Abstract
Serine/threonine kinase 33 (STK33), a member of the calcium/calmodulin‐dependent kinase (CAMK), plays vital roles in a wide spectrum of cell processes. The present study was designed to investigate whether STK33 expressed in the mammalian cochlea and, if so, what effect STK33 exerted on aminoglycoside‐induced ototoxicity in House Ear Institute‐Organ of Corti 1 (HEI‐OC1) cells. Immunofluorescence staining and western blotting were performed to investigate STK33 expression in cochlear hair cells (HCs) and HEI‐OC1 cells with or without gentamicin treatment. CCK8, flow cytometry, immunofluorescence staining and western blotting were employed to detect the effects of STK33 knockdown, and/or U0126, and/or N‐acetyl‐L‐cysteine (NAC) on the sensitivity to gentamicin‐induced ototoxicity in HEI‐OC1 cells. We found that STK33 was expressed in both mice cochlear HCs and HEI‐OC1 cells, and the expression of STK33 was significantly decreased in cochlear HCs and HEI‐OC1 cells after gentamicin exposure. STK33 knockdown resulted in an increase in the cleaved caspase‐3 and Bax expressions as well as cell apoptosis after gentamicin damage in HEI‐OC1 cells. Mechanistic studies revealed that knockdown of STK33 led to activated mitochondrial apoptosis pathway as well as augmented reactive oxygen species (ROS) accumulation after gentamicin damage. Moreover, STK33 was involved in extracellular signal‐regulated kinase 1/2 pathway in primary culture of HCs and HEI‐OC1 cells in response to gentamicin insult. The findings from this work indicate that STK33 decreases the sensitivity to the apoptosis dependent on mitochondrial apoptotic pathway by regulating ROS generation after gentamicin treatment, which provides a new potential target for protection from the aminoglycoside‐induced ototoxicity.
Highlights
Hearing loss is regarded as a very common sensory disorder in humans all over the world, which seriously affects the quality of human life
Previous studies have confirmed that Serine/threonine kinase 33 (STK33) is highly expressed in testis,[5] and in this work, positive immunostaining was found in testis, mouse cochlea and House Ear Institute‐Organ of Corti 1 (HEI‐OC1) cells, whereby indicating that STK33 truly exists in the cochlea and HEI‐OC1 cells
Mice treated with gentamicin showed elevated Auditory brainstem response (ABR) thresholds in all frequencies examined and severe hair cells (HCs) loss, indicating that an animal model with hearing impairment has been set up successfully, which is conductive to the subsequent experiments
Summary
Hearing loss is regarded as a very common sensory disorder in humans all over the world, which seriously affects the quality of human life. It has been documented that MAPKs, which are composed of a serine/threonine protein kinases, represent the classical signal transduction pathways.[12,13] Extracellular signal‐regulated kinase 1/2 (ERK1/ 2), one of the members of the MAPK family, is able to mediate many physiological and pathological processes, such as cell proliferation, protein synthesis, cell survival, differentiation, migration and apoptosis.[14,15,16,17] Previous studies have shown that activated ERK1/2 is present in both the cytoplasm and the nucleus,[18] whereas inhibiting (MEK1/2) activation significantly reduces inner hair cells (IHCs) death caused by neomycin treatment, suggesting that ERK activation in supporting cells might promote HC death.[15,18] ERK1/2 inhibition in cochlear explants causes a loss of outer hair cells (OHCs).[18,19] Of note, STK33‐ retarded apoptosis is possibly linked to ERK1/2 pathway activation relevant to the accumulation of reactive oxygen species (ROSs).[13]. The present study was designed to investigate whether STK33 expressed in cochlear HCs and House Ear Institute‐Organ of Corti 1 (HEI‐OC1) cells and, if so, what effect STK33 exerted on gentamicin‐induced damage in cochlear HCs and HEI‐OC1 cells, with special attention given to the possible relationship between STK33 and ERK1/2 signalling pathway
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