AbstractThyroid-stimulating hormone (TSH) treatment activates inhibitor of NF-κB/nuclear factor κB (IκB/NFκB) and extracellular signal-regulated kinase (ERK)–P38 in macrophages, but how these pathways are activated, and how they contribute to the proinflammatory effect of TSH on macrophages remain unknown. The TSH receptor (TSHR) is coupled to 4 subfamilies of G proteins (Gs, Gi/o, Gq/11, and G12/13) for its downstream signaling. This study investigated the G protein subtypes responsible for the proinflammatory effect of TSH on macrophages. qPCR showed that Gi2, Gi3, Gas, Gq, G11, G12, G13, and G15 are abundantly expressed by macrophages. The contribution of different G protein pathways to the proinflammatory effect was studied by the corresponding inhibitors or siRNA interference. While TSH-induced IκB phosphorylation was not inhibited by Gs inhibitor NF449, Gi inhibitor pertussis toxin, or Gq or G11 siRNA, it was blocked by phospholipase C inhibitor U73122 or G15 siRNA interference. TSH-induced ERK and P38 phosphorylation was blocked by G13 but not G12 siRNA interference. Interference of either G13 or G15 could block the proinflammatory effect of TSH on macrophages. The present study demonstrate that TSH activates macrophage inflammation by the G13/ERK–P38/Rho GTPase and G15/phospholipase C (PLC)/protein kinases C (PKCs)/IκB pathways.