Role of Matrix Metalloproteinase-3 in Copper Oxide Nanoparticle-Induced Alterations of Tight Junction-Associated Proteins In Vitro and In Vivo

Abstract The extensive utilization of copper oxide nanoparticles (Nano-CuO) in industrial applications has heightened the concerns about human health effects after occupational or nonoccupational exposure. Previous studies demonstrated that Nano-CuO exposure caused pulmonary injury, inflammation, and fibrosis. However, the underlying mechanisms have not been fully understood. Here, we proposed that matrix metalloproteinase-3 (MMP-3) might play an important role in Nano-CuO-induced disruption of tight junction-associated proteins. Our results demonstrated that exposure of BEAS-2B cells to Nano-CuO, but not Nano-TiO2, caused a significant dose-dependent increase in MMP-3 production and activity, and MMP-3 activity reached the highest level at 12 h after exposure. In addition, Nano-CuO exposure also disrupted the expression of tight junction-associated proteins (ZO-1, occludin, and claudin-1), whereas MMP-3 siRNA transfection effectively reversed this downregulation in BEAS-2B cells. In vivo, Nano-CuO exposure caused increased expression of MMP-3 protein and reduced expression of tight junction-associated proteins (ZO-1 and occludin) in mouse lungs, thereby leading to epithelial barrier dysfunction and increased lung permeability. Knocking down MMP-3 significantly attenuated Nano-CuO-induced lung permeability through restoration of ZO-1 and occludin expression in mouse lungs. These findings suggest that Nano-CuO exposure significantly upregulates MMP-3 in BEAS-2B cells and mouse lungs, which mechanistically disrupts tight junction-associated proteins and compromises epithelial barrier integrity. These findings provide critical insights into the pathogenesis of pulmonary inflammation and fibrosis caused by metal nanoparticle exposure.