A nitric oxide/Ca 2+ /calmodulin/ERK1/2 mitogen‐activated protein kinase pathway is involved in the mitogenic effect of IL‐1β in human astrocytoma cells

Background and purpose: Evidence is accumulating to support a role for interleukin‐1β (IL‐1β) in astrocyte proliferation. However, the mechanism by which this cytokine modulates this process is not fully elucidated. Experimental approach: In this study we used human astrocytoma U‐373MG cells to investigate the role of nitric oxide (NO), intracellular Ca 2+ concentration ([Ca 2+ ] i ), and extracellular signal‐regulated protein kinase (ERK) in the signalling pathway mediating IL‐1β‐induced astrocyte proliferation. Key results: Low IL‐1β concentrations induced dose‐dependent ERK activation which paralleled upregulation of cell division, whereas higher concentrations gradually reversed both these responses by promoting apoptosis. Pretreatment with the nonspecific NOS inhibitor, N ‐ω‐nitro‐l‐arginine methyl ester ( L ‐NAME) or the selective iNOS inhibitor, N ‐[[3‐(aminomethyl)phenyl]methyl]‐ethanimidamide dihydrochloride (1400W), antagonized ERK activation and cell proliferation induced by IL‐1β. Inhibition of cGMP formation by the guanylate cyclase inhibitor, 1 H ‐[1,2,4]oxadiazolo[4,3‐ a ]quinoxalin‐1‐one (ODQ), partially inhibited ERK activation and cell division. Functionally blocking Ca 2+ release from endoplasmic reticulum with ryanodine or 2‐aminoethoxydiphenylborane (2‐APB), inhibiting calmodulin (CaM) activity with N ‐(6‐aminohexyl)‐5‐chloro‐1‐naphthalenesulphonamide hydrochloride (W7) or MAPK kinase activity with 1,4‐diamino‐2,3‐dicyano‐1,4‐bis[2‐aminophenylthiol]butadiene (U0126) downregulated IL‐1β‐induced ERK activation as well as cell proliferation. The cytokine induced a transient and time‐dependent increase in intracellular NO levels which preceded elevation in [Ca 2+ ] i . Conclusions and implications: These data identified the NO/Ca 2+ /CaM/ERK signalling pathway as a novel mechanism mediating the mitogenic effect of IL‐1β in human astrocytes. As astrocyte proliferation is a hallmark of reactive astrogliosis, our results reveal a new potential target for therapeutic intervention in neuroinflammatory disorders. British Journal of Pharmacology (2008) 153 , 1706–1717; doi: 10.1038/bjp.2008.40 ; published online 25 February 2008