Clin Exp Otorhinolaryngol.  2018 Mar;11(1):9-16. 10.21053/ceo.2017.00626.

The Protective Effect of Egb 761 Against 3-Nitropropionic Acid-Induced Hearing Loss: The Role of Sirtuin 1

• Chang MY
• Rhee J
• Kim SH
• Kim YH

• KMID: 2407786
• DOI: http://doi.org/10.21053/ceo.2017.00626

Abstract

OBJECTIVES
Local administration of 3-nitropropionic acid (3-NP) to the inner ear induces sensorineural hearing loss. Several studies have shown the otoprotective effects of ginkgo biloba extract EGb 761. Moreover, EGb 761 has been reported to activate Sirtuin 1 (SIRT1). The present study was designed to investigate whether EGb 761 prevents 3-NP-induced sensorineural hearing loss and determine its effects on the expression of SIRT1.
METHODS
Sprague Dawley rats were divided into four experimental groups: control group receiving vehicle of 3-NP, EGb group receiving EGb 761, 3-NP group receiving 3-NP, and EGb+3-NP group receiving EGb 761 and 3-NP. EGb 761 was given orally for 5 days. The 3-NP solution was injected into the tympanum 3 days after the start of EGb 761 administration. The auditory brainstem response was recorded before and after the injection. At 4 weeks after the administration of 3-NP or vehicle of 3-NP, cochleae were harvested, and hematoxylin and eosin staining and immunohistochemistry for SIRT1 antibody were performed.
RESULTS
EGb+3-NP group showed significantly lower threshold shifts than 3-NP group. There was a significant preservation of type II fibrocytes and spiral ganglion cells in EGb+3-NP group than in 3-NP group. In EGb+3-NP group, there was a significantly greater number of SIRT1 immunopositive type II fibrocytes and spiral ganglion cells than in 3-NP group. Calculating the percentage of SIRT1 immunoreactive type II fibrocytes and spiral ganglion cells in viable type II fibrocytes and spiral ganglion cells, respectively, EGb+3-NP group showed significantly higher SIRT1 immunoreactive cells than 3-NP group.
CONCLUSION
These results suggest that EGb 761 may prevent hearing loss induced by 3-NP in an acute ototoxic animal model, which appears to be related with SIRT1 expression.

Keyword

Cochlea; Ginkgo Biloba Extract; Sirtuin 1; 3-Nitropropionic Acid

MeSH Terms

Cochlea
Ear, Inner
Ear, Middle
Eosine Yellowish-(YS)
Evoked Potentials, Auditory, Brain Stem
Ginkgo biloba
Hearing Loss*
Hearing Loss, Sensorineural
Hearing*
Hematoxylin
Immunohistochemistry
Models, Animal
Rats, Sprague-Dawley
Sirtuin 1*
Spiral Ganglion
Eosine Yellowish-(YS)
Hematoxylin
Sirtuin 1

Figure

• 

  Fig. 1. Threshold shifts in auditory brainstem responses (ABRs) at 2 weeks (A) and 4 weeks (B) after the baseline ABRs and treatment in each group. There was a significant difference of the threshold shifts among the groups (P<0.001). In a comparison of ABRs threshold shifts between EGb+3-NP group and 3-NP group, the threshold shifts in EGb+3-NP group were significantly lower (P<0.001). Error bar is a standard error. 3-NP, 3-nitropropionic acid.

• 

  Fig. 2. Light microscopic images of the spiral ligament and type II fibrocyte counts in the second turn of the cochlea. Compared with control group (A), fibrocytes in EGb group (B) were not reduced, while fibrocytes in 3-NP group (C) were reduced remarkably. The fibrocytes in EGb+3-NP group (D) were remarkably preserved compared with 3-NP group (A-D; H&E, ×100). (E) Fibrocytes in 3-NP group were reduced significantly compared with control group. Fibrocytes in EGb group were not reduced compared with control group. Fibrocytes in EGb+3-NP group were significantly preserved compared with 3-NP group. Error bar is a standard error. 3-NP, 3-nitropropionic acid. *Statistically significant difference between the two groups (P<0.05).

• 

  Fig. 3. Light microscopic images of the spiral ganglion and spiral ganglion cell counts in the second turn of the cochlea. Compared with control group (A), spiral ganglion cells in EGb group (B) were not reduced, while spiral ganglion cells in 3-NP group (C) were reduced remarkably. Spiral ganglion cells in EGb+3-NP group (D) were remarkably preserved compared with 3-NP group (A-D; H&E, ×100). (E) Spiral ganglion cells in 3-NP group were reduced significantly compared with control group. Spiral ganglion cells in EGb group were not reduced compared with control group. Spiral ganglion cells in EGb+3-NP group were significantly preserved compared with 3-NP group. Error bar is a standard error. 3-NP, 3-nitropropionic acid. *Statistically significant difference between the two groups (P<0.05).

• 

  Fig. 4. Representative staining of Sirtuin 1 in the spiral ganglion (A) and spiral ligament (B) from EGb+3-NP group. 3-NP, 3-nitropropionic acid (immunohistochemistry for SIRT1, ×100).

• 

  Fig. 5. The mean cell counts (A) and proportion (B) of SIRT1 immunoreactive type II fibrocyte. Three-NP group showed significantly lower SIRT1 immunoreactive type II fibrocytes than control, EGb, and EGb+3-NP groups (P<0.001, P=0.007, and P=0.011, respectively). The proportion of SIRT1 immunoreactive type II fibrocytes in 3-NP group was significantly lower than EGb and EGb+3-NP groups (P =0.001 and P<0.001, respectively). The mean cell counts (C) and percentage (D) of SIRT1 immunoreactive spiral ganglion cell. Three-NP group showed significantly lower SIRT1 immunoreactive spiral ganglion cell than control, EGb, and EGb+3-NP groups (P<0.001, P<0.001, and P=0.011, respectively). The proportion of SIRT1 immunoreactive spiral ganglion cells in 3-NP group was significantly lower than EGb+3-NP group (P =0.009). Error bar is a standard error. SIRT1, Sirtuin 1; 3-NP, 3-nitropropionic acid. *Statistically significant difference between the two groups (P<0.05).

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