Characterization of Cathepsin B in Mediating Silica Nanoparticle-Induced Macrophage Pyroptosis via an NLRP3-Dependent Manner.

INTRODUCTION

Silica nanoparticles (SiNPs) are one of the most widely used inorganic nanomaterials, and exposure to SiNP has been demonstrated to induce pulmonary inflammation, primarily promoted by the NLRP3-mediated macrophage pyroptosis. However, mechanisms underlying the activation of NLRP3 signaling are complex, and whether cathepsin B (CTSB), an enzyme released by the ruptured lysosome, could trigger NLRP3 assembly is controversial.

METHODS

To further characterize the role of CTSB in silica-induced pyroptosis, we conducted this study by establishing SiNP exposure models in vitro. The morphological features of SiNPs were exhibited by the SEM and TEM, and the effects of SiNPs' internalization on macrophages were examined by the TEM and immunofluorescent staining. Moreover, Western blot was performed to detect the expression of proteins related to pyroptosis and CTSB after blocking the expression of NLRP3 and CTSB.

RESULTS

We found that SiNPs internalization caused the rupture of macrophage membrane and promoted the aging of cells with increased intracellular vacuoles. Also, the expression of NLRP3, ASC, Caspase-1, GSDMD, Pro-IL-1β, IL-1β, and CTSB increased under the stimulation of SiNP, which could be suppressed by additional treatment with MCC950, an NLRP3-specific inhibitor. Besides, we found SiNP joint treatment with leupeptin, a CTSB inhibitor, could inhibit the expression of CTSB, but it had no effect on the expression of NLRP3, ASC, and Caspase-1, and the process of macrophage pyroptosis was also not affected.

CONCLUSION

SiNP exposure induces rupture of macrophages and the release of lysosomal CTSB, but CTSB fails to specifically act on the NLRP3 inflammasome to induce pyroptosis which is causally linked to lung inflammation and fibrosis.