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GPRC6A mediates Alum-induced Nlrp3 inflammasome activation but limits Th2 type antibody responses.

ABSTRACT: Alum adjuvanticity is still an unknown mechanism despite the frequent use as vaccine adjuvant in humans. Here we show that Alum-induced inflammasome activation in vitro and in vivo is mediated by the G protein-coupled receptor GPRC6A. The Alum-induced humoral response in vivo was independent of the inflammasome because Nlrp3-/- and ASC-/- mice responded normally to Alum and blockade of IL-1 had no effect on antibody production. In contrast, Alum adjuvanticity was increased in GPRC6A-/- mice resulting in increased antibody responses and increased Th2 cytokine concentrations compared to wildtype mice. In vitro activation of GPRC6A-/- splenic B cells also induced increased IgG1 concentrations compared to wildtype B cells. For the first time, we show GPRC6A expression in B cells, contributing to the direct effects of Alum on those cells. B cell produced immunostimulatory IL-10 is elevated in GPRC6A-/- B cells in vitro and in vivo. Our results demonstrate a dual role of GPRC6A in Alum adjuvanticity. GPCR6A activation by Alum leads to the initiation of innate inflammatory responses whereas it is an important signal for the limitation of adaptive immune responses induced by Alum, partially explained by B cell IL-10.

SUBMITTER: Quandt D

PROVIDER: S-EPMC4658484 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

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GPRC6A mediates Alum-induced Nlrp3 inflammasome activation but limits Th2 type antibody responses.

Quandt Dagmar D Rothe Kathrin K Baerwald Christoph C Rossol Manuela M

Scientific reports 20151125

Alum adjuvanticity is still an unknown mechanism despite the frequent use as vaccine adjuvant in humans. Here we show that Alum-induced inflammasome activation in vitro and in vivo is mediated by the G protein-coupled receptor GPRC6A. The Alum-induced humoral response in vivo was independent of the inflammasome because Nlrp3-/- and ASC-/- mice responded normally to Alum and blockade of IL-1 had no effect on antibody production. In contrast, Alum adjuvanticity was increased in GPRC6A-/- mice resu ...[more]

PMID: 26602597

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Project description:Paclitaxel is a chemotherapeutic drug commonly used to treat different types of cancer. In addition to its antitumor effect, paclitaxel is also known to promote Toll-like receptor (TLR) 4-dependent inflammatory responses, which may lower its chemotherapeutic efficacy. However, it remains unclear whether paclitaxel is able to affect inflammasome signaling in myeloid or cancer cells. Therefore, we examined the potential effect of paclitaxel on the activation of an inflammasome complex by examining caspase-1 activation and interleukin (IL)-1β secretion in bone marrow-derived macrophages (BMDMs). The results showed that treatment with paclitaxel alone or following LPS priming failed to trigger the secretion of active caspase-1 and IL-1β from BMDMs. However, paclitaxel could induce robust activation of caspase-1 in BMDMs in the presence of NLRP3 inflammasome-activating signal 2, such as ATP or nigericin. This paclitaxel/ATP-mediated inflammasome activation was completely abrogated in Nlrp3-deficient macrophages. Mechanistically, paclitaxel treatment induced robust activation of the TLR4 signaling cascade, including phosphorylation of IκB and JNK and upregulation of proinflammatory cytokine mRNA levels in a TLR4-dependent manner. In contrast, paclitaxel treatment alone did not induce mitochondrial damages such as the loss of the mitochondrial membrane potential and production of mitochondrial ROS. These findings suggest that paclitaxel can drive the priming of signal-mediated events for NLRP3 activation but not a second signal-triggered phenomenon such as mitochondrial damage. This suggestion was supported by the observations that paclitaxel treatment caused robust IL-1β production in macrophages in the presence of cell-free medium derived from growth of injured cells and also in the spleen of mice. Collectively, our data strongly indicate that paclitaxel is able to facilitate the activation of NLRP3 inflammasome signaling in a certain physiological environment.

Project description:Intrauterine Listeria monocytogenes (L. monocytogenes) infections pose a major threat during pregnancy via affecting placental immune responses. However, the underlying mechanisms of placental defense against this pathogen remain ill-defined. Therefore, this study aims to investigate the function and the mechanism of inflammasomes on against L. monocytogenes infection during pregnancy. A listeriosis murine model and cell culture system was used to investigate the role of trophoblastic nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) in orchestrating innate immune responses to L. monocytogenes infection. Caspase-1 activity was determined using a caspase-1 activity colorimetric kit. NLRP3 and apoptosis-associated speck-like protein containing a CARD (ASC) in placental tissue was detected by immunohistochemistry. NLRP3 in HTR-8/SVneo cells was also detected by immunofluorescence. The expression of interleukin 1β (IL-1β), NLRP3, ASC, and caspase-1 was detected by Western blot. We characterized the NLRP3 inflammasome in trophoblast cells according to whether L. monocytogenes infection increased the activation of caspase-1 and the release of IL-1β. For human or mouse IL-1β in the culture supernatants and mouse tissue lysates were analyzed using ELISA Kits. Trophoblast cells constitutively expressed the components of the NLRP3 inflammasome. In vitro, L. monocytogenes triggers NLRP3 inflammasome activation in trophoblast cells by inducing caspase-1 activation, increasing the NLRP3 protein levels, IL-1β maturation and secretion in HTR-8/SVneo cells. In vivo, L. monocytogenes induces fetal resorption and IL-1β processing in pregnant mice. In addition, NLRP3-deficient mice were more prone to fetal loss than their wild-type counterparts following infection with L. monocytogenes at a lower infective dose. We conclude that trophoblast cells respond to L. monocytogenes infection through the NLRP3 receptor, resulting in inflammasome activation and IL-1β production, which prevents listeriosis during pregnancy.

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GPRC6A mediates Alum-induced Nlrp3 inflammasome activation but limits Th2 type antibody responses.

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GPRC6A mediates Alum-induced Nlrp3 inflammasome activation but limits Th2 type antibody responses.

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