HgCl2-induced alteration of actin filaments in cultured primary rat proximal tubule epithelial cells labelled with fluorescein phalloidin

Abstract

When proximal tubule epithelial cells are exposed to HgCl₂, cytoplasmic blebs are formed. These represent on early, potentially reversible response to injury. These blebs are accompanied by reorganization of cytoskeletal proteins, and pre-sumably by alternations in cytoskeletal-plasma membrane interactions. Ca²⁺-activated proteinases, such as calpain, are known to affect cytoskeletal proteins and to be involved in diverse cellular processes. However, the role of calpains in cytotoxicity d due to HgCl₂ is unknown. To determine the relationship between Factin, calpain, and HgCl₂ toxicity, cells were stained with fluorescein phalloidin before and after treatment with HgCl₂. Cells were grown on coverslips and exposed to HgCl₂ (10 or 25 μM) in the presence or absence of the calpain inhibitor, leupeptin. Untreated cells were flat, polygonal, and contained many fluorescent-stained cables of actin filaments. Generally, cells exposed to HgCl₂ became pleomorphic and contracted as the blebs formed. These cells showed fewer actin cables and fluorescence was seen mostly as either compact areas of dense stain or as peripheral rings. In many cells, actin cables and filaments were completely absent. Disappearance of F-actin was initially seen by 2 min after exposure to HgCl₂. Thus, disruption of the actin cytoskeleton and blebbing were found to be early events in HgCl₂ toxicity. When leupeptin was used with HgCl₂ treatment, the actin staining appeared similar to that of untreated cells. These findings clearly illustrate that HgCl₂ injury to proximal tubule epithelial cells causes rearrangement and alteration of F-actin which may involve the activation of calpain.