Gallein but not fluorescein enhances the PGD₂-stimulated synthesis of osteoprotegerin and interleukin-6 in osteoblasts: Amplification of osteoprotegerin/interleukin-6 by gallein

Gallein, a small molecule related to fluorescein, is established as an inhibitor of Gβγ subunits to inhibit G protein (Gs) signaling. This agent is providing a potential therapeutic strategy to ameliorate organ dysfunctions especially involved in inflammation, however; the effects on bone metabolism have not yet been clarified. Prostaglandins (PGs) play important roles as autacoids including osteoblasts, and d-type prostanoid (DP) receptor, a member of G protein-coupled receptor specific to PGD₂, is expressed on osteoblasts. We previously reported that prostaglandin D₂ (PGD₂) induces the syntheses of osteoprotegerin (OPG) and interleukin-6 (IL-6), essential factors in bone remodelling process, and p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), and p44/p42 MAPK are involved in the signal transduction of PGD₂ in osteoblast-like MC3T3-E1 cells. Thus, we investigated in this study that the effect and the underlying mechanism of gallein, an inhibitor Gβɤ subunits, on the syntheses of OPG and IL-6 induced by PGD₂ in these cells. The cultured cells were treated with gallein or fluorescein, a structurally related compound inactive to Gβɤ subunits, and subsequently stimulated with PGD₂. Not fluorescein but gallein amplified the PGD₂-stimulated releases of OPG and IL-6. Gallein enhanced the PGD₂-upregulated mRNA expression levels of OPG and IL-6. Regarding the signaling mechanism, gallein did not affect the PGD₂-induced phosphorylation of p38 MAPK, JNK, or p42 MAPK. In conclusion, gallein upregulates the PGD₂-stimulated syntheses of OPG and IL-6 by the specific effect to inhibit Gβγ subunits in osteoblasts, but the effect is not exerted at the upstream of p38 MAPK, JNK, or p44/p42 MAPK activation.