Korean J Occup Environ Med.  2009 Sep;21(3):259-266.

Induction of Inducible Nitric Oxide Synthase Expression by Manganese in C6 Glioma Cells

• Yu GI
• Lee DH
• Chung IS
• Lee MY
• Shin DH

Abstract

OBJECTIVE
It is well established that manganese neurotoxicity is associated with clinical symptoms similar to those of idiopathic Parkinson's disease. Recent research has shown that the exposure to manganese (MnCl2) leads to induction of iNOS in BV2 microglial cells via iNOS transcriptional up-regulation and activation of both MAPKs and PI3K/Akt signaling pathways. Here, we further investigated the effect and the action mechanism of MnCl2 on iNOS expression in C6 glioma cells.
METHODS
Western blot analyses demonstrated that treatment with MnCl2 at 250 micronmeter was sufficient to induce iNOS at both the protein and mRNA levels in C6 cells.
RESULTS
These studies demonstrated that the induction of iNOS protein and mRNA was visible after 4h- and 2 h-treatment with MnCl2, respectively. MnCl2 treatment led to strong phosphorylation of JNKs and ERKs, members of MAP kinases (MAPKs), and Akt, a PI3-kinase (PI3K) downstream effector, in C6 cells. MnCl2 treatment had no effect on I kappa B-alpha in C6 cells. Notably, pretreatment with LY294002 (a PI3K inhibitor), which inhibited phosphorylation of Akt by MnCl2, caused strong suppression of MnCl2- induced iNOS protein and mRNA expression in C6 cells. Moreover, pretreatment with SP600125 (an inhibitor of JNKs) and PD98050 (an inhibitor of ERKs), which respectively interfered with MnCl2-mediated phosphorylation of JNKs and ERKs, led to the partial suppression of MnCl2-induced iNOS protein. Interestingly, pretreatment with LY294002 inhibited phosphorylation of not only Akt, but also ERKs and JNKs, in response to MnCl2. Moreover, there was an effective suppression of MnCl2-mediated phosphorylation of AKT by SP600125.
CONCLUSION
These results collectively suggest that MnCl2 induces iNOS expression in C6 glioma cells via activation of PI3K/Akt and JNK-ERK MAPK signaling proteins, whose activations seem to be mutually interconnected in response to MnCl2.

Keyword

Manganese; Inducible nitric oxide synthase; C6 glioma cells

MeSH Terms

Anthracenes
Blotting, Western
Chlorides
Chromones
Glioma
Manganese
Manganese Compounds
Morpholines
Nitric Oxide Synthase Type II
Parkinson Disease
Phosphatidylinositol 3-Kinases
Phosphorylation
Phosphotransferases
Proteins
RNA, Messenger
Up-Regulation
Anthracenes
Chlorides
Chromones
Manganese
Manganese Compounds
Morpholines
Nitric Oxide Synthase Type II
Phosphatidylinositol 3-Kinases
Phosphotransferases
Proteins
RNA, Messenger