Abstract
Inflammasome activation is associated with numerous diseases. However, invivo detection of the activated inflammasome complex has been limited by a dearth of tools. We have developed transgenic mice that ectopically express the fluorescent adaptor protein, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) and characterized the formation of assembled inflammasome complexes ("specks") in primary cells and tissues. In addition to hematopoietic cells, we have found that a stromal population in the lung tissues formed specks during the early phase of influenza infection, whereas myeloid cells showed speck formation after 2days. In a peritonitis and group B streptococcus infection model, a higher percentage of neutrophils formed specks at early phases of infection, while dendritic cells formed specks at later time points. Furthermore, speck-forming cells underwent pyroptosis and extensive release of specks to the extracellular milieu invivo. These data underscore the importance of free specks during inflammatory processes invivo.
Highlights
Inflammasomes are key signaling platforms that detect pathogenic microorganisms and sterile stressors and control the caspase-1-dependent maturation of the highly pro-inflammatory cytokines interleukin-1b (IL-1b) and IL-18
We generated a transgenic mouse expressing mouse a caspase recruitment domain (ASC)-citrine fusion protein in the Rosa26 locus. This system contains a knockin of the ASC-citrine gene and a proximal loxP-flanked stop site, allowing us to conditionally express ASC-citrine in a lineage-specific manner (Figure S1A)
The embryonic stem cell (ESC) clones that harbored the expected ASC-citrine fragment were confirmed by Southern blot analysis (Figure S1B), and the targeted ESC clones were injected into blastocysts to generate chimeric mice
Summary
Inflammasomes are key signaling platforms that detect pathogenic microorganisms and sterile stressors and control the caspase-1-dependent maturation of the highly pro-inflammatory cytokines interleukin-1b (IL-1b) and IL-18. Both IL-1b and IL-18 have numerous functions, including activation of feed-forward pathways that result in the production of more cytokines, such as tumor necrosis factor alpha (TNF-a). Inflammasomes consist of three proteins: (1) a receptor that recognizes danger signals, (2) the adaptor protein ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain), and (3) the enzyme caspase-1 (Latz, 2010). The autocatalytically activated form of caspase-1 converts pro-IL-1b and pro-IL-18 into the corresponding mature active cytokines
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