A nitric oxide/Ca(2+)/calmodulin/ERK1/2 mitogen-activated protein kinase pathway is involved in the mitogenic effect of IL-1beta in human astrocytoma cells.
OBJECTIVE
Evidence is accumulating to support a role for interleukin-1beta (IL-1beta) in astrocyte proliferation. However, the mechanism by which this cytokine modulates this process is not fully elucidated.


METHODS
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-1beta-induced astrocyte proliferation.


RESULTS
Low IL-1beta 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-omega-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-1beta. Inhibition of cGMP formation by the guanylate cyclase inhibitor, 1H-[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-1beta-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
These data identified the NO/Ca(2+)/CaM/ERK signalling pathway as a novel mechanism mediating the mitogenic effect of IL-1beta 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.
Meini
A
A
Department of Biomedical Sciences, University of Siena, Siena, Italy.
Sticozzi
C
C
Massai
L
L
Palmi
M
M
D001254:Astrocytoma / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D006801:Humans; D053583:Interleukin-1beta / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D008938:Mitosis / Q000502:physiology; D009569:Nitric Oxide / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors
D002147:Calmodulin; D053583:Interleukin-1beta; D009569:Nitric Oxide; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D002118:Calcium