星形胶质细胞培养液及抗呆Ⅰ号对模拟损伤神经元的影响

Influence of astrocyte-conditioned medium and Kangdai-1 on neurons damaged by simulated cerebral ischemia/reperfusion

目的 探讨体外模拟脑缺血再灌注损伤条件下星形胶质细胞条件培养液(ACM)、经抗呆Ⅰ号作用的星形胶质细胞条件培养液(ACMK)及ACM与抗呆Ⅰ号联合应用(ACM+K)对损伤神经元的影响。方法 先分别进行大鼠大脑皮质星形胶质细胞(Ast)和神经元的分离纯化培养及体外模拟脑缺血再灌注损伤模型的建立,然后用再灌18 h后收集的ACM、ACMK和ACM+K以1∶5的浓度来培养损伤后的神经元,最后测定其神经元的活性、存活率、死亡率、培养液乳酸脱氢酶(LDH)的漏出率和NOS强阳性细胞的表达量。结果 ACM、ACMK和ACM+K均能使脑缺血再灌注损伤后神经元的活性和存活率明显提高,使死亡率、细胞培养液LDH的漏出率和NOS强阳性细胞的表达量显著降低,其作用 ACMK>ACM+K>ACM。结论 ①经体外模拟脑缺血再灌注损伤的神经元呈损伤→代偿→再损伤→恢复的时相变化;②ACM对受损的神经元具有重要的保护和修复作用;③Ast在脑缺血预适应(BIP)中发挥重要的作用;④抗呆Ⅰ号可通过Ast间接地保护和修复受损的神经元。

Objective To investigate the influence of astrocyte-conditioned medium (ACM), ACM treated with Kangdai-1, and ACM plus Kangdai-1 on the neurons damaged by simulated cerebral ischemia/reperfusion in vitro.Method First, the isolation, purification and cultivation of rat cortical astrocytes and neurons, and establishment of the model of stimulated cerebral damage by ischemia/reperfusion were carried out. Then, the ACM, ACM treated with Kangdai-1, and ACM plus Kangdai-1 collected 18 hours after ischemia/reperfusion, were used to culture the damaged neurons in a concentration of 1∶5. Last, the activity, survival rate and death rate of the cultured neurons, the culture fluid leakage rate of LDH, and the expressed amount of NOS-intense positive cells, were detected. Results All ACM, ACM treated with Kangdai-1, and ACM plus Kangdai-1 could markedly increase the activity and survival rate of the damaged neurons, and markedly decrease the death rate of the damaged neurons, the culture fluid leakage rate of LDH, and the expression of NOS-intense positive cells, with a sequence of potency as: ACM treated with Kangdai-1>ACM plus Kangdai-1>ACM. Conclusion (1) The neurons damaged by simulated cerebral ischemia/reperfusion in vitro showe a phasic change of damage-compensation-redamage-restoration; (2) ACM showes significant protective and repairing effects on damaged neurons; (3) Astrocytes play an important role in the process of brain ischemic preconditioning; (4) Kandai-1 may indirectly protect and repair damaged neurons through astrocytes.