An Investigation On The Protective Effect Of Sound Conditioning From Hearing Loss And Its Anti-ROS Mechanism In Acoustic Trauma

Hearing loss due to exposure to explosive loud noise or chronic exposure to sound is called Noise-induced hearing loss, NIHL. The World Health Organization estimates that more than 12% of the world population is at risk for developing noise-induced hearing loss. Hearing protection becomes a importunate problem. In 1963, it was found by Miller's team that, in cat, exposure to low level noise prior to exposure to high intensity resulted in marked reduction in threshold shift. It was suggested that prior exposure to low level noise protests hearing from subsequent traumatic noise. Then, The first convincing evidence of it was obtained by Canlon et al. This phenomenon was termed as "toughening", "sound conditioning" and so on. Since sound conditioning presents one means of reducing the deleterious effects of noise trauma, at present, it has become a hotspot in the field of hearing research.Evidence showed that high level noise exposures cause trauma to the organ of Corti and Spiral Ganglion Cells by means of mechanical trauma, disturbance of circulationr and metabolism disorder. Recently,the influence of free oxygen radicals in tissue damage has been shown and discussion has been focused on the effect of noise-related free oxygen radicals in cochlear damage.It has been reported that the increase in free oxygen radical formation and weakness in antioxidant system may lead to noise induced hearing loss.A variety of sound conditioning paradigms have been proven successful in preventing hearing damage, however, the underlying mechanism has not been clarified. Proposed mechanisms include the upregulation of endogeneous antioxidants, the number of NMDA receptors, heat shock proteins, apoptosis, calcium buffering systems, and efferent system. It was reported that An upregulation of antioxidant enzymes in cochlear tissues have been demonstrated after sound conditioning (Jacono et al., 1998). The upregulation of theseenzymes would be expected to attenuate threshold shifts induced by noise exposure,and it suggested that one possible mechanism was to cut down the excess production of free oxygen radicals.In this study, to explore the mechanism in protective effect of sound conditioning.we observed the effect of sound conditioning on the imbalance of oxidation reduction system caused by intense noise. Methods of biochemistry, immunohistochemistry and molecular biology were introduced to investigate the changes in protein and gene expression after sound conditioning and noise exposure to provide some theoretical and experimental evidences to prevent hearing loss induced by intense noise. Statistical analysis was performed using ANOVA and SNK-q, depending on the type of data. P values below 0.05 were considered significant.Methods and results:1. Experimental grouping and treatment of animalsHealthy albino guinea pigs(weight 250g-350g), were divided randomly into four groups: Group A, sound conditioning+intensive noise exposure;Group B, intensive noise exposure;Group C, sound conditioning;Group D, quiet control. The guinea pigs of the group A were exposed to weak noise first (87dB SPL white noise 5h/d for 10 days), and then exposed to intense noise (HOdB SPL white noise 5h). The animals of the group B were exposed to the same intense noise only. The animals of the group D was kept in quiet cages(background noise 40dB SPL).2. The effect of sound conditioning against hearing loss induced by intense noise exposure in electrophysiologyHearing thresholds of auditory brainstem responses (ABRs) of all animals were measured before and after each treatment to evaluate hearing function. The hearing thresholds of guinea pigs in control group were stable. The thresholds (click and tone burst) of animals in group C changed little after treatment. The average threshold shift of the group A was 12.5dB(click) and 13.75-16.25dB(4,6,8kHz) 24h after exposure to intense noise.The threshold shifts of the group B at click and at tone(4,6,8kHz)were 30.83dB and35-35.83dB,much higher than those of group A (P<0.05).3. The effect of sound conditioning on changes in concentration of MDA caused by intense noiseImmediately after the animals were sacrificed, the cochlear were removed and made into tissue homogenate for examining the content of MDA with TBA(thiobarbituric acid) fluorophotometric method. The MDA level in the quiet control was (146±ll)nmol/L. After noise exposure, the MDA level was (188±24)nmol/L in group B, (164±18)nmol/L in group A, (149±14)nmol/L in group C, with MDA level in group B significantly higher than those in group A (P<0.05), and there is no difference in MDA level in group C and D(P>0.05).4. The effect of sound conditioning on iNOS expression in cochlear caused by intense noiseImmediately after the animals were sacrificed, the cochlear were removed and processed for examination of iNOS distribution and expression via immunohistochemistry method. Immunohistochemistry show no positive staining in group D, and very weak staining in group C, H scores were not different between them. There was positive staining in the cells of Corti, stria vascularis and spiral ganglion in group A and B, which showed theexpression of iNOS in these cells. The H score of group B(H±s=2.79±0.12) was much higher than that of group A(H±s=1.27±0.43) (P<0.05).5. The effect of sound conditioning on changes in mRNA expression and activity of SOD Immediately after the animals were sacrificed, the cochlear were removed and made into tissue homogenate. The mRNA expression and activity of SOD in Spiral Ganglion was determined by quantitive realtime-PCR and xianthine-xanthine oxidase method respectively. The expression of SOD mRNA was upregulated by 30.09% and 37.84% relative to control at Oh and 4h after intense noise exposure, and back to baseline at 12h, 24h and 36h. The expression of SOD mRNA was significantly upregulated by sound conditioning, and the increase of concentration in group A, B and C was 68.74%, 30.09%and 72.19% respectively compared with control group. The activity of SOD was ( 156.40±9.25 ) U/mgprot in group A, ( 119.50±6.40 ) U/mgprot in group B, (203.54+14.41) U/mgprot in group C and (209.40+7.81) U/mgprot in group D, withactivity of SOD in group B significantly lower than those of group A, C and D(P<0.05),but no significant differences were found between group C and D.In this study, the animal model of sound conditioning was established to observe the protective effect of sound conditioning against noise induced hearing loss. And We investigated the mechanism of the phenomenon with regard to reactive oxygen species, also the role of ROS in NIHL was demonstrated. Techniques of biochemistry, immunology and molecular biology were employed to the changes of proteins or gene expression in NIHL and the effect of sound conditioning on them. Thus the study provides pragmatic method to prevent noise-induced hearing loss, and introduced theoretical and experimental evidences in mechanism and prevention of hearing loss induced by intense noise.