br Acknowledgments We thank Dr Kathleen Sullivan of Merck
Acknowledgments We thank Dr Kathleen Sullivan of Merck Research Laboratories for providing the CHO-K1 PD98059 expressing an NFAT-β-lactamase reporter and the Gα subunit Gqi5, Dr. Chen Liaw (Arena Pharmaceutical), Ms. Rebecca Kaplan, Mr. Frank Xiaodong Gan for technical assistance, Dr. Andrew Howard for comments on the manuscript, and Drs. John S. Mudgett, Kimberly L. Folander and Robert J. Driscoll for coordinating the effort between Deltagen and Merck, and Dr. Daniel Connolly for comments and support.
The inflammatory cascade initiated by sterile inflammation and mediated by the inflammasome is a central component in a wide range of diseases., , , , Inflammasome activation has a minimal requirement for 2 signals. Signal 1 typically is provided by activation of Toll-like receptors (TLRs), and results in transcriptional up-regulation of procytokines and inflammasome components. These receptors include TLR4 and TLR9, which sense a number of ligands including extracellular high-mobility group box protein 1 and double-stranded DNA released from damaged host cells, respectively., Signal 2 can be provided by diverse stimuli released from damaged cells and results in activation of inflammasome components culminating in activation of caspase-1 (CASP1). Active CASP1 in turn proteolytically cleaves and activates pro-interleukin (Il)1β into a mature form. TLR4, TLR9, IL1β, and the inflammasome components NACHT, LRR, and pyrin domain–containing protein 3 (NLRP3) and CASP1 are all strong determinants of inflammation and organ damage in acute injury of the liver and pancreas., , , The broad importance of these pathways to tissue injury has made identification of mechanisms that regulate sterile inflammation and their development as therapeutic targets a high priority. Activation of a proinflammatory signaling pathway also stimulates metabolic pathways. Specifically, TLR activation in macrophages causes an increase in aerobic glycolysis through many steps, including up-regulation of the enzymes of glycolysis and down-regulation of Krebs cycle enzymes, with a resultant increase in lactate production. This switch to glycolysis results in the accumulation of metabolites such as lactate, which can be used for the synthesis of immune mediators. Lactate is a carboxylic acid (pKa 3.86), and almost completely disassociates to the lactate anion near a physiologic pH. Lactate has several chemical and metabolic effects including reduction of the extracellular pH and conversion back to glucose by the Cori cycle within the liver. Recently, lactate was identified as the ligand for the plasma membrane G-protein–coupled receptor 81 (GPR81). GPR81 is one of a large family of GPRs with low affinity for hydroxy-carboxylic acid structure ligands. Several GPRs recently have been identified to negatively regulate TLR-mediated inflammation through interactions with intracellular protein arrestin β-2. Arrestin β-2 (ARRB2) is known to interact with GPR81-related proteins, specifically GPR40 and GPR120, and is required for their immunomodulatory effects on TLR and NLRP3 signaling through suppression of transforming growth factor-β-activated kinase 1 (TAKI) inflammatory signaling and through direct binding of NLRP3, respectively., Activation of GPR81 on adipocytes inhibits lipolysis, but its actions on macrophages and other immune cells are not known., Previous in vitro studies differed substantially from the current investigation because they did not interrogate the effects on inflammation of short-term lactate exposure in primary innate immune cells, and no in vivo studies have addressed the effect on inflammation of short-term lactate exposure as occurs in moderately intense exercise., , We hypothesized that lactate could signal though GPR81 to down-regulate NLRP3 inflammasome activity in macrophages and thus provide an important negative regulatory feedback to limit sterile inflammation. By extension, we further hypothesized that brief, high-concentration lactate exposure could activate GPR81-mediated pathways and limit sterile inflammation in vivo in the liver and pancreas.