右美托咪定对氧糖剥夺后星形胶质细胞反应性增殖的影响∗

摘要

Objective To investigate the effects of dexmedetomidine on reactive astrocyte proliferation after oxygen-glucose deprivation.Methods Primary astrocytes were harvested from newborn SD rats of postnatal day 1-2.They were randomly di-vided into 5 groups:control group (Group C),oxygen-glucose deprivation/reperfusion (OGD/R)group (Group OGD/R), dexmedetomidine+OGD/R group (Group Dex+OGD/R),yohimbine+OGD/R group (Group YHB+OGD/R)and dexme-detomidine+yohimbine+OGD/R group (Group Dex+YHB+OGD/R).The model of OGD/R was established by replacing the standard astrocyte culture medium (DMEM/F12)with glucose-free medium and then immediately exposing cells to 95%N2 and 5% CO2 in the anaerobic chamber at 37°C for 1 h,followed by a return to standard culture conditions for another 24 h.The astrocytes in Group OGD/R were subj ected to OGD/R treatment as described above.Those in Group C were not ex-posed to OGD.The treatment for Group Dex+OGD/R was the same as for Group OGD/R,except that 10μmol/L dexmedeto-midine was added to the normal medium during reoxygenation.The treatment for Group YHB+OGD/R was the same as for Group OGD/R,except that 10 μmol/L ofα2 adrenergic receptor antagonist yohimbine was added to medium 30 min before OGD/R treatment.In Group Dex+YHB+OGD/R,10μmol/L yohimbine was added to medium 30 min before OGD/R treat-ment and 10μmol/L dexmedetomidine to medium during reoxygenation.The 5-ethynyl-2’-deoxyuridine (EdU)assay was used to evaluate the astrocyte proliferation after OGD/R treatment.Western blotting was applied to detect the protein levels of pro-liferating cell nuclear antigen (PCNA)and glial fibrillary acidic protein (GFAP),phospho-signal transducer and activator of transcription 3 (STAT3).ELISA assay was used to determine the concentrations of interlukin-6 (IL-6)and ciliary neurotrophic factor (CNTF)in the medium.Results The proportion of EdU-positive cells,the protein levels of PCNA,GFAP,p-STAT,and the release of IL-6,CNTF were increased in group OGD/R when compared with group C.They were significantly decreased in group Dex+OGD/R as compared with group OGD/R and group Dex+YHB+OGD/R.Conclusion Dexmedetomidine is capa-ble of attenuating reactive astrocyte proliferation after oxygen-glucose deprivation.%目的：探讨右美托咪定对氧糖剥夺后星形胶质细胞反应性增殖的影响。方法采用出生1~2 d的 SD大鼠,提取原代星形胶质细胞,将其随机分为5组：对照组(C 组)、氧糖剥夺/复氧(OGD/R)组、右美托咪定+OGD/R 组(Dex+OGD/R组)、α2肾上腺能受体抑制剂育亨宾+OGD/R 组(YHB+OGD/R 组)和右美托咪定+育亨宾+OGD/R 组(Dex+YHB+OGD/R组)。缺糖缺氧1 h后,复糖复氧培养24 h制备OGD/R模型。C组星形胶质细胞仅做正常培养；OGD/R组星形胶质细胞制备 OGD/R模型；Dex+OGD/R组星形胶质细胞于 OGD处理1 h后,于复糖的培养液中加入终浓度为10μmol/L的右美托咪定；YHB+OGD/R组星形胶质细胞于 OGD/R处理前30 min向培养液中加入终浓度为10μmol/L的育亨宾,之后制备 OGD/R模型；Dex+YHB+OGD/R 组于 OGD/R 处理前30 min向培养液中加入终浓度为10μmol/L的育亨宾,其它处理同Dex+OGD/R 组。EdU 法检测细胞增殖能力；Western blot 法测定增殖细胞核抗原(PCNA)、胶质纤维酸性蛋白(GFAP)、磷酸化 STAT3(p-STAT3)的蛋白表达水平；ELISA 法测定培养液中白细胞介素-6(IL-6)和睫状神经营养因子(CNTF)的浓度。结果与C组比较,OGD/R组 EdU 阳性细胞率,PCNA、GFAP、p-STAT蛋白表达水平,培养液中 IL-6和CNTF释放量均显著增高(均P<0.05)；与 OGD/R组比较,Dex+OGD/R组EdU阳性细胞率,PCNA、GFAP、p-STAT蛋白表达水平,培养液中 IL-6和 CNTF释放量较低(均P<0.05)；与 Dex+OGD/R组比较,Dex+YHB+OGD/R组 EdU 阳性细胞率,PCNA、GFAP、p-STAT3蛋白表达水平,培养液中 IL-6和CNTF释放量均较高(均P<0.05)。结论右美托咪定可缓解氧糖剥夺后星形胶质细胞反应性增殖。