Silica phagocytosis causes apoptosis and necrosis by different temporal and molecular pathways in alveolar macrophages

Abstract

Chronic inhalation of crystalline silica is an occupational hazard that results in silicosis due to the toxicity of silica particles to lung cells. Alveolar macrophages play an important role in clearance of these particles, and exposure of macrophages to silica particles causes cell death and induction of markers of apoptosis. Using time-lapse imaging of MH-S alveolar macrophages, a temporal sequence was established for key molecular events mediating cell death. The results demonstrate that 80 % of macrophages die by apoptosis and 20 % by necrosis by clearly distinguishable pathways. The earliest detectable cellular event is phago-lysosomal leakage, which occurs between 30 and 120 min after particle uptake in both modes of death. Between 3 and 6 h later, cells undergoing apoptosis showed a dramatic increase in mitochondrial transmembrane potential, closely correlated with activation of both caspase-3 and 9 and cell blebbing. Externalization of phosphatidyl serine and nuclear condensation occurred 30 min-2 h after the initiation of cell blebbing. Cells undergoing necrosis demonstrated mitochondrial membrane depolarization but not hyperpolarization and no caspase activation. Cell swelling followed the decrease in mitochondrial membrane potential, distinguishing necrosis from apoptosis. All cells undergoing apoptosis followed the same temporal sequence, but the time lag between phago-lysosomal leakage and the other events was highly variable from cell to cell. These results demonstrate that crystalline silica exposure can result in either apoptosis or necrosis and each occurs in a well-defined but temporally variable order. The long time gap between phago-lysosomal leakage and hyperpolarization is not consistent with a simple scenario of phago-lysosomal leakage leading directly to cell death. The results highlight the importance of using a cell by cell time-lapse analysis to investigate a complex pathway such as silica induced cell death.

Responses of A549 human lung epithelial cells to cristobalite and α-quartz exposures assessed by toxicoproteomics and gene expression analysis

“…) indicated that CR and MI can cause damage to the cell membrane (an indicator of cell death), decreased BrdU incorporation (an indicator of decreased cell proliferation) and lowered cellular ATP level (an indicator of energy content) in A549 cells, which were consistent with the observed proteomic results. In addition, these findings were supported by a large body of evidence in the literature that silica particles (mostly α‐quartz in the form of Min‐U‐Sil) can induce cell death (Cassel et al ., ; Chao et al ., ; Iyer et al ., ; Joshi & Knecht, ) and inflammatory response (Cassel et al ., ; Dostert et al ., ; Hornung et al ., ; Peeters et al ., ). These cellular responses were reported in respiratory tract macrophages and epithelial cells.…”

Nanoparticle uptake: The phagocyte problem

“…Crystalline silica nanoparticles initiate both necrotic and apoptotic cell death mechanisms, a result of mitochondrial damage. Early stages of toxicity with these particles show phagolysosmal escape and cellular damage, resulting in drastic changes in mitochondrial potential [195]. These temporal changes elicited hyperpolarization or depolarization, causing apoptotic or necrotic cell death, respectively.…”

Systematic in vitro nanotoxicity study on anodic alumina nanotubes with engineered aspect ratio: Understanding nanotoxicity by a nanomaterial model

“…During LMP the green fluorescence intensity is reduced significantly due to the leakage of lysosomal components into the cytoplasm [38]. Hydrogen peroxide was used here as a prototypical LMP inducer (Fig.…”