.-In mammals, estrogens have dose-and cell-type-specific effects on immune cells and may act as pro-and anti-inflammatory stimuli, depending on the setting. In the bivalve mollusc Mytilus, the natural estrogen 17-estradiol (E2) has been shown to affect neuroimmune functions. We have investigated the immunomodulatory role of E 2 in Mytilus hemocytes, the cells responsible for the innate immune response. E2 at 5-25 nM rapidly stimulated phagocytosis and oxyradical production in vitro; higher concentrations of E 2 inhibited phagocytosis. E2-induced oxidative burst was prevented by the nitric oxide (NO) synthase inhibitor N G -monomethyl-L-arginine and superoxide dismutase, indicating involvement of NO and O 2 Ϫ ; NO production was confirmed by nitrite accumulation. The effects of E2 were prevented by the antiestrogen tamoxifen and by specific kinase inhibitors, indicating a receptor-mediated mechanism and involvement of p38 MAPK and PKC. E2 induced rapid and transient increases in the phosphorylation state of PKC, as well as of a aCREB-like (cAMP responsive element binding protein) transcription factor, as indicated by Western blot analysis with specific anti-phospho-antibodies. Localization of estrogen receptor-␣-and --like proteins in hemocytes was investigated by immunofluorescence confocal microscopy. The effects of E2 on immune function were also investigated in vivo at 6 and 24 h in hemocytes of E2-injected mussels. E2 significantly affected hemocyte lysosomal membrane stability, phagocytosis, and extracellular release of hydrolytic enzymes: lower concentrations of E2 resulted in immunostimulation, and higher concentrations were inhibitory. Our data indicate that the physiological role of E2 in immunomodulation is conserved from invertebrates to mammals. estrogen; innate immunity; kinase-mediated cell signaling; Mytilus IN MAMMALS, ESTROGENS EXERT a broad spectrum of activities on a wide variety of cells and tissues, including the immune system. Estrogens have dose-and cell-type-specific effects on immune cells and may act as pro-and anti-inflammatory stimuli, depending on the setting (5,22,24,43). 17-Estradiol (E 2 ) can modulate the function of neutrophil granulocytes, monocytes, and macrophages; these cell types have been reported to express intracellular and membrane estrogen receptors (ERs), and their response to E 2 can be mediated by nuclear, classical or "genomic" pathways, as well as by rapid, "nongenomic" mechanisms of action (2,3,24,45). These latter mechanisms may be initiated at membrane or cytosolic locations (29,39,49) and can result in direct local effects (e.g., modification or ion fluxes) and regulation of gene transcription secondary to activation of cytosolic kinase cascades (5,22,24,29,43).Among invertebrates, estrogens have been identified in bivalve molluscs (25,34,40,43), in which their role has been mainly investigated in the control of gametogenesis (20,21,30,33,34). However, there is evidence that estrogen can represent an important signaling molecule that is involved in roles ot...
