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目的寻找能激活MN9D细胞内源性孤儿核受体(Nurrl)表达的信号分子,研究Nurrl表达增高对酪氨酸羟化酶(TH)表达的影响,探讨激活Nurrl表达的信号机制。方法用蛋白激酶A激动剂Forskolin或蛋白激酶C激动剂PMA作用于MN9D细胞,用免疫荧光细胞化学及Westernblot方法检测MN9D细胞内源性Nurrl表达的变化,以及Nurrl表达增加对TH表达的影响。利用PKA信号转导通路的特异性抑制剂H89探讨激活Nurrl表达的信号机制。结果1·从加入Forskolin1h起,直至6h,MN9D细胞Nurrl表达比未加Forskolin组明显增加(P<0·05);Forskolin主要增加MN9D细胞核内Nurrl含量,而细胞浆内Nurrl含量变化不明显;Forskolin作用后MN9D细胞TH表达无明显变化。2·用蛋白激酶A的特异性抑制剂H89预处理后,Forskolin仍具有增加Nurrl表达的作用。结论Forskolin可增加MN9D细胞内源性Nurrl表达及核转位,单纯Nurrl表达及核转位增加不影响MN9D细胞TH的表达,TH表达的激活可能还需要其他特殊的细胞(或神经元)环境或共激活因子的共同作用。Forskolin增加MN9D细胞内源性Nurrl表达及核转位的作用不是主要通过激活PKA信号转导通路完成的。
OBJECTIVE: To investigate the effect of Nurrl on the expression of tyrosine hydroxylase (TH) in MN9D cells, and to explore the signal mechanism of activating Nurrl expression. Methods MN9D cells were treated with protein kinase A agonist Forskolin or protein kinase C agonist PMA. The changes of endogenous Nurrl expression in MN9D cells and the effect of Nurrl on TH expression were detected by immunofluorescence cytochemistry and Western blot. The signal mechanism of activating Nurr1 expression was explored by H89, a specific inhibitor of PKA signaling pathway. Forskolin mainly increased the Nurrl content in MN9D cells, while the content of Nurrl in cytoplasm did not change significantly. Forskolin increased the expression of Nurrl in MN9D cells, but not in Forskolin group (P <0.05) After treatment, there was no significant change in the TH expression in MN9D cells. 2. Forskolin still has the effect of increasing Nurrl expression after preconditioning with protein kinase A specific inhibitor H89. Conclusion Forskolin can increase endogenous Nurrl expression and nuclear translocation in MN9D cells. Nurrl expression and nuclear translocation alone do not affect the expression of TH in MN9D cells. Activation of TH expression may also require other special cellular (or neuronal) Coactivator co-operation. Forskolin increased MN9D cells endogenous Nurrl expression and nuclear translocation role not through the activation of PKA signal transduction pathway to complete.