Dataset Information

Globular Adiponectin Inhibits Lipopolysaccharide-Primed Inflammasomes Activation in Macrophages via Autophagy Induction: The Critical Role of AMPK Signaling.

ABSTRACT: The inflammasome acts as a key platform for the activation of pro-inflammatory cytokines. Adiponectin exhibits potent anti-inflammatory properties. However, the effect of adiponectin on the modulation of the inflammasome has not been explored. Herein, we show that globular adiponectin (gAcrp) suppressed lipopolysaccharide (LPS)-primed inflammasomes activation in murine peritoneal macrophages judged by prevention of interleukin-1β (IL-1β) maturation, caspase-1 activation, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) speck formation, and pyroptotic cell death. Interestingly, pretreatment with 3-methyl adenine, a pharmacological inhibitor of autophagy, abrogated the suppressive effects of gAcrp on IL-1β secretion and caspase-1 activation, indicating the crucial role of autophagy induction in gAcrp-modulation of the inflammasome activation. In addition, inhibition of 5'Adenosine monophaspahate (AMP)-activated protein kinase (AMPK) signaling abolished suppressive effect of gAcrp on inflammasomes activation. Furthermore, autophagy induction or inhibition of the inflammasome activation by gAcrp was not observed in macrophages deficient in AMPK. Taken together, these results indicate that adiponectin inhibits LPS-primed inflammasomes activation in macrophages via autophagy induction and AMPK signaling-dependent mechanisms.

SUBMITTER: Kim MJ

PROVIDER: S-EPMC5486097 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:The free-radical-operated mechanism of death of activated macrophages at sites of inflammation is unclear, but it is important to define it in order to find targets to prevent further tissue dysfunction. A well-defined model of macrophage activation at sites of inflammation is the treatment of RAW 264.7 cells with lipopolysaccharide (LPS), with the resulting production of reactive oxygen species (ROS). ROS and other free radicals can be trapped with the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO), a cell-permeable probe with antioxidant properties, which thus interferes with free-radical-operated oxidation processes. Here we have used immuno-spin trapping to investigate the role of free-radical-operated protein oxidation in LPS-induced cytotoxicity in macrophages. Treatment of RAW 264.7 cells with LPS resulted in increased ROS production, oxidation of proteins, cell morphological changes and cytotoxicity. DMPO was found to trap protein radicals to form protein-DMPO nitrone adducts, to reduce protein carbonyls, and to block LPS-induced cell death. N-Acetylcysteine (a source of reduced glutathione), diphenyleneiodonium (an inhibitor of NADPH oxidase), and 2,2'-dipyridyl (a chelator of Fe(2+)) prevented LPS-induced oxidative stress and cell death and reduced DMPO-nitrone adduct formation, suggesting a critical role of ROS, metals, and protein-radical formation in LPS-induced cell cytotoxicity. We also determined the subcellular localization of protein-DMPO nitrone adducts and identified some candidate proteins for DMPO attachment by LC-MS/MS. The LC-MS/MS data are consistent with glyceraldehyde-3-phosphate dehydrogenase, one of the most abundant, sensitive, and ubiquitous proteins in the cell, becoming labeled with DMPO when the cell is primed with LPS. This information will help find strategies to treat inflammation-associated tissue dysfunction by focusing on preventing free radical-operated proteotoxic stress and death of macrophages.

Project description:Astragalus membranaceus (AM), used in traditional Chinese medicine, has been shown to enhance immune functions, and recently, its anti-inflammatory effects were identified. However, the mechanisms of action remain unclear. Most studies have shown that autophagy might be involved in the immune response of the body, including inflammation. Here, we developed an inflammatory model by stimulating macrophages with lipopolysaccharides (LPS) to explore the anti-inflammatory effect and mechanisms of AM injection from the perspective of the regulation of autophagy. Immunoblot, immunofluorescence, and ELISA were used to determine the effects of AM injection on the production of interleukin-6 (IL-6) and alterations of autophagy markers. It was found that AM injection reduced the expression of IL-6 in LPS-stimulated macrophages and reversed the LPS-induced inhibition of cellular autophagy. After treatment with inhibitors of signaling pathways, it was shown that LPS downregulated autophagy and upregulated the production of IL-6 in macrophages via the protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway. AM injection reversed the effects of LPS by activating the AMP-activated protein kinase (AMPK) instead of inhibiting Akt. These results were further confirmed by testing activators and siRNA silencing of AMPK. Hence, these 2 distinct signaling molecules appear to exert opposite effects on mTOR, which integrates information from multiple upstream signaling pathways, negatively regulating autophagy. In addition, we demonstrated that autophagy might play a key role in regulating the production of IL-6 by testing activators and inhibitors of autophagy and siRNA silencing of ATG5. These findings showed that AM injection might enhance autophagy by activating AMPK and might further play a repressive effect on the LPS-stimulated expression of IL-6. This study explored the relationship between autophagy, signaling pathways, and the production of inflammatory factors in a model of endotoxin infection and treatment with AM injection.

Dataset Information

Globular Adiponectin Inhibits Lipopolysaccharide-Primed Inflammasomes Activation in Macrophages via Autophagy Induction: The Critical Role of AMPK Signaling.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets