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Abstract Objective To investigate the effect of peroxynitrite on the cultured cochlear hair cells of C57BL/6 P3 mice in vitro as well as the role of Wnt3a, as an activator of the canonical Wnt signaling pathway, underlying the action of such an oxidative stress. Methods The in vitro primary cultured cochlear hair cells were subjected to l00 μM peroxynitrite and l00 μM peroxynitrite +25 ng/mL Wnt3a for 24 h, the cell survival and morphological changes were examined by immunofluorescence and transmission electron microscopy. Results The number of surviving hair cells was significantly reduced in the 100 μM peroxynitrite group, while it was significantly higher in the Wnt3a + peroxynitrite treated group compared with the peroxynitrite treated group. The transmission electron microscopy showed that exposure to peroxynitrite induced a dramatic decrease in the number of mitochondria and severely disrupted mitochondrial ultrastructure, while Wnt3a clearly diminished the disruption of mitochondrial structure and preserved a higher number of mitochondria. Conclusion These results indicated that peroxynitrite could cause oxidative damage to the cochlear hair cells, and low concentrations of Wnt3a has a protective effect against oxidative damage. Level of evidence: Level 2.
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Abstract Introduction Distortion product otoacoustic emissions (DPOAE) and their suppression may be considered useful in monitoring cochlear function and the efferent auditory pathway inhibitory effect. Nonetheless, the establishment of reliable parameters of response variations is of great importance. Objectives To verify the replicability of test and retest in the research of the inhibitory effect of the efferent pathway using contralateral suppressing stimulus during DPOAE recording for clinical applicability. Methods Cross-sectional study with 48 volunteers, aged 18 to 30 years, with normal audiometric thresholds. The procedures included were audiometric and immittance measures to overrule any conductive or sensorineural conditions and DPOAE recordings without and with contralateral suppression with a 60 dBHL white noise. Distortion product otoacoustic emissions amplitudes were analyzed and compared in both conditions with Wilcoxon test, and the Spearman correlation test was used to assess test-retest reliability. Results The comparative analysis showed differences between amplitudes in test and retest conditions only in 1,500 Hz for DPOAE measures with all other tested frequencies showing no differences, and no difference was observed in all recorded frequencies in the test and retest comparison for DPOAE suppression. The degree of correlation between test and retest of DPOAE amplitude was good at 6,000 Hz and strong (r > 0.880) at the other frequencies. For DPOAE with suppression, all frequencies presented strong correlation between test and retest: 1,500 Hz (r = 0.880), 2,000 Hz (r = 0.882), 3,000 Hz (r = 0.940), and 6,000 Hz (r = 0.957). Conclusions The study found good replicability in contralateral suppression of DPOAE with potential clinical applicability, and we recommend conducting the test from 2000Hz to higher frequencies for more reliable results.
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Objective:To investigate the influence of endoplasmic reticulum stress (ERS) in the degeneration of cochlear hair cells in the type 2diabetic mice, and to clarify its mechanism.Methods:Twenty clean Kun Ming male mice aged one month were selected and randomly divided into control group and model group (n=10) .The mice in model group were injected with STZ (40 mg·kg-1) to establish the type 2diabetic models.The fasting blood glucose levels of the mice were measured through collecting the vena caudalis blood of the mice.Auditory brain stem response (ABR) was used to detect the ABR threshold of the mice.Otoacoustic emission (OAE) test was used to detect the OAE threshold of mice.The defect rate of mouse cochlear outer hair cells was calculated by the mouse cochlear spreading technique.The expression levels of GRP78, caspase-12, p-ERK and Nrf2proteins were detected by Western blotting method.Results:Compared with control group, the fasting blood glucose levels of the mice in model group at the 7th and the 14th days had no significant differences (P＞0.05) , but the levels were increased significantly at the 21th, 28th and 35th days and the level reached the highest value at the 35th day.The ABR thresholds of the mice in model group at 8, 12, and 24kHZ were increased significantly compared with control group (P＜0.05) .Under the stimulation of low frequency, there was no significant change in the OAE threshold of the mice in model grouop compared with control group.The OAE thresholds of the mice in model group were increased significantly under the medium frequency and high frequency stimulation compared with control group (P＜0.05) .The defects of the cochlear hair cells were mainly concentrated on the bottom of gyrus of the mice, and the defects in middle temporal gyrus and parietal gyrus were less.Compared with control group, the defect rate in the bottom of gyrus of the mice in model group was increased significantly (P＜0.05) ;the defect rates in the middle temporal gyrus and parietal gyrus were increased, but there was no significant difference (P＞0.05) .The expression levels of p-ERK and Nrf2in the cochlear hair cells of the mice in model group were lower than those in control group (P＜0.05) , and the expression levels of GRP78and caspase-12were higher than those in control group (P＜0.05) .Conclusion:ERS can result in the increase of defect rate of cochlear outer hair cells and ABR brainstem hearing threshold of the diabetic mice and decrease the expression levels of p-ERK and Nrf2proteins, suggesting that ERS can promote the degenerative lesions of cochlear hair cells in the type 2diabetic mice.
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Objective: To investigate the influence of endoplasmic reticulum stress (ERS) in the degeneration of cochlear hair cells in the type 2 diabetic mice∗ and to clarify its mechanism. Methods: Twenty clean Kun Ming male mice aged one month were selected and randomly divided into control group and model group (n= 10). The mice in model group were injected with STZ (40 mg • kg ) to establish the type 2 diabetic models. The fasting blood glucose levels of the mice were measured through collecting the vena caudalis blood of the mice. Auditory brain stem response (ABR) was used to detect the ABR threshold of the mice. Otoacoustic emission (OAE) test was used to detect the OAE threshold of mice. The defect rate of mouse cochlear outer hair cells was calculated by the mouse cochlear spreading technique. The expression levels of GRP78, caspase-12, p-ERK and Nrf2 proteins were detected by Western blotting method. Results: Compared with control group, the fasting blood glucose levels of the mice in model group at the 7th and the 14th days had no significant differences ( P>0. 05) , but the levels were increased significantly at the 21th,28th and 35th days and the level reached the highest value at the 35th day. The ABR thresholds of the mice in model group at 8, 12, and 24 kHZ were increased significantly compared with control group ( P0. 05). The expression levels of p-ERK and Nrf2 in the cochlear hair cells of the mice in model group were lower than those in control group (P<0.05), and the expression levels of GRP78 and caspase-12 were higher than those in control group (P<0. 05). Conclusion: ERS can result in the increase of defect rate of cochlear outer hair cells and ABR brainstem hearing threshold of the diabetic mice and decrease the expression levels of p-ERK and Nrf2 proteins, suggesting that ERS can promote the degenerative lesions of cochlear hair cells in the type 2 diabetic mice.
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Objective To obtain an easy and high efficient method for gene transfer to cochlear hair cells ,by comparing three mediating green fluorescent protein (GFP) methods (electroporation ,adenovector and lentivirus vector) .Methods Cochlear sensory epithelium was dissected from anaesthetized P2 mice .Sensory epithelia were transferred onto poly -L -lysine treated cover slides and cultured overnight .Gene transfer was performed by elec‐troporation in medium containing pGPHI/GFP/Neo plasmid or by incubation with diluted recombined adenovirus/lentivirus vector .After 48 hours ,green fluorescence was checked under fluorescence microscope .To confirm the ef‐ficiency of exogenous gene transfer ,real-time PCR was performed using specific primers .Results The transfec‐tion efficiencies of electroporation and lentivirus vector mediated gene transfer were very low .Both immunofluores‐cence and real - time PCR results showed that the transfection efficiency of adenovirus mediated GFP and Bmi1 transfer were relatively higher .The proportion of GFP positive cells in outer hair cells and inner hair cells of middle turn were 90 .0% ± 4 .1% and 5% ± 0 .4% ,respectively .Conclusion Adenovector is more efficient for exogenous gene transfer to cochlear hair cells ,thus adenovector is a good carrier for gene transfer to cochlear hair cells .
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Objective To observe the effect of Jingui Shenqi Wan (JSW) on the content of calcium channel protein CaV1.3 of guinea pigs with ototoxic deafness induced by gentamicin (GM). Methods Forty healthy guinea pigs were randomly divided into normal group, model group and high-, middle-, and low-dose JSW treatment groups. The model group was given intramuscular injection of GM ( 120 mg/kg) per day for 10 continuous days. The high-, middle-, and low-dose JSW treatment groups were given intramuscular injection of GM (120 mg/kg) and intragastric administration of JSW in the dosage of 20.3, 13.5, 6.08 g·kg-1·d-1 respectively per day for 10 days. Immuno histochemistry was used to detect the mean optical density value of CaV1.3 expression in cochlear hair cells. Results There were significant differences of the optical density value of CaV1.3 in cochlear hair cells between the model group and normal group (P<0.01), and between the model group and high-dose JSW treatment group (P<0.05). Conclusion JSW has certain effect on preventing and treating guinea pig with ototoxic deafness induced by GM, thus to protect the hearing function.
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El interés por la fisiología y patología del sistema auditivo ha crecido en los últimos años, y no sin razón, en Estados Unidos grados variables de hipoacusia afectan al doble de la población afectada por ceguera. El sistema auditivo presenta algunas características fascinantes en su funcionamiento, la cóclea de los mamíferos, por ejemplo, es capaz de responder a vibraciones de tan solo ±0,3nm, el diámetro de un átomo y de detectar estímulos en humanos de hasta 20 KHz. El propósito del órgano de la audición es transformar la energía sonora en un impulso eléctrico que se transmite por el nervio coclear hacia el Sistema Nervioso Central. Esta revisión describe la fisiología coclear haciendo énfasis en la correlación morfofisiológica subyacente, tanto a nivel celular como molecular, intentando seguir la secuencia temporal de eventos mediante la cual un estímulo acústico se traduce en una respuesta neural.
Interest in the physiology and pathology of the auditory system has grown in recent years, not unreasonably, in the United States, variable degrees of hearing loss affect twice the population affected by blindness. The auditory system has some fascinating characteristics in their function, the cochlea of mammals, for example, is able to, answer to vibrations of ± 0,3nm, the diameter of an atom and to detect stimulations in humans up to 20 KHz. The purpose of the hearing organ is to convert sound energy into an electrical impulse that is transmitted by the cochlear nerve to the central nervous system. This review describes the cochlear physiology, making emphasis on the underlying morphophysiological correlation, in a cellular and molecular level, trying to follow the temporal sequence of events, through which, an acoustic stimulus resulting in a neural response.
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Cochlea , Auditory Perception , Hair Cells, AuditoryABSTRACT
Objective: To observe the preventive effect of Bu Shen Jian Pi Capsule on age-related cochlear hair cells apoptosis in C57 BL /6J mice,and research its mechanism of action.Methods: 30 new born C57 BL /6J mice were divided into 2 groups,with 10 animals in control group(CG) and 20 animals in Bu Shen Jian Pi Capsule (experimental group,EG).The animals in CG were fed regularly,while those in EG were fed regularly at the first one months after the birth and then,commonly fed water was substituted by the medical solution of Bu Shen Jian Pi Capsule for these animals in EG.The experimental course lasted for seven months.The cochleas of the two groups of mice were taken out for cochlear surface preparation,then being observed under optical microscope with a comparative analysis on the status of degenerative changes in the hair cells of cochlear between the tow groups.Results: Cochlear hair cells damage of CG animals showed aggravating tendency with the increase of the age,and followed the developmental rules of lesion progression in cochlear hair cells from the basal turn to the apical turn and with the number of outer hair cells loss greater than that of inner hair cells.In contrast,the cochlear damage of animals of EG was relatively milder than that in animals of CG,compared among the animals at the same age between them.Conclusion: This study showed that Bu Shen Jian Pi Capsule could postpone age-related cochlear hair cells apoptosis in C57 BL /6J mice,therefore,it suggested that Bu Shen Jian Pi Capsule might possess the pharmacological action of preventing presbyacusis.We assume the possible protective mechanism of Bu Shen Jian Pi Capsule might be related to increase of cells cAMP,regulation of metabolism of nucleic acid,and protectoin or stabilization of DNA structure.
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Objective: To study quantitative changes of cochlear hair cells and spiral ganglion cells after blast exposure in guinea pigs. Methods: The number of hair cells was calculated using the surface specimen technique and computer image analysis technique after blast exposure. The number of spiral ganglion cells was calculated using pathologic technique and computer image analysis technique after blast exposure. Results: Quantitative observation was carried out 21 days after blast exposure. The number of total hair cells in uncus, the first and second gyre in normal control group and in blast group was 3 599?159.6 and 6 022?98.4 respectively, and the number of spiral ganglion cells in the 2 low parts of cochlear in normal control group and in blast group was 51? 4.72 and 27? 6.94 respectively, the difference in mean value between the groups being significant. Conclusion: Not only the hair cells reduce, but also the spiral ganglion cells are severely damaged after blast exposure. [