Heterotrimeric G protein-dependent WNT-5A signaling to ERK1/2 mediates distinct aspects of microglia proinflammatory transformation.
BACKGROUND
WNT-5A signaling in the central nervous system is important for morphogenesis, neurogenesis and establishment of functional connectivity; the source of WNT-5A and its importance for cellular communication in the adult brain, however, are mainly unknown. We have previously investigated the inflammatory effects of WNT/β-catenin signaling in microglia in Alzheimer's disease. WNT-5A, however, generally recruits β-catenin-independent signaling. Thus, we aim here to characterize the role of WNT-5A and downstream signaling pathways for the inflammatory transformation of the brain's macrophages, the microglia.


METHODS
Mouse brain sections were used for immunohistochemistry. Primary isolated microglia and astrocytes were employed to characterize the WNT-induced inflammatory transformation and underlying intracellular signaling pathways by immunoblotting, quantitative mRNA analysis, proliferation and invasion assays. Further, measurements of G protein activation by [γ-(35)S]GTP binding, examination of calcium fluxes and cyclic AMP production were used to define intracellular signaling pathways.


RESULTS
Astrocytes in the adult mouse brain express high levels of WNT-5A, which could serve as a novel astroglia-microglia communication pathway. The WNT-5A-induced proinflammatory microglia response is characterized by increased expression of inducible nitric oxide synthase, cyclooxygenase-2, cytokines, chemokines, enhanced invasive capacity and proliferation. Mapping of intracellular transduction pathways reveals that WNT-5A activates heterotrimeric G(i/o) proteins to reduce cyclic AMP levels and to activate a G(i/o) protein/phospholipase C/calcium-dependent protein kinase/extracellular signal-regulated kinase 1/2 (ERK1/2) axis. We show further that WNT-5A-induced ERK1/2 signaling is responsible for distinct aspects of the proinflammatory transformation, such as matrix metalloprotease 9/13 expression, invasion and proliferation.


CONCLUSIONS
Thus, WNT-5A-induced and G protein-dependent signaling to ERK1/2 is important for the regulation of proinflammatory responses in mouse primary microglia cells. We show for the first time that WNT-5A/G protein signaling mediates physiologically important processes in primary mammalian cells with natural receptor and G protein stochiometry. Consequently, WNT-5A emerges as an important means of astrocyte-microglia communication and we, therefore, suggest WNT-5A as a new player in neuroinflammatory conditions, such as neurodegenerative disease, hypoxia, stroke, injury and infection.
Dept. Physiology & Pharmacology, Sec. Receptor Biology & Signaling, Karolinska Institutet, Nanna Svartz väg 2, Stockholm, S-17177, Sweden.
Carina
C
Halleskog
Jacomijn Petronella
JP
Dijksterhuis
Michaela Brita Christina
MB
Kilander
Javier
J
Becerril-Ortega
Juan Carlos
JC
Villaescusa
Eva
E
Lindgren
Ernest
E
Arenas
Gunnar
G
Schulte
D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D020962:Heterotrimeric GTP-Binding Proteins / Q000502:physiology*; D018836:Inflammation Mediators / Q000502:physiology*; D020935:MAP Kinase Signaling System / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000473:pathology* / Q000502:physiology; D051153:Wnt Proteins / Q000502:physiology*; D000071818:Wnt-5a Protein
D018836:Inflammation Mediators; D051153:Wnt Proteins; D000071818:Wnt-5a Protein; C438615:Wnt5a protein, mouse; D020962:Heterotrimeric GTP-Binding Proteins