神经钙调蛋白介导IL-1β诱导的NF-κB p65的表达及神经毒性

目的：探讨神经钙调蛋白在白细胞介素-1β(IL-1β)和兴奋性氨基酸（NMDA）诱导皮层神经元NF-κΒ表达及神经损伤机制的作用。 方法： 应用IL-1β或NMDA诱导原代培养的胎鼠皮层神经元损伤；通过MTT法、LDH释放率测定观察细胞生存力和损伤程度；通过Western blotting分析NF-κB p65的表达；通过膜联蛋白（Annexin V）和碘化丙啶（PI）免疫荧光法观察细胞凋亡或坏死程度。 结果： MTT和LDH释放率测定结果表明， IL-1β和NMDA都可分别引起神经细胞的生存力明显下降，并呈量效依赖关系。IL-1β和NMDA在引起细胞损伤的同时都可促进NF-κΒ p65的表达。应用神经钙调蛋白抑制剂CsA明显抑制IL-1β引起的细胞损伤，也明显抑制IL-1β诱导的NF-κB p65表达增加。但是，环孢菌素A(CsA)不能明显抑制NMDA诱导的细胞损伤（P>0.05），对NMDA诱导的NF-κΒ p65的表达的抑制作用也不明显。Annexin V 和 PI免疫荧光检测表明，IL-1β诱导的神经损伤以神经元凋亡为主，而NMDA则主要引起坏死。 结论： 神经钙调蛋白参与介导IL-1β诱导的NF-κΒ p65的表达及神经细胞凋亡，但对NMDA诱导的神经元坏死则不起主要作用。说明神经钙调蛋白是细胞凋亡信号通路中重要分子。

Calcineurin mediates the NF- κB p65 expression and neurotoxicity induced by interleukin- 1β

AIM: To explore the role of calcineurin in the expression of NF-κB and the neurotoxicity in cultured cortical neurons treated with interleukin-1β (IL-1β) and NMDA. METHODS: The cultured rat cortical neurons were used in the experiment, damage of neurons was induced by interleukin-1β(IL-1β) or excitatory amino acid (NMDA). The degree of neuron damage was examined with the methods of MTT assay and LDH releasing rate assay, as well as the Annexin V and PI immunofluorescence. The expression of NF-κB p65 on the neurons was tested by the Western blot analysis. RESULTS: Viability of neurons was obviously lower in the IL-1β group and NMDA group respectively than that in control group (P<0.05). The higher expression of NF-κB p65 was induced by IL-1β or NMDA, which were coincident with the damage of neurons. The calcineurin inhibitor cyclosporine A (CsA) inhibited the higher expression of NF-κB p65 and neuron damage induced by IL-1β (P<0.05) but not the NMDA (P>0.05). Annexin V and PI immunofluorescence showed that IL-1β mainly induced the neuron apoptosis, and NMDA induced the neuron necrosis. CONCLUSIONS: The calcineurin mediates the higher expression of NF-κB p65 and neuron damage induced by IL-1β, but not play a critical role in the necrosis induced by NMDA in the cultured cortical neurons. These results indicate that calcineurin is the key molecule in the apoptotic signaling pathway.