增加(P<0.05)。结论 阻断P2X7R可能参与了OSAHS引起的认知功能障碍的保护机制,该机制可能与M1型向M2型小胶质细胞的转变,降低海马组织内促炎症因子的表达水平有关。

Objective To investigate the role of P2 X7 receptor(P2 X7 R) in cognitive dysfunction induced by obstructive sleep apnea hypopnea syndrome(OSAHS) in rats.Methods A total of 72 Wistar rats were randomly divided into three groups:control + normal saline group(group C),50 ml/L intermittent hypoxia+ normal saline group(group IH) and 50 ml/L intermittent hypoxia + brilliant blue G(BBG) group(group BBG) with 24 rats in each group.Each group was divided into intermittent hypoxia for 7 days and 21 days,with 12 rats in each group.Oxygen concentration in the chamber was reduced and maintained at 50 ml/L by filling nitrogen and air,then the oxygen concentration was restored to 210 ml/L for 2 minutes per cycle and 8 hours per day to construct the OSAHS rat model.Each group was given BBG(selective P2 X7 R antagonist) 50 mg/kg or equivalent normal saline every day until the model was completed.After the model was established,the learning and memory ability of rats were evaluated by Morris water maze.After behavioral test,6 rats in each group were collected for brain extraction by perfusion.The expressions of P2 X7 R,M1 microglia marker CD68 and M2 microglia marker CD206 in hippocampus were detected by immunofluorescence assay.Another 6 rats in each group were decapitated and their brains were harvested under anesthesia,tumor necrosis factor alpha(TNF-α) and interleukin 10(IL-10) concentration in hippocampus tissues were measured by enzyme linked immunosorbent assay(ELISA).Results Compared with group C,the escape latency in Morris water maze was significantly increased in the group IH(P<0.05),and the number of crossing platform was significantly reduced(P<0.05);the expression of P2 X7 R,CD68 and the concentration of TNF-α in hippocampus tissues were significantly increased(P<0.05),while IL-10 and CD206 were significantly decreased(P<0.05).Compared with group IH,the escape latency in Morris water maze was significantly decreased in group BBG(P<0.05),and the number of crossing platform was significantly increased(P<0.05),the expression of P2 X7 R,CD68 and the concentration of TNF-α in hippocampus tissues were significantly decreased(P<0.05),while IL-10 and CD206 were significantly increased(P<0.05).Conclusion Blocking P2 X7 R may be involved in the protective mechanism of OSAHS-induced cognitive dysfunction,which may be related to the transformation of M1-type to M2-type microglia and the reduction of pro-inflammatory cytokines in hippocampus tissues.