Research paper
Noise-induced changes in gene expression in the cochleae of mice differing in their susceptibility to noise damage

https://doi.org/10.1016/j.heares.2010.12.014Get rights and content

Abstract

The molecular mechanisms underlying the vast differences between individuals in their susceptibility to noise-induced hearing loss (NIHL) are unknown. The present study demonstrated that the effects of noise over-exposure on the expression of molecules likely to be important in the development of NIHL differ among inbred mouse strains having distinct susceptibilities to NIHL including B6 (B6.CAST) and 129 (129X1/SvJ and 129S1/SvImJ) mice. The noise-exposure protocol produced a loss of 40 dB in hearing sensitivity in susceptible B6 mice, but no loss for the two resistant 129 substrains. Analysis of gene expression in the membranous labyrinth 6 h following noise exposure revealed upregulation of transcription factors in both the susceptible and resistant strains. However, a significant induction of genes involved in cell-survival pathways such as the heat shock proteins HSP70 and HSP40, growth arrest and DNA-damage-inducible protein 45β (GADD45β), and CDK-interacting protein 1 (p21Cip1) was detected only in the resistant mice. Moreover, in 129 mice significant upregulation of HSP70, GADD45β, and p21Cip1 was confirmed at the protein level. Since the functions of these proteins include roles in potent anti-apoptotic cellular pathways, their upregulation may contribute to protection from NIHL in the resistant 129 mice.

Highlights

► Different noise effects in gene expression accompany distinct functional outcomes. ► Expression of strong modulators of apoptosis is induced by noise in resistant mice. ► Induced p21cip1 and Gadd45β protein levels may contribute to protection from NIHL.

Introduction

Exposure to intense noise may produce either a temporary or permanent hearing loss depending upon multiple factors. Such factors involve the physical parameters of the noise stimulus including its intensity, duration, and frequency range as well as an inherent, genetically determined susceptibility to noise-induced hearing loss (NIHL). For example, a great variability in susceptibility to NIHL reflecting differences in the underlying genetic background has been reported for both humans (Davis et al., 2003, Fortunato et al., 2004) and mice (Ohlemiller and Gagnon, 2007, Sliwiniska-Kowalska et al., 2006, Sliwinska-Kowalska et al., 2006, Van Laer et al., 2006). Moreover, certain inbred mouse strains such as the 129Sv/Ev (Yoshida et al., 2000), 129X1/SvJ, and MOLF/EiJ (Candreia et al., 2004) exhibit a very high resistance to noise damage.

It has long been known that acoustic overstimulation induces adverse changes to the morphology and function of the inner ear (Engstrom et al., 1970). At the cellular level, excessive noise exposure produces permanent damage to the organ of Corti including destruction of the outer hair cells (OHCs), hair cell stereocilia, and occasionally inner hair cells (IHCs) (Hu et al., 2002, Ou et al., 2000, Wang et al., 2002, Yang et al., 2004). In addition, the pattern of damage depends upon the genetic background of the individual (Hu et al., 2002, Ohlemiller and Gagnon, 2007, Yang et al., 2004, Zhu et al., 2002). Over the past decade or so, the most intensely investigated mechanisms assumed to underlie the noise-induced degeneration of hair cells have included the production of reactive nitrogen and oxygen species as well as an overload of Ca2+ that leads to the triggering of apoptosis, the latter being one of the pathways to noise-induced hair cell death (Bohne et al., 2007, Henderson et al., 2006, Kopke et al., 1999, Ohinata et al., 2000, Ohlemiller et al., 1999b, Yamane et al., 1995). In addition, several studies of the ultrastructural changes associated with acoustic over-exposure have described an inflammatory response that involves the appearance of a phagocytic cell population in the cochlea (Fredelius, 1988, Fredelius and Rask-Andersen, 1990, Hirose et al., 2005).

An increase in reactive oxygen species, which has been detected after sound overstimulation, is thought to play a major role in the development of NIHL (Henderson et al., 2006, Ohlemiller et al., 1999a, Ohlemiller et al., 1999b). However, an increase in the activity of enzymes of the antioxidant defense system after noise exposure has also been reported, specifically enhanced glutathione reductase, γ-glutamyl cysteine synthetase, and catalase activities (Jacono et al., 1998). Other efforts exploring noise susceptibility have also focused on defining the molecular changes induced by intense sounds. For example, Cho et al. (2004) demonstrated that several immediate early genes including transcription factors and cytokines were induced 3 h after a noise exposure that resulted in permanent hearing loss. In contrast, upregulation of these genes did not occur in response to a milder noise exposure that caused a temporary, but not a permanent shift in hearing thresholds. Other studies have demonstrated induction of heat shock proteins (HSPs) after intense noise exposure (Lim et al., 1993). In addition, Kirkegaard et al. (2006) found significant early upregulation of inflammatory-response genes and genes involved in cellular antioxidant defense following over-exposure to noise. Thus, it appears that a large number of genes from various interlocked pathways are likely to make significant contributions to the development of NIHL. Particularly, the stress-associated c-Jun N-terminal kinase (JNK) signaling pathway, known to contribute to neuronal cell death induced by a variety of stressful stimuli (Derijard et al., 1994, Kyriakis et al., 1994), was demonstrated to be important in the development of NIHL. Blockade of this particular pathway provided in vivo protection from NIHL (Ahn et al., 2005, Pirvola et al., 2000, Wang et al., 2003, Wang et al., 2007, Zine and van de Water, 2004). Additionally, antisense oligonucleotides that prevent the upregulation of the JNK target gene c-Jun protected cultured spiral ganglia neurons from oxidative-stress damage, a known mediator of NIHL (Scarpidis et al., 2003).

Nevertheless, given that the pathophysiological processes of NIHL are complex, it is difficult to discern a coherent profile of alterations in gene expression with molecular methods such as the Northern blot analysis or the reverse transcriptase polymerase chain reaction. Most significantly, these techniques preclude the simultaneous analysis of large numbers of genes. The advent of cDNA-microarray technology has afforded an efficient and reliable tool for quantifying the expression of many genes simultaneously. Indeed, several studies, some of which were noted above, have described the noise-induced changes in gene expression in the cochleae of various animal species using this strategy (Cho et al., 2004, Kirkegaard et al., 2006, Lomax et al., 2001, Taggart et al., 2001).

The knowledge that some inbred mice exhibit a very high resistance to the adverse effects of noise overstimulation is intriguing. The aim of the present study was to further our understanding of the endogenous molecular mechanisms that confer such protection. Here, the results of a microarray analysis of gene expression in microdissected membranous labyrinths from different mouse strains representing unique susceptibilities to noise damage are described for a time period of 6 h after the noise exposure. Thus, changes in gene expression were studied at a period of time for which no loss of hair or supporting cells is expected which could otherwise invalidate the gene-expression experiments (Wang et al., 2002). The major finding was that exposure to excessive noise differentially affected the expression of molecules likely to be important in the development of NIHL in inbred mouse strains that are distinct in their susceptibility to NIHL. Thus, this study may provide valuable insights with respect to the future design of targeted protective interventions regarding NIHL.

Section snippets

Mice

The B6.CAST-Cdh23CAST/J (B6) strain used in this study is a congenic strain derived from the C57BL/6J but corrected for the age-related hearing loss of the parental strain by replacing its defective ahl allele with the wildtype Ahl of the Cast/Ei (Johnson et al., 1997). The ahl allele of the 129X1/SvJ (129X1) is the ahl allele common to most laboratory mouse strains including other 129 strains. This ahl allele is different from the Cast/Ei’s and also different from the defective C57BL/6J’s (

Inbred strains of mice differ in susceptibility to noise damage

The hearing sensitivity of each mouse used was evaluated within one week prior to the noise exposure. The average pre-exposure ABR thresholds at 8, 16, and at 32 kHz, were 32 ± 3, 21 ± 2, and 24 ± 2 dB SPL (±SD) for the B6, 29 ± 7, 16 ± 3, and 44 ± 5 for the 129X1 and 47 ± 10, 37 ± 10, and 38 ± 8 for the 129S1 mice. Fig. 1 illustrates the effects of the 1-h, 105-dB SPL, 10-kHz OBN exposure on the auditory function of the B6 (black bars), 129X1 (light gray bars), and 129S1 (dark gray bars) mice.

Inbred mice differ with respect to their susceptibility to NIHL

The pre-exposure ABR thresholds of the mice used in this study were in close agreement to previously reported values (Johnson et al., 1997). The B6 mice is a congenic strain corrected for the age-related hearing loss exhibited by the C57/BL6J; as expected, no loss of hearing sensitivity was noted in these mice by ten weeks of age. On the other hand, by this age, the 129 mice exhibited slightly elevated ABR thresholds, as reported previously (Zheng et al., 1999). Susceptibility to noise damage

Acknowledgements

We would like to thank Barden B. Stagner for assistance with the preparation of the illustrations. This research was supported by the Public Health Service NIH-NIDCD grants DC006442 (MAG) and DC005578 (AEV).

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    Current address: Department of Otolaryngology-Head, Neck Surgery, St Louis University, St Louis, MO, USA.

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