Hyperoxia induces the neuronal differentiated phenotype of PC12 cells via a sustained activity of mitogen-activated protein kinase induced by Bcl-2

We previously reported that rat pheochromocytoma PC12 cells express the neu ronal differentiated phenotype under hyperoxia through the production of re active oxygen species (ROS). In the present study, we found that in this ph enotype, Bcl-2, an apoptosis inhibitor, affects mitogen-activated protein ( MAP)kinase activity, which is known as a key enzyme of the signal-transduct ion cascade for differentiation. When PC12 cells were cultured under hypero xia, a rapid increase in MAP-kinase activity, including that of both p42 an d p44, was observed. Although the activity level then decreased quickly, ac tivity higher than the control level was observed for 48 h. PD98059, an inh ibitor of MAP kinase, suppressed the hyperoxia-induced neurite extensions, suggesting the involvement of MAP-kinase activity in the mechanism of diffe rentiation induced by ROS. An elevation of Bcl-2 expression was observed af ter culturing PC12 cells for 24 h under hyperoxia. This Bcl-2 elevation was not affected by treatment with PD98059, suggesting that it did not directl y induce neurite extension under hyperoxia. However, the blockade of the Bc l-2 elevation by an antisense oligonucleotide inhibited the sustained MAP-k inase activity and neurite extensions under hyperoxia. Further, in PC12 cel ls highly expressing Bcl-2, the sustained MAP-kinase activity and neurite e xtensions under hyperoxia were enhanced. These results suggested that MAP k inase is activated through the production of ROS, and the subsequent elevat ion of Bcl-2 expression sustains the MAP-kinase activity, resulting in the induction of the neuronal-differentiation phenotype of PC12 cells under hyp eroxia.