Alterations of inner ear morphology in experimental hypercholesterolemia.

Previous clinical and experimental studies have indicated that auditory function may be compromised by hypercholesterolemia. In this investigation, inner ear tissue from chinchillas maintained on a cholesterol-supplemented diet for 3 months was examined for morphological alterations which might underlie the physiological changes observed with this condition in earlier studies. Ultrastructural analysis of cochleas from 16 hypercholesterolemic chinchillas revealed alterations in both the stria vascularis and outer hair cells. Strial marginal cells throughout the cochlea and outer hair cells of the apical turn, contained electron-lucent patches of an amorphous material. These patches had the morphological characteristics and histochemical properties of glycogen. Mild extracellular edema and increased numbers of lysosomes were also noted in the stria vascularis of experimental animals. These alterations suggest that chronic hypercholesterolemia metabolically stresses inner ear tissue. It is hypothesized that such changes could increase susceptibility of the cochlea to ototraumatic agents.     

Effect of a beta-stimulant on the inner ear stria vascularis

In vivo and in vitro experiments were conducted to investigate the effect of alpha-isoproterenol on the inner ear stria vascularis with intracellular cytochrome oxidase activity used as an index. Intraperitoneal injection of alpha-isoproterenol (5 mg/kg) was performed in 10 rats, and that of physiological saline in 4 rats, for 21 consecutive days. After the 3-week treatment, bilateral cochleas were excised for frozen sections and stained for cytochrome oxidase. The staining density of the stria vascularis for the enzyme was analyzed with a computer. Electron microscopic observation was also performed for some specimens. As for the in vitro experiments, bilateral cochleas from 6 normal rats were excised for cell culture. Cochlear cells from the right ear were cultured with medium containing alpha-isoproterenol (10-micromol/L concentration), and those from the left ear with medium alone. After 3-day culture, the enzyme activity of cytochrome oxidase in the stria vascularis was quantified by the same method used for the in vivo experiments. Cytochrome oxidase activity was markedly elevated in the alpha-isoproterenol group. The activity tended to be higher in the lower turns of the cochlea. Electron microscopy revealed that numerous mitochondria were present in marginal cells that protruded into the endolymphatic space. The enzyme activity was also elevated in the stria vascularis from cochlear specimens in the alpha-isoproterenol group of the in vitro experiment. The above results suggest that alpha-isoproterenol accelerated the metabolic activity of the cells that constitute the stria vascularis. The increase in activity was probably attributable to direct pharmaceutical effects of the beta-stimulant, rather than an increase in blood flow. It is possible that the cells that constitute the stria vascularis may have beta-receptors.     

The cochlea of the spontaneously diabetic mouse

Summary We used transmission electron microscopy to examine the cochleae of non-obese diabetic mice as animal models for human type I or non-insulin-dependent diabetes mellitus. Pathological changes were observed in the organ of Corti of the basal turn and in the stria vascularis of each turn. Major findings in the stria vascularis were protrusion or condensation of marginal cells, swelling of intermediate cells, and widening of the intercellular spaces. Principal findings in the organ of Corti involved degenerative changes of the outer and inner hair cells and replacement of hair cells by supporting cells. No prominent pathological changes were observed in the capillaries. The possible mechanism of diabetic involvement in cochlear pathology is discussed.     

The cochlea of the spontaneously diabetic mouse. II. Electron microscopic observations of non-obese diabetic mice

We used transmission electron microscopy to examine the cochlea of non-obese diabetic mice as animal models for human type I or non-insulin-dependent diabetes mellitus. Pathological changes were observed in the organ of Corti of the basal turn and in the stria vascularis of each turn. Major findings in the stria vascularis were protrusion or condensation of marginal cells, swelling of intermediate cells, and widening of the intercellular spaces. Principal findings in the organ of Corti involved degenerative changes of the outer and inner hair cells and replacement of hair cells by supporting cells. No prominent pathological changes were observed in the capillaries. The possible mechanism of diabetic involvement in cochlear pathology is discussed.     

Ultrastructural localization of calmodulin in gerbil cochlea by immunogold electron microscopy

Localization of calmodulin, a calcium binding protein, was identified in adult gerbil cochleas using paraffin section immunohistochemistry and immunogold electron microscopy with monoclonal antibody against bovine calmodulin. Immunoreactive calmodulin was abundant in inner hair cells (IHCs), outer hair cells (OHCs) and Boettcher cells of the cochleas. Other cell types containing calmodulin were marginal cells and basal cells of the stria vascularis, fibrocytes in the spiral ligament, spiral ganglion neurons and vascular smooth muscle cells. Immunogold labeling for calmodulin was observed in cuticular plate, stereocilia, and within cytoplasm of IHCs and OHCs. In OHCs the labeling was mostly observed in the region underlying lateral wall corresponding to subsurface cisterna. In IHCs the staining was diffuse in the cytoplasm and denser than that in OHCs. Boettcher cells showed dense staining along the microvillous projections facing to the intercellular spaces between Boettcher cells and Claudius cells and between the neighboring Boettcher cells. These distributions of calmodulin in the hair cells consist with the assumption that IHCs act as a true neurotransducer and OHCs as an active bi-directional mechanotransducer. The rich presence of calmodulin in Boettcher cells suggests that the cells may involve in mediating Ca(2+) regulation and play a distinctive active role in ion transport.     

Ultrastructural pathology in the stria vascularis of the MRL-Fasl(lpr) mouse.

The MRL-Fas(lpr) mouse, a model of multisystemic, organ nonspecific autoimmune disease, has been proposed as a model of immune-mediated inner ear disease. A preliminary study employing light microscopy indicated that it develops cochlear pathology that appeared most striking in the stria vascularis, where cells underwent edema and degeneration. However, other structures, including the inner and outer hair cells and the supporting cells, also appeared to display pathology. The current study analyzed cochlear ultrastructure using transmission electron microscopy to better delineate the cochlear lesions found in these animals. MRL-Fas(lpr) animals were allowed to develop systemic disease (20 weeks old) and then had auditory brainstem response (ABR) thresholds determined. Animals were then killed and their cochleas prepared for electron microscopy. Age-matched MRL-+/+ and BALB/c mice served as controls. Results indicated that MRL-Fas(lpr) mice demonstrated elevated ABR thresholds. In contrast to a preliminary report, the cochlear pathology was observed exclusively in the stria vascularis, where cells demonstrated hydropic degeneration. Strial capillary structure was normal as were the rest of the cellular cochlear constituents. No inflammatory infiltrate was noted. These studies confirm that the MRL-Fas(lpr) mouse develops cochlear abnormalities focused in the stria vascularis. Whether the mechanism of the cellular degeneration involves autoimmune, genetic, or uremic processes has yet to be determined.     

噪声刺激对耳蜗一氧化氮合酶的影响

目的：探讨一氧化氮（ＮＯ）在噪声性聋发病中的作用。方法：用中高频连续稳态噪声制作噪声性聋的动物模型,用ＮＡＤＰＨ－黄递酶组织化学、原位杂效和Ｎｏｒｔｈｅｒｎ印迹法,观察噪声刺激对耳蜗一氧化氮合酶（ＮＯＳ）表达的影响。结果：组织化学法显示ＮＯＳ主要分布于内外毛细胞、螺旋神经节细胞和血管纹边缘细胞;原位杂效法发现ＮＯＳｍＲＮＡ在内外毛细胞、螺旋神经节细胞胞浆内均可见阳性染色,但血管纹边缘细胞无阳性染色     

Development of the human stria vascularis.

Fifteen human fetal cochleas were investigated by light microscopy and transmission electron microscopy in order to observe the development of the stria vascularis. The earliest signs of strial cell differentiation take place during the 11th week of gestation. Subsequently, the first stages of the stria vascularis development occur quickly. At week 14 the three types of cells, namely, marginal, intermediate and basal cells are discernable. Moreover at this stage, signs of specific activity are already present. The adult-like appearance of the stria vascularis is reached by week 21 but its maturation is completed only during the last trimester of pregnancy. This is in good agreement both with the development of the organ of Corti structures and with the maturation of the human auditory function.     

Susceptibility of organ of Corti with or without melanin to acoustic overstimulation]

Albino and pigmented guinea pigs were compared in terms of susceptibility to acoustic trauma. The animals were exposed to a 4 kHz pure tone of 120 dB for 60 min. N1 thresholds of CAP were measured before and after the acoustic exposure. Changes in the outer hair cell and stria vascularis were studied using SEM and TEM. After acoustic trauma, N1 thresholds were more elevated in the albino than in the pigmented guinea pigs. Also, pathological changes in the outer hair cell and stria vascularis were more severe in the albino animals. A noteworthy finding in the stria vascularis was that the melanin in intermediate cells had moved into marginal cells. This melanin migration may be possibly involved in mechanisms underlying prevention of acoustic trauma.     

Atrophy of the stria vascularis, a common cause for hearing loss

Atrophy of the stria vascularis is a genetically determined deafness of aging characterized by a bilateral symmetrical sensori-neural hearing loss showing flat audiometric patterns and excellent speech discrimination. The temporal bones of individuals exhibiting this type of hearing loss were studied by serial sections and surface preparations for light microscopy and by electron microscopy. The atrophic changes are most severe in the apical regions of the cochleas and involve the marginal, intermediate and basal cells in that order of severity. It seems probable that atrophy of the stria vascularis causes some deficiency in the quality of endolymph throughout the cochlear duct, regardless of the location of the atrophy. Typically all other structures of the cochlear duct are normal, and the sense organ when stimulated within its sensitivity range is capable of normal stimulus coding, thus accounting for the usually excellent speech discrimination.     

Degeneration of the organ of Corti following intravenous administration of cisplatin.

The development of acute morphological changes in the cochlea was studied in guinea pigs given one intravenous high-dose injection of cisplatin. In the light microscope three major stages of degeneration in the organ of Corti could be recognized: 1) an initial swelling of the Hensen's cells and protrusion of the Deiters' cells into the space of Nuel enclosing the outer hair cells, 2) a gradual degeneration of the outer hair cells together with a vacuolization in the region of the base of the inner hair cells, 3) a collapse of the Reissner's membrane and the entire organ of Corti with different degrees of damage to the inner hair cells. Sporadic bulging of the marginal cells of the stria vascularis into the endolymphatic space could be observed 4 days after injection.     

Elemental composition of individual cells and tissues in the cochlea

Localization of elements at the cellular and sub-cellular levels was performed with the energy dispersive X-ray microanalysis technique, using shock-frozen, freeze-dried and araldite-embedded mouse (CBA/CBA) cochleae sectioned dry. Anatomical identification occurred in the STEM (scanning transmission electron microscopy) mode. In inner hair cell stereocilia the K/Na ratio was 70:1 but only 20:1 in the cytoplasm. In outer hair cell cytoplasm the K/Na ratio was 11:1 while the ratio in stereocilia was similar to that in inner hair cells. Ca was identified in stereocilia and the upper part of the cytoplasm of both outer and inner hair cells. The elemental composition in the subtectorial space is endolymph-like and that in the inner tunnel of the organ of Corti is similar to extracellular fluid. Considerable regional differences in elemental composition occur in the tectorial membrane with regard to P, K and Ca. The highest concentration of Ca occurs in the basal part of the tectorial membrane towards the sensory hairs. The highest concentration of K occurs in the basal and outer parts whereas the middle part of the tectorial membrane contains low levels of both K and Ca. The elemental composition changes in two main directions: 1) from the limbal (growing) region to the tip of the tectorial membrane, and, 2) from upper to lower surfaces. The three cell types of the stria vascularis differ considerably in elemental composition. The highest concentration of K occurs in marginal cells. The basal cells contain more K than do the intermediate cells. A significantly higher concentration of Ca, Cl and Na occurs in marginal cell cytoplasm than in any other cell type in the stria vascularis.     

Establishment of inner ear epithelial cell culture: isolation, growth and characterization

Select epithelial regions of the bovine inner ear were established and maintained in cell culture. Marginal cells from the stria vascularis and dark cells from the posterior wall of the utricle were isolated, dissociated and placed in culture medium. Within 24 h, cellular islands of hexagonal-shaped, epithelial-like cells from both the stria vascularis and posterior utricular wall were readily identifiable by inverted light microscopy. Ultrastructural examination of both the cultured stria marginal cells and utricular dark cells revealed that both cell types had numerous microvilli on their apical surfaces and interdigitating infoldings of their basolateral surfaces. Apical tight junctional complexes were present between apposing cells. These findings demonstrate that inner ear bovine epithelial cells can be successfully isolated and maintained in culture, and that such cells retain certain of their in vivo morphological characteristics.     

Cochlear ultrastructure in two-phase endolymphatic hydrops in the guinea-pig

Endolymphatic hydrops continues to be considered as a pathological factor in the etiology of Menière’s disease. We have developed the two-phase endolymphatic hydrops model, which seems to represent a functional model combining multiple etiologies, and which may resemble the fluctuating characteristics of Menière’s disease. A transmission electron microscopic study was performed on the endolymphatic sacs of four groups of guinea-pig cochleas: (1) controls, (2) non-operated, aldosterone-treated cochleas, (3) operated (dissection of the endolymphatic sac) cochleas, (4) operated and aldosterone-treated cochleas. Light and electron microscopy showed a normal morphology in the controls. Aldosterone as a single treatment resulted in an increased activity of the marginal cells in the stria vascularis. Dissection induced a gradient of degenerative effects and cell loss in the intracellular and extracellular structures of the sensory cells, the stria vascularis and Reissner’s membrane, which may be reversible. Subsequent administration of aldosterone induced severe damage and increased cell loss, which may be irreversible. Our findings demonstrate changes that may be reversible due to the compromising effect of a single treatment, and irreversible changes due to interaction of both compromising factors. These findings support our two-phase concept. Received: 30 October 2000 / Accepted: 14 June 2001     

Electron microscopic temporal bone histopathology in experimental pneumococcal meningitis.

Bacterial meningitis is one of the most common causes of acquired profound sensorineural deafness in children. Measurement of hearing and examination of the cochlea is limited in patients suffering from acute meningitis. A rabbit model of pneumococcal meningitis was developed to identify the temporal bone histopathologic changes that occur in meningogenic labyrinthitis caused by Streptococcus pneumoniae. Light microscopy was previously performed on temporal bones from acutely meningitic rabbits with profound hearing loss as determined electrophysiologically. Extensive inflammation of the cochlea with endolymphatic hydrops was observed. The organ of Corti, however, showed preserved architecture in the majority of these animals. In order to further investigate these findings, a protocol was used to create meningitic rabbits with hearing loss ranging from early high-frequency loss to profound deafness. The temporal bones from 7 rabbits were examined by transmission electron microscopy. In cases of mild hearing loss, partial degeneration of the inner row of outer hair cells, as well as edema of efferent cochlear nerve endings and marginal cells of the stria vascularis, was seen. With increasing degrees of hearing loss, the remainder of the organ of Corti and intermediate cells of the stria showed ultrastructural abnormalities. Spiral ganglion cells and basal cells of the stria vascularis remained intact in all subjects. This study provides unique information regarding the histology and pathophysiology of meningogenic deafness. The clinical implications of these findings are discussed, with an emphasis on potentially reversible changes and therapeutic intervention.     

The influence of A chronic vitamin a deficiency on the rat cochlea

Summary Rats were fed a vitamin-A-free diet and the cochleas of these animals were studied under the light and electron microscope. The cochlear function was tested by means of the electrocochleography. No pathologic changes could be found in the organ of Corti; the stria vascularis and the cochleas showed normal development. The recordings of the deficient rats are identical in configuration to controls, but they are shifted to higher intensities due to an otitis media found in the tested animals. These findings suggest that vitamin A does not have an important function in the inner ear.Presented at the 18th Workshop on Inner Ear Biology in Montpellier/La Grande Motte, September 14–16, 1981     

Effect of estrogen and antiestrogens on the estrogen receptor content in the cochlea of ovariectomized rats

Older women in the normal population tend to develop less severe hearing loss as compared to males in the same age. In Turner syndrome (45,X), estrogen deficiency is one of the predominant problems. Ear and hearing problems are common among these patients. Does estrogen have an impact on the hearing organ? Twenty-four rats were ovariectomized and treated with vehicle (controls), estradiol or selective estrogen receptor modulators such as tamoxifen and ICI182780, in order to study the effects on the estrogen receptor levels and distribution in the inner ear. The cochleas were stained immunohistochemically using antibodies against estrogen receptor alpha and beta. No major difference in estrogen receptor content in the cochleas was observed among groups. There was however a potential down regulation of estrogen receptor alpha in the marginal cells of stria vascularis in the rats that were substituted with ICI182780 (pure antiestrogen) as compared to those given estradiol or tamoxifen. When investigating the tissues with light microscopy no change in inner ear anatomy could be observed.     

The histologic study of the inner ear in guinea pigs on anaphylatoxin

Complement is known to relate to many inflammatory reactions. C4a, C3a and C5a, known as anaphylatoxins, are known to cause strong inflammatory reactions. In this study, the role of anaphylatoxins on the pathogenesis in the cochlea was examined. On hundred forty six male Harley guinea pigs, weighing about 350 grs, all susceptible to preyer's reflex, were used in this study. Anaphylatoxins were made from guinea pig serum treated with zymosan, and inoculated into the carotid artery of the guinea pigs. Parts of these animals were sacrificed and examined at ten minutes, one day, two days, three days, seven days, ten days and fifteen days after injection of anaphylatoxins. Pathological changes in inner ears were observed by light microscopy. After 10 minutes, inner ears were found morphologically normal. After one day, inner ears were found to be almost morphologically normal but the stria vascularis was observed with cystic formation. After two days, cystic formations in the stria vascularis were enlarged and Reissner's membranes were collapsed in some other animals. After three days, the stria vascularis in the various cochlear turns except in the basal turn, were extremely atrophied, some cochlear nerves showed degeneration and some cochlea showed endolymphatic hydrops. After seven days, ten days and fifteen days, the morphological changes showed atrophy in the stria vascularis similar to the results observed on the third days. Atrophy in the stria vascularis was improved gradually with time, but the degeneration of the cochlear nerve was not improved. Opinions have been divided on the cause of inner ear disease including Meniere's disease. Many authors have reported that infectious diseases, for example mumps, measles and cytomegalovirus infection, have caused human sensorineural hearing loss. These diseases have been reported to result in atrophy in the stria vascularis, degeneration of the cochlear nerve and some other pathological changes. In this study, it was clearly observed that the atrophy of the stria vascularis, the endolymphatic hydrops and other morphological changes were caused by introduction of anaphylatoxins. These results were similar to the pathological changes observed in inner ear diseases in human beings. Therefore, inflammatory substances, including anaphylatoxins, were closely related to the cause of inner ear diseases. The animal model used in this report is considered to be important for elucidating the pathogenesis of inner ear diseases.     

川芎嗪对庆大霉素耳中毒耳蜗外毛细胞和血管纹的保护作用

目的：探讨在庆大霉素（gentamycin，GM）耳中毒情况下，川芎嗪（tetramethylpyrazine，TMP）对豚鼠耳蜗外毛细胞和血管纹边缘细胞的保护作用。方法：12只豚鼠随机分为GM组、联合用药组、TMP组及对照组，用药十天后处死，采用透射电镜观察耳蜗外毛细胞及血管纹边缘细胞超微结构，扫描电镜观察血管纹边缘细胞表面形态。结果：透射和扫描电镜显示，联合用药组外毛细胞及血管纹边缘细胞超微及表面结构破坏不均明显轻于庆大霉素组，特别是其中的线粒体结构破坏与数目减少更显著轻于庆大霉素组。结论：川芎嗪具有保护庆大霉素耳中毒耳蜗外毛细胞和血管纹结构的作用，从而拮抗庆大霉素耳毒性。     

Effects of type 2 diabetes mellitus on cochlear structure in humans

OBJECTIVE: To evaluate the effects of type 2 diabetes mellitus on cochlear elements in humans. DESIGN: Comparative study of the histopathologic characteristics of human temporal bones. SETTING: Otopathology laboratory in a tertiary academic medical center. PATIENTS: Temporal bones from 18 patients with type 2 diabetes mellitus were divided into 2 groups according to the method of management of diabetes: insulin in 11 patients (mean age, 51.9 years; age range, 44-65 years) and oral hypoglycemic agents in 7 patients (mean age, 54.4 years; age range, 45-64 years). The diabetic groups and 26 age-matched controls (mean age, 52.9 years) were examined using light microscopy, and the cochlear changes were compared between groups. MAIN OUTCOME MEASURES: Morphometric measurements of vessel wall thickness in the basilar membrane and stria vascularis were made in all turns of the cochlea at the midmodiolar level. Area measurements of the stria vascularis were made in all turns of the cochlea at the midmodiolar level. Cochlear reconstructions and standard cytocochleograms were prepared using an oil immersion objective. The number of spiral ganglion cells was determined for each segment of the cochlea. Comparisons were made in each segment between diabetic and control groups. RESULTS: In the insulin group, walls of the vessels of the basilar membrane and stria vascularis in all turns were significantly thicker than those of controls. Walls of the vessels of the stria vascularis in the basal turn were also significantly thicker in the oral hypoglycemic group than in controls. Atrophy of the stria vascularis in most turns of the insulin group and the lower middle turn of the oral hypoglycemic group was significantly greater than in the controls. Loss of cochlear outer hair cells was significantly greater in the lower and upper basal turns in both diabetic groups. No significant difference was found in the number of spiral ganglion cells or inner hair cells between groups. CONCLUSION: This study demonstrates that cochlear microangiopathy and degeneration of the stria vascularis and cochlear outer hair cells are found in patients with type 2 diabetes mellitus.