Protein kinase Cα requirement in the activation of p38 mitogen-activated protein kinase, which is linked to the induction of tumor necrosis factor α in lipopolysaccharide-stimulated microglia

Activated microglia have been suggested to produce a cytotoxic cytokine, tumor necrosis factor alpha (TNFalpha), in many pathological brains. Thus. determining the molecular mechanism of this induction and suppression has been the focus of a great deal of research. Using lipopolysaccharide (LPS) as an experimental inducer of TNFalpha, we investigated the regulatory mechanism by which TNFalpha is induced or suppressed in microglia. We found that LPS-induced TNFalpha is suppressed by pretreatment with the p38 mitogen-activated protein kinase (p38MAPK) inhibitor SB203580. Similar suppression was achieved by pretreatment with specific protein kinase C (PKC) inhibitors, Go6976, myristoylated pseudosubstrate (20-28), and bisindolylmaleimide. These results suggest that PKCalpha activity as well as p38MAPK activity is associated with TNFalpha induction in LPS-stimulated microglia. The requirement of PKCalpha in LPS-dependent TNFalpha induction was verified in PKCalpha-downregulated microglia which could be induced by phorbol-12-myristate-13-acetate pretreatment. Simultaneously, PKCalpha was found to be requisite for the activation of p38MAPK in LPS-stimulated microglia. In addition, the PKCalpha levels in the LPS-stimulated microglia were observed to decrease in response to the p38MAPK inhibitor, indicating that the PKCalpha levels are regulated by the p38MAPK activity. We therefore concluded that PKCalpha and p38MAPK are interactively linked to the signaling cascade inducing TNFalpha in LPS-stimulated microglia, and that in this cascade, PKCalpha is requisite for the activation of p38MAPK, leading to the induction of TNFalpha. (C) 2003 Elsevier Ltd. All rights reserved.