Cystine crystal-induced reactive oxygen species associated with NLRP3 inflammasome activation: implications for the pathogenesis of cystine calculi.

PURPOSE

To investigate whether cystine crystal-induced production of reactive oxygen species (ROS) and activation of NLRP3 inflammasome contribute to cystine calculi formation.

METHODS

Slc7a9-knockout rats were created as cystine calculi animal models. Kidney histological examination using TEM and immunohistochemistry were performed. The protein expression of NLRP3 and IL-1β and the concentrations of oxidative stress markers such as ROS, MDA and H2O2 in kidney tissues were estimated. In parallel, HK-2 human renal proximal tubule cells were exposed to cystine crystals and NAC treatment. The protein and mRNA expression levels of NLRP3 were evaluated. Finally, cell apoptosis and cystine crystal adherence were also assessed.

RESULTS

Activation of the NLRP3 inflammasome and marked elevations in MDA, H2O2 and ROS levels were observed both in vivo and in vitro. In particular, the protein and mRNA expression of NLRP3 was significantly increased by cystine crystals, but could be restored by an inhibitor of ROS. In addition, cell apoptosis and cystine crystal adherence were promoted by the NLRP3 inflammasome. The expression of CD44, OPN and HA in HK-2 cells was markedly increased by cystine crystals, but could be decreased by NLRP3 siRNA treatment.

CONCLUSION

Notably, we found that the activation of NLRP3 by cystine crystal-induced ROS production was of major importance in the pathogenesis of cystine calculi formation.