Background: Kinins are pro-inflammatory peptides that mediate numerous vascular and
pain responses in tissue injury. Kinins exert their biological functions via two G-protein-coupled
receptors: Bradykinin 1 Receptor (B1R) and Bradykinin 2 Receptor (B2R). We previously demonstrated
the up-regulation of B2R after Hypoxia/Reoxygenation (H/R) injury in primary cultured
cortical neurons. However, the role of B2R in inflammasome-induced pyroptosis remains unknown.
Methods: We induced H/R neuronal injury in primary cultured cortical neurons harvested from
embryonic day 17 brains. Next, we examined the neuroprotective function of B2R in H/R-induced
neuronal apoptosis or necrosis using an annexin V FITC/Propidium Iodide (PI) double-staining
technique. The pyroptosis signaling cascade, including caspase-1, IL-1β and IL-18 levels and
Cleaved Gasdermin D (GSDMD) expression was examined by real-time quantitative reverse transcription
polymerase chain reaction (RT-qPCR) and western blotting to explore the underlying
Results: H/R injury significantly increased B2R protein expression (P<0.05) as well as the percentage
of early apoptotic and necrotic or late apoptotic neurons as verified by the annexin V
FITC/PI flow cytometric analysis. Bradykinin (BK), a specific B2R agonist, caused a significant
decrease in apoptotic neuronal death after H/R injury, while HOE140, a specific B2R antagonist,
markedly reduced the neuroprotective effect of B2R. Following H/R injury, BK downregulated the
caspase-1, IL-1β and IL-18 levels (P<0.01). In contrast, pretreatment with HOE140 significantly
increased caspase-1, IL-1β, and IL-18 levels (P<0.01). Further analysis revealed that GSDMD, a
key executioner of pyroptosis, is a target for B2R-mediated inhibition of neuronal pyroptosis.
Cleaved GSDMD expression was significantly inhibited by BK pretreatment and significantly enhanced
by HOE140 pretreatment (P<0.01).
Conclusion: These results indicate that activation of B2R plays an important role in pyroptosis
mediated by H/R injury.