Receptor-interacting protein kinases modulate noise-induced sensory hair cell death

Abstract

Receptor-interacting protein (RIP) kinases promote the induction of necrotic cell death pathways. Here we investigated signaling pathways in outer hair cells (OHCs) of adult male CBA/J mice exposed to noise that causes permanent threshold shifts, with a particular focus on RIP kinase-regulated necroptosis. One hour after noise exposure, nuclei of OHCs in the basal region of the cochlea displayed both apoptotic and necrotic features. RIP1 and RIP3 protein levels increased and caspase-8 was activated. Treatment with pan-caspase inhibitor ZVAD blocked the activation of caspase-8 and reduced the number of apoptotic nuclei, while increasing levels of RIP1, RIP3, and necrotic OHCs. Conversely, treatment with necrosis inhibitor necrostatin-1 (Nec-1) or RIP3 siRNA (siRIP3) diminished noise-induced increases in RIP1 and RIP3, and decreased necrotic OHC nuclei. This treatment also increased the number of apoptotic nuclei without increasing activation of caspase-8. Consistent with the elevation of levels of RIP1 and RIP3, noise-induced active AMPKα levels increased with ZVAD treatment, but decreased with Nec-1 and siRIP3 treatment. Furthermore, treatment with siRIP3 did not alter the activation of caspase-8, but instead increased activation of caspase-9 and promoted endonuclease G translocation into OHC nuclei. Finally, auditory brainstem response functional measurements and morphological assessment of OHCs showed that ZVAD treatment reduces noise-induced deficits. This protective function is potentiated when combined with siRIP3 treatment. In conclusion, noise-induced OHC apoptosis and necrosis are modulated by caspases and RIP kinases, respectively. Inhibition of either pathway shifts the prevalence of OHC death to the alternative pathway.

Emerging therapeutic interventions against noise-induced hearing loss

“…In addition, noise exposure causes the release of cytochrome C and the translocation of endonuclease G in apoptotic OHCs 19–21 . Swollen nuclei, a marker of necrotic cell death, in OHCs after noise exposure indeed suggest involvement of necrosis 18 . Recent studies have added necroptosis to this spectrum in which signaling pathways like that of the receptor-interacting protein (RIP) kinases are induced, followed by caspase activation.…”

“…Recent studies have added necroptosis to this spectrum in which signaling pathways like that of the receptor-interacting protein (RIP) kinases are induced, followed by caspase activation. Caspases 3, 8, and 9 have been found in OHCs with condensed nuclei after noise exposure 18, 22 . While an intervention into a cell death sequence may attenuate noise trauma, pathway interactions add a layer of complexity: inhibition of noise-induced apoptosis shifts the prevalence of OHC death to necrosis 18 .…”

“…This assessment is based on morphological and molecular features where apoptotic cell death is actively regulated and characterized by condensed nuclei with activation of multiple cell death pathways that are primarily regulated by caspases 18–20 . In addition, noise exposure causes the release of cytochrome C and the translocation of endonuclease G in apoptotic OHCs 19–21 .…”

Necroptosis: Mechanisms and Relevance to Disease

“…Similarly, Ripk3 −/− mice exhibit a sizeable reduction in astrocytic demise upon spinal cord injury as compared with WT mice, which correlates with an improved preservation of neurotrophic function (159). Moreover, the administration of a Ripk3 -targeting siRNA limits the demise of outer hair cells exposed to noise that causes a permanent threshold shift in hearing, an otoprotective effect that can be increased by the concomitant administration of Z-VAD-fmk (161). Taken together, these observations suggest that necroptosis may play an etiological role in both acute and chronic neurological conditions.…”

Resveratrol Promotes Recovery of Hearing following Intense Noise Exposure by Enhancing Cochlear SIRT1 Activity

“…We quantified immunolabeled signals from surface preparations or cryosections using a protocol that has been described previously [Hill et al, 2016;Zheng et al, 2014]. Briefly, immunolabeled signals of outer hair cells (OHCs) in surface preparations and spiral ganglion neurons (SGNs) in cryosections were quantified from original confocal images.…”