Role of JNK/AP-1 signaling pathway in paraquat-induced activation of NLRP3 inflammasome in microglia

Background Paraquat (PQ) is one of the environmental factors that can cause sporadic Parkinson's disease (PD). Microglia-mediated neuroinflammation plays an important role in the occurrence and development of PD. Our previous studies have found that low doses of PQ can activate BV-2 microglia to the M1 phenotype and exert pro-inflammatory effects, but the associated mechanism is not clear yet.

Objective To explore the role of c-Jun N-terminal kinase (JNK)/activator protein 1 (AP-1) signaling pathway in PQ-induced activation of the NOD-like receptor thermal protein domain associated protoin 3 (NLRP3) inflammasome in microglia.

Methods An in vitro microglia model was established. The cells were treated with 0, 0.03, 0.06,and 0.12 μmol·L⁻¹ PQ for 24 h, the whole cell protein was extracted. The relative expression levels of JNK, AP-1 constituent proteins (c-Jun, c-Fos), NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), caspasse-1 precursor (pro caspase-1), interleukin-18 (IL-18), and interleukin-1β (IL-1β) were evaluated by Western blotting, to observe the effects of PQ exposure on JNK/AP-1 signaling pathway and NLRP3 inflammasome. After the treatment of 20 μmol·L⁻¹ JNK inhibitor SP600125, the above proteins were detected again, to explore the driving effect of JNK/AP-1 signaling pathway on NLRP3 inflammasome activation.

Results After PQ exposure, the relative expression levels of key proteins of JNK, c-Jun, and c-Fos, NLRP3, ASC, and pro caspase-1 in the 0.06 μmol·L⁻¹ PQ group and the 0.12 μmol·L⁻¹ PQ group were higher than those in the 0 μmol·L⁻¹ PQ group (P＜0.05), and the relative expression levels of IL-18 and IL-1β increased with higher exposure (P＜0.05). After the treatment of JNK inhibitor SP600125, the relative expression levels of key proteins of JNK/AP-1 signaling pathway (JNK, c-Jun, and c-Fos), NLRP3 inflammasome (NLRP3, ASC, and Pro caspase-1), and inflammatory factors (IL-18 and IL-1β) in the control group, the 20 μmol·L⁻¹ SP600125 group, and the 20 μmol·L⁻¹ SP600125+0.06 μmol·L⁻¹ PQ group were lower than those in the 0.06 μmol·L⁻¹ PQ group (P＜0.05).

Conclusion PQ exposure can activate the JNK/AP-1 signaling pathway and subsequently drive the activation of NLRP3 inflammasome in BV-2 microglia to mediate neuroinflammatory responses..