Characterization of CoCl2-induced reactive oxygen species (ROS): Inductions of neurite outgrowth and endothelin-2/vasoactive intestinal contractor in PC12 cells by CoCl2 are ROS dependent, but those by MnCl2 are not

CoCl2 and MnCl2 are hypoxic mimetic agents. We previously found that expression of ET-2/VIC, one of hypoxia-related factors, and the induction of neurite outgrowth in PC12 cells through ROS induced by CoCl2. MnCl2 also are known to induce neurite outgrowth in PC12 cells. However, it is unclear whether the mechanism of the effect induced by these metals is same. In the present study, we evaluated biological effects induced by MnCl2 and compared with those induced by CoCl2. Furthermore, we analyzed sources of CoCl2-induced ROS generation. MnCl2 up-regulated ET-2/VIC gene expression and ET-2/VIC peptide production as CoCl2 did, but not affect ET-1 gene expression, in the neurite outgrowth of PC12 cells. NAC did not at all inhibit the effects induced by MnCl2. Furthermore, addition of MnCl2 to the culture medium did not generate ROS as CoCl2 did. These results indicate that ET-2/VIC expression is a common pathway in neurite outgrowth induced by CoCl2 and MnCl2, but the effects induced by CoCl2 are ROS dependent, whereas the effects induced by MnCl2 are ROS independent. Taken together, the mechanism for the effects by CoCl2 was different from that by MnCl2. The ROS, were not decomposed by catalase or SOD, were rapidly generated by reaction of CoCl2 mainly with components of HS rather than with FBS or DMEM. Some ROS generated by reaction of CoCl2 with components of HS may participate in the observed neurite outgrowth of PC12 cells.