Cycloastragenol Protects Against Cisplatin-Induced Cochlear Hair Cell Apoptosis via the PI3K/Akt/mTOR Pathway

Cycloastragenol protects cochlear hair cells against cisplatin-induced ototoxicity by preserving mitochondrial function, suppressing apoptosis, and activating the PI3K/Akt/mTOR signaling pathway. These findings highlight the potential of CAG as a therapeutic candidate for preventing cisplatin-induced hearing loss. CAG significantly improved cell viability and reduced cisplatin-induced apoptosis in cochlear hair cells, as demonstrated by decreased TUNEL positivity, reduced apoptotic rates, and modulation of apoptosis-related proteins. CAG also preserved hair-cell marker expression and maintained cochlear hair-cell morphology. Furthermore, CAG restored mitochondrial function by increasing ATP production, maintaining ΔΨm, reducing ROS accumulation, and enhancing respiratory chain complex activities, while preventing mitochondrial fragmentation. In vivo, CAG markedly attenuated cisplatin-induced hearing loss, as indicated by improved ABR thresholds. Mechanistically, CAG activated the PI3K/Akt/mTOR signaling pathway, and inhibition of this pathway abolished its protective effects. Cisplatin-induced ototoxicity, a major adverse effect of chemotherapy, results in irreversible sensorineural hearing loss primarily due to apoptotic loss of cochlear sensory hair cells. Cycloastragenol (CAG), a naturally occurring triterpenoid saponin derived from Astragalus, possesses antioxidant and anti-apoptotic properties. However, its potential protective effects against cisplatin-induced cochlear injury and the underlying mechanisms remain unclear. HEI-OC1 cells, cochlear explant cultures (including the ex vivo cochlear explant model), and a mouse model were used to evaluate the protective effects of CAG against cisplatin-induced ototoxicity. Cell viability was assessed using the CCK-8 assay, while apoptosis was evaluated by TUNEL staining, flow cytometry, and Western blotting of apoptosis-related proteins. Hair-cell markers (Myo7a and Prestin) and cochlear hair-cell morphology were examined by immunofluorescence staining. Mitochondrial function was assessed by measuring ATP levels, mitochondrial membrane potential (ΔΨm), reactive oxygen species (ROS), and respiratory chain complex activities, along with mitochondrial morphology analysis using immunofluorescence and transmission electron microscopy. Auditory function in mice was evaluated by auditory brainstem response (ABR) measurements. The involvement of the PI3K/Akt/mTOR pathway was analyzed by Western blotting and inhibition assays using the PI3K inhibitor LY294002.