Calcium oxalate monohydrate crystal disrupts tight junction via F-actin reorganization

Abstract

Tight junction is an intercellular protein complex that regulates paracellular permeability and epithelial cell polarization. This intercellular barrier is associated with actin filament. Calcium oxalate monohydrate (COM), the major crystalline composition in kidney stones, has been shown to disrupt tight junction but with an unclear mechanism. This study aimed to address whether COM crystal disrupts tight junction via actin deregulation. MDCK distal renal tubular epithelial cells were treated with 100 μg/ml COM crystals for 48 h. Western blot analysis revealed that level of a tight junction protein, zonula occludens-1 (ZO-1), significantly decreased, whereas that of β-actin remained unchanged after exposure to COM crystals. Immunofluorescence study showed discontinuation and dissociation of ZO-1 and filamentous actin (F-actin) expression at the cell border. In addition, clumping of F-actin was found in some cytoplasmic areas of the COM-treated cells. Moreover, transepithelial resistance (TER) was reduced by COM crystals, indicating the defective barrier function of the polarized cells. All of these COM-induced defects could be completely abolished by pretreatment with 20 μM phalloidin, an F-actin stabilizer, 2-h prior to the 48-h crystal exposure. These findings indicate that COM crystal does not reduce the total level of actin but causes tight junction disruption via F-actin reorganization.