| Objectives:The membranous labyrinth is in the osseous labyrinth,and the space between the membranous labyrinth and osseous labyrinth is filled with perilymph,and the membranous labyrinth is filled with endolymph.In the normal inner ear,inner ear cells play a role in stable lymphatic fluid,which maintains a constant volume and a specific concentration of sodium,potassium,chloride,and other electrolytes.The factors like genetic factors,inner ear ischemia,virus,immune response and endocrine disorders,etc.,cause excessive endolymphatic generation or endolymphatic absorption disorders,and affect the endolymphatic circulation leading to endolymphatic hydrops（EH）,which impairs the inner ear’s physiology and function.The mechanism of EH is not clear,which may be the result of multiple mechanisms.EH causes characteristic paroxysmal vertigo,accompanied by tinnitus,fluctuating hearing loss or ear tightness.The basic research of EH,especially the establishment of animal models,has become the key to study the mechanism and treatment of vertigo.EH are the same pathological changes after injury of the inner ear in many otologic diseases.Vertigo has been reported before and after otosclerosis surgery,but EH is rarely reported.The causal relationship between the two entities remains unclear.The aim of this study was to investigate the clinical features of otosclerosis associated with EH by magnetic resonance imaging and blood flow measurement of inner ear,to reveal the characteristics of EH in otosclerosis,it is helpful to discover the mechanism of EH in otosclerosis.At present,the mechanism of EH is as follows:mechanical obstruction of endolymphatic sac and endolymphatic duct,increase of Na+-K+-ATPase activity,change of Ca2+concentration,action of nitric oxide and nitric oxide synthase,deposition of immune complex and increased activity of aquaporins.The subtypes of aquaporin are widely distributed in the cochlea,vestibule,endolymphatic sac and endolymphatic duct of the inner ear,which play an important role in the transmembrane transport of fluid and the maintenance of fluid balance in inner ear.AQP4（aquaporin 4）is a water channel protein with the highest water permeability.The whole AQP4 is a three-dimensional"funnel model",which is composed of the wide-open ends on both sides of the membrane and the constricted NPA conserved sequence in the central membrane.AQP4 is closely related to the occurrence of many diseases and most widely studied in the central nervous system and plays an important role in the development of vasogenic and cytotoxic brain edema,it is also associated with a variety of autoimmune diseases.At present,the research of AQP4 in inner ear mainly focuses on hearing.In order to study the role of AQP4 in EH,the immune EH model was used to evaluate whether the model had EH symptoms and morphological changes,to detect the location of AQP4 expression in inner ear and compare whether the expression of AQP4 is increased in this model,and to explore the possible role of AQP4 in EH.It is hoped that this study will provide some help for the further study of the pathogenesis of EH and provide a new direction for the treatment of vertigo.Methods:Part Ⅰ:The clinical charts of 46 ears in 37 cases diagnosed as having otosclerosis were collected from 2012 to 2017 in the department of otorhinolaryngology of Nagoya University hospital in Japan.The subjects were divided into EH group and without EH group.Preoperative pure tone audiometry and middle ear CT were performed.3T magnetic resonance imaging（MRI）was performed 4 hours after intravenous gadolinium injection.Laser doppler flow measurements were performed in 23 ears of 22 patients undergoing stapes surgery.The type and degree of EH in otosclerosis was evaluated,and the relationship between the degree of EH and CT grade of otosclerosis was compared.The pure tone threshold and blood flow at AOW and PT in the two groups were compared.The study was approved by the Nagoya University Hospital Ethics Committee.Part Ⅱ:Thirty healthy female guinea pigs were randomly divided into normal group,model group and intervention group.1.The immune EH guinea pigs’model was established after the first systemic immunization,enhanced immunity and local immunity（epidural approach was used）.The guinea pigs in intervention group was given intraperitoneal injection of TGN020 solution.2.The vestibular symptoms were observed in each group.3.After five weeks,the average auditory threshold of experimental animals was tested by ABR test.4.The degree of cochlear EH and vestibular EH were observed and calculated by hematoxylin-eosin staining（HE）.5.The expression and distribution of AQP4 in cochlea were detected by immunohistochemistry.6.Western blot was used to compare the expression of AQP4 in three groups.Results:Part Ⅰ:The overall incidence of EH was 58.7%（27 of 46 ears）.Mild or significant EH in the cochlea was observed in 17/46（37.0%）or 7/46（15.2%）,respectively,while mild or significant EH in the vestibule was observed in 6/46（13.0%）or 6/46（13.0%）,respectively.EH in the cochlea was more common.Average thresholds in ears with significant EH were significantly higher at 500 and 1000 Hz on AC or 1000,2000,and4000 Hz on BC than in those with no or mild EH.There was no linear trend between the presence of EH and the CT grade,significantEH was more frequent in those with advanced stages.The average blood flow values of AOW with and without EH were 6.03 and 2.86 AU（ml/min/100 g）,respectively.The average blood flow values of PT with and without EH were 6.01 and 7.37AU（ml/min/100g）,respectively.The blood flow at AOW was higher in EH group.But there was no significant difference in blood flow at PT between the two groups.Part Ⅱ:After the first week of immunization,vestibular dysfunction did not appear in the experimental animals,but in the second week,the incidence of vestibular dysfunction in the model group and intervention group was 20%,40%-50%,respectively.The incidence of vestibular disorders was 50-70%in the third week after immunization.The incidence of vestibular disorders was still 70%at the fifth week in the model group.2.The average hearing threshold in ABR of the three groups was 12.5 d B,25 d B,and 20 d B,respectively.The average hearing threshold of the model group was higher than that of the normal group,but there was no significant difference between the intervention group and model group.The immune EH model could simulate the clinical features of hearing loss.The degree of hearing loss was mainly mild and moderate hearing loss.3.In the model group,EH was found in all cochlear gyrus,showing moderate and severe degree,and the EH of apical gyrus was not aggravated compared with other gyrus.In the intervention group,moderate EH was observed in each cochlea section.Cochlea EH was reduced after administration of antagonist.In model group,vestibular EH was moderate or severe.The degree of vestibular EH in the intervention group was not reduced than that in the model group.4.AQP4 is expressed in the basilar membrane of the cochlea and extends to the perilymph secreted by the stria vascularis of the spiral ligament and in the model group,AQP4 was characteristically expressed in the stria vascularis of cochlea.5.Compared with normal group,AQP4 expression increased in model group and intervention group,while AQP4expression decreased in intervention group compared with model group.Conclusions:1.EH was frequently present in ears that were candidates for stapes surgeries,especially those with severe hearing loss or advanced disease on CT.2.The existence of EH was not related to the extent of the lesions and could not predictable from clinical features.3.The more vascularity of AOW may play an important role to cause EH in Otosclerosis.4.The immune EH model of guinea pigs can simulate the clinical features of vestibular dysfunction and hearing loss.Moderate and severe EH can be seen in cochlea and vestibule.5.AQP4 is expressed in the cochlea and characteristically expressed in the stria vascularis.6.The expression of AQP4 increased in the inner ear of the model group,and the EH was alleviated after injection of antagonist,but the clinical symptoms were not relieved.7.AQP4 plays a negative role in EH and its mechanism needs further study. |
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