Ultrastructural investigation of canine osteosarcoma cells killed by 17-AAG (17-allylamino-17-demethoxygeldanamicin) through autophagy/apoptosis/necrosis.

Introduction: 17-AAG, an Hsp90 inhibitor, exerts cytotoxic effects on several human transformed cells and the aim of this study was to investigate the mechanism of cell death induced by 17-AAG on D22, a canine osteosarcoma (OSA) cell line, using transmission electron microscopy (TEM).Materials and Methods: The D22 cell line was treated with 1, 3 or 5 μM of 17-AAG for 24 and 48 h, fixed in 2.5% gluteraldehyde, embedded in epoxy resin and ultrathin sections, stained with uranyl acetate and lead citrate, were analyzed with a Zeiss EM900.Results: 17-AAG-treated cells were pleomorphic, with variable number of lamellipodia, surface bubbles and blisters, cytoplasmic vacuoles, increased RER, mitochondrial degeneration, numerous lysosomes and free ribosomes. Morphological signs of mitochondrial autophagy (mitochondria-RER complexes, isolation membranes and autophagosomes) first appeared at 24 h with 1 μM 17-AAG, while apoptosis was prevalent at 3 μM (24 h) and necrosis at 5 μM (24 h). Ultrastructural features of the three mechanisms of cell death appeared early after 48 h treatment.Conclusions: 17-AAG exhibits time- and dose-dependent selective cytotoxicity for OSA cells inducing different types of cell death, including the recently discovered ‘mitophagy’, providing support for its potential therapeutic application in clinical settings.[...]