Clin Exp Otorhinolaryngol.  2008 Jun;1(2):80-85. 10.3342/ceo.2008.1.2.80.

Apoptotic Pattern of Cochlear Outer Hair Cells and Frequency-specific Hearing Threshold Shift in Noise-exposed BALB/c Mice

• Lim HW
• Choi SH
• Kang HH
• Ahn JH
• Chung JW

• KMID: 1486058
• DOI: http://doi.org/10.3342/ceo.2008.1.2.80

Abstract

OBJECTIVES
Apoptosis of outer hair cell (OHC) can be identified through nuclear staining by specific nuclear changes. The change of filamentous actin (F-actin) is also involved in early cell death process. The study was designed to investigate OHC death along the whole length of the organ of Corti. METHODS: BALB/c hybrid mice were used in this study. The noise group was exposed to white noise of 120 dB SPL for 3 hr per day for 3 consecutive days. The tone burst auditory brainstem response (ABR) test was conducted and cochleas from each group were obtained for the immunostaining of FITC phalloidin for F-actin and propidium iodide (PI) for nuclei. RESULTS: ABR threshold of the noise group significantly increased after noise exposure (P<0.001). No threshold shift was found in the control group. Threshold shift of the noise group constantly increased from 4 to 16 kHz, but threshold shifts at 16 kHz and 32 kHz were similar. Patterns of OHC staining were subclassified as FITC+PI- cells, FITC+ PI+ cells, FITC-PI+ cells and missing cells. Proportion of normal live OHCs (FITC+PI-) rapidly decreased from the apex to the base. In the basal turn, FITC-PI+ cells and vacancy OHC (missing cells) were observed easily. Apoptotic and missing cells were most abundant at 60% of the whole length of the Corti organ. CONCLUSION: We could subclassify morphologic changes in OHC death after noise exposure. Quantitative changes in OHCs along the whole Corti organ showed a plateau pattern similar to that of a frequency-specific threshold shift.

Keyword

Noise-induced hearing loss; Apoptosis; Auditory brainstem evoked potential; Cell death

MeSH Terms

Actins
Animals
Apoptosis
Cell Death
Chimera
Cochlea
European Continental Ancestry Group
Evoked Potentials, Auditory, Brain Stem
Fluorescein-5-isothiocyanate
Hair
Hearing
Hearing Loss, Noise-Induced
Humans
Mice
Noise
Organ of Corti
Phalloidine
Propidium
Actins
Fluorescein-5-isothiocyanate
Phalloidine
Propidium

Figure

• 

  Fig. 1 Images of immunoreactivity for PI and FITC-phalloidin in the organ of Corti of a noise-exposed mouse (×400). (A) Apical turn. There shows a nuclear condensation in a fluorescent image for PI (PI+, arrow, upper panel). In an image for FITC-phalloidin, the corresponding cell shows normal appearance (FITC+, arrow, lower panel). Other cells are normal live cells (FITC+, PI-). (B) Middle turn. There is a cell with no nuclear staining (arrow, upper panel) and no fluorescence for FITC-phalloidin (arrow, lower panel). This cell is considered as missing. (C) Basal turn. Nuclear condensation (PI+, arrow, upper panel) with an intact structure (FITC+, arrow, lower panel) was noted in one cell. Adjacent cells (asterisk in both panels) show nuclear condensation and a deformed structure (FITC-, PI+). The other cells (thick arrow in both panels) have no nucleus and show deformed structures (missing). Scale bar: 10 µm.

• 

  Fig. 2 Distribution of several OHC conditions along the whole length of the cochlear OHC layer in the noise group mice after noise exposure. Error bars represent standard deviation.

• 

  Fig. 3 Distribution of damaged OHCs in the noise group mice after noise exposure.

• 

  Fig. 4 ABR threshold of the noise and control groups. ABR threshold of noise group was significantly increased after noise exposure (P<0.001). ABR threshold of the noise group after noise exposure and that of the control group after silent condition within noise booth were significantly different (P<0.001). Error bars represent standard deviation.

• 

  Fig. 5 ABR threshold shift after noise exposure. Threshold shift of the noise group was constantly increased from 4 to 16 kHz, but threshold shifts at 16 and 32 kHz were similar. Error bars represent standard deviation.

Reference

1. Hamernik RP, Turrentine G, Roberto M, Salvi R, Henderson D. Anatomical correlates of impulse noise-induced mechanical damage in the cochlea. Hear Res. 1984; 3. 13(3):229–247. PMID: 6735931.

2. Wyllie AH, Kerr JF, Currie AR. Cell death: the significance of apoptosis. Int Rev Cytol. 1980; 68:251–306. PMID: 7014501.

3. Saunders JC, Dear SP, Schneider ME. The anatomical consequences of acoustic injury: a review and tutorial. J Acoust Soc Am. 1985; 9. 78(3):833–860. PMID: 4040933.

4. Henderson D, Spongr V, Subramaniam M, Campo P. Anatomical effects of impact noise. Hear Res. 1994; 6. 01. 76(1-2):101–117. PMID: 7928703.

5. Hu BH, Guo W, Wang PY, Henderson D, Jiang SC. Intense noise-induced apoptosis in hair cells of guinea pig cochleae. Acta Otolaryngol. 2000; 1. 120(1):19–24. PMID: 10779180.

6. Hu BH, Henderson D, Nicotera TM. Involvement of apoptosis in progression of cochlear lesion following exposure to intense noise. Hear Res. 2002; 4. 166(1-2):62–71. PMID: 12062759.

7. Robertson JD, Orrenius S, Zhivotovsky B. Review: nuclear events in apoptosis. J Struct Biol. 2000; 4. 129(2-3):346–358. PMID: 10806085.

8. Atencia R, Asumendi A, García-Sanz M. Role of cytoskeleton in apoptosis. Vitam Horm. 2000; 58:267–297. PMID: 10668402.

9. Asumendi A, Andollo N, Boyano MD, Hilario E, Perez-Yarza G, Atencia R, et al. The role of cleavage of cell structures during apoptosis. Cell Mol Biol (Noisy-le-grand). 2000; 2. 46(1):1–11. PMID: 10726967.

10. Rao JY, Jin YS, Zheng Q, Cheng J, Tai J, Hemstreet GP 3rd. Alterations of the actin polymerization status as an apoptotic morphological effector in HL-60 cells. J Cell Biochem. 1999; 12. 15. 75(4):686–697. PMID: 10572251.

11. Hu BH, Henderson D, Nicotera TM. F-actin cleavage in apoptotic outer hair cells in chinchilla cochleas exposed to intense noise. Hear Res. 2002; 10. 172(1-2):1–9. PMID: 12361861.

12. Yang WP, Henderson D, Hu BH, Nicotera TM. Quantitative analysis of apoptotic and necrotic outer hair cells after exposure to different levels of continuous noise. Hear Res. 2004; 10. 196(1-2):69–76. PMID: 15464303.

13. Ding D, McFadden SL, Salvi RJ. Willott JF, editor. Cochlear hair cell densities and inner-ear staining techniqiues. Handbook of mouse auditory research: from behavior to molecular biology. 2001. 1st ed. Florida: CRC press;p. 189–204.

14. Mashima T, Naito M, Tsuruo T. Caspase-mediated cleavage of cytoskeletal actin plays a positive role in the process of morphological apoptosis. Oncogene. 1999; 4. 15. 18(15):2423–2430. PMID: 10229193.

15. Maruyama W, Irie S, Sato TA. Morphological changes in the nucleus and actin cytoskeleton in the process of Fas-induced apoptosis in Jurkat T cells. Histochem J. 2000; 8. 32(8):495–503. PMID: 11095075.

16. Ohlemiller KK, Wright JS, Heidbreder AF. Vulnerability to noise-induced hearing loss in 'middle-aged' and young adult mice: a dose-response approach in CBA, C57BL, and BALB inbred strains. Hear Res. 2000; 11. 149(1-2):239–247. PMID: 11033262.

17. Santi PA, Blair A, Bohne BA, Lukkes J, Nietfeld J. The Digital Cytocochleogram. Hear Res. 2004; 6. 192(1-2):75–82. PMID: 15157965.