Live-cell imaging of macrophage phagocytosis of asbestos fibers under fluorescence microscopy

Abstract

Abstract Background Frustrated phagocytosis occurs when an asbestos fiber > 10 μm in length is engulfed imperfectly by a macrophage, and it is believed to be associated with chromosomal instability. Few studies have focused on dynamic cellular imaging to assess the toxicity of hazardous inorganic materials such as asbestos. One reason for this is the relative lack of fluorescent probes available to facilitate experimental visualization of inorganic materials. We recently developed asbestos-specific fluorescent probes based on asbestos-binding proteins, and achieved efficient fluorescent labeling of asbestos. Results Live-cell imaging with fluorescent asbestos probes was successfully utilized to dynamically analyze asbestos phagocytosis. The fluorescently labeled asbestos fibers were phagocytosed by RAW 264.7 macrophages. Internalized fibers of 10 μm in length that were localized in the proximity of the intercellular bridge. Conclusions Fluorescently labeled asbestos facilitated visualization of the dynamic biological processes that occur during and after the internalization of asbestos fibers, and indicated that (i) frustrated phagocytosis itself does not lead to immediate cell death unless the asbestos fiber is physically pulled from the cell by an external force, and (ii) macrophages that have phagocytosed asbestos can divide but sometimes the resulting daughter cells fuse, leading to the formation of a binucleated cell. This fusion only seemed to occur when a comparatively long asbestos fiber (> 10 μm) was shared by two daughter cells.

Keywords

• Asbestos
• Frustrated phagocytosis
• Multinucleation