L-ergothioneine reduces mitochondrial-driven NLRP3 activation in gestational diabetes mellitus

Background: Maternal hyperglycaemia has a significant impact on placental metabolism and mitochondrial function. The NLRP3 inflammasome is responsive to endogenous signals of mitochondrial dysfunction. We tested our hypothesis that mitochondrial dysfunction orchestrates activation of the NLRP3 inflammasome and contributes to inflammation in gestational diabetes mellitus (GDM). Methods: Fasting blood, omental and placental tissue were collected on the day of caesarean section from nulliparous women with normal glucose tolerant (NGT) (n = 30) and GDM (n = 27) pregnancies. Cell-free mitochondrial DNA (cf-mtDNA) copy number was quantified by real-time PCR. M1-like (CD14⁺CD86⁺CD206^-) and M2-like (CD14⁺CD86⁺CD206⁺) macrophage populations were characterized by flow cytometry. Immunoblotting for protein expression of NLRP3, ASC and caspase-1 was performed in maternal BMI and age-matched tissue samples. IL-1β and IL-18 were measured by multiplex ELISA. Placental explants from GDM participants were cultured for 24 h with 1 mM L-ergothioneine (antioxidant) and 1 µM MCC950 (NLRP3 inhibitor). Results: Cf-mtDNA copy numbers were significantly higher in GDM compared to NGT participants (p = 0.002). Placental populations of CD14⁺ (p = 0.02) and CD14⁺CD86⁺CD206^- (p = 0.03) macrophages produced significantly increased levels of mitochondrial superoxide in GDM compared to NGT participants. Placental production of IL-18 (p = 0.04) was significantly increased in GDM. This increase in placental IL-18 was attenuated by treatment with 1 µM MCC950 (p = 0.0005), and 1 mM L-ergothioneine (p = 0.007). Conclusion: Placental inflammation is significantly increased in women with GDM. Furthermore, this increase may be initiated by elevated production of mitochondrial superoxide by macrophage subpopulations and orchestrated by the NLRP3 inflammasome. The mitochondrial antioxidant, L-ergothioneine, ameliorates NLRP3-induced placental inflammation in GDM, identifying a potential therapeutic role.