New Target of Oxidative Stress Regulation in Cochleae：Alternative Splicing of the p62/Sqstm1 gene

Abstract To examine the oxidative stress and the antioxidant response of p62-Keap1-Nrf2 pathway in cochleae during age-related hearing loss (ARHL) and noise-induced hearing loss (NIHL), and then elucidate the function of full-length and variant p62/Sqstm1 (referred to here as p62) in the regulation of Nrf2 activation. Cochlear damage was assessed by analyzing auditory brainstem response (ABR) and by counting hair cells (HCs). For oxidative stress detection, the malondialdehyde (MDA) levels and 7, 8-dihydro-8-oxoguanine (8-oxoG) expression in the cochleae was measured by assay kits and immunohistochemistry, respectively. The expression of full-length and variant p62 in cochleae, hippocampus (HIP) and auditory cortex (AC) was found by western blotting; For the Keap1-Nrf2 pathway activation, the Nrf2 nuclear translocation and the Nrf2 target genes HO-1/NQO-1 expression was detected by western blotting and qRT-PCR, respectively. The oxidative function of full-length and variant p62 was examined in HEI-OC-1 cells by flow cytometry. The results showed hearing loss and cochlear hair cell loss was associated with MDA accumulation and 8-oxoG expression during ARHL and NIHL. Nrf2 showed no obvious changes in nuclear protein. Expression levels mRNA for HO-1 and NQO1 was lower in old mice and mildly greater in AT Mice. The expression of p62 splicing variant lacking the Keap1-interacting region was greater than full-length p62 in cochleae. However, the expression of p62 splicing variant was lesser than full-length p62 in HIP and AC. For HEI-OC-1 cells, high expression of full-length p62 decreased ROS levels induced by H2O2. Oxidative stress is closely related to ARHL and NIHL. Changing the ratio of full-length to variant p62 protein expression may be a new target to reduce the level of oxidative stress in cochleae.