Secondary apoptosis of spiral ganglion cells induced by aminoglycoside: Fas-Fas ligand signaling pathway.

Objectives/hypothesis: Hair cell loss results in the secondary loss of spiral ganglion neurons (SGNs), over a period of several weeks. The death of the SGNs themselves results from apoptosis. Previous studies have shown that several molecules are involved in the apoptosis of SGNs that occurred secondary to hair cell loss. However, the precise mechanism of apoptosis of the SGNs remains unclear. The aim of this study was to ascertain the secondary apoptosis of spiral ganglion cells induced by aminoglycoside and to investigate the role of the Fas-FasL signaling pathway using guinea pigs as an experimental animal model.

Study Design: Laboratory study using experimental animals.

Methods: Guinea pigs weighing 250 to 300 g (n = 21) from 3 to 4 weeks of age were used. Gentamicin (60 microL) was injected through a cochleostomy site on their left side. At 1 (n = 7), 2 (n = 7), and 3 (n = 7) weeks after gentamicin treatment, their cochleas were obtained from their temporal bone. Hematoxylin and eosin and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining were performed to observe apoptosis. To investigate the involvement of the Fas-FasL signaling pathway in the secondary apoptosis of SGNs, we performed reverse transcription-polymerase chain reaction (RT-PCR), western blotting, and immunohistochemistry.

Results: A progressive loss of spiral ganglion cells with increasing time after gentamicin treatment was observed on light microscopic examination. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling staining demonstrated induction of apoptotic cell death in SGNs after gentamicin treatment. Expression of FasL increased over time after gentamicin treatment as determined by RT-PCR and western blotting. On immunohistochemical staining, we observed the localization of FasL in the SGNs. The proapoptotic molecules Bax and Bad were increased, but levels of the antiapoptotic molecule Bcl-2 were decreased at increasing survival times after gentamicin treatment on RT-PCR. The gentamicin-treated group displayed initial activation of caspase-8 and increased the cleavage of caspase-3, caspase-8, and PARP protein in a time-dependent manner.

Conclusions: The secondary apoptosis of SGNs could be a result of the apoptotic Fas-FasL signaling pathway. Blocking the Fas-FasL signaling pathway could be considered as a method for preventing secondary degeneration of SGNs, and further studies are needed to confirm this.