Immunogold TEM of otoconin 90 and otolin - relevance to mineralization of otoconia, and pathogenesis of benign positional vertigo
Immunogold TEM of otoconin 90 and otolin - relevance to mineralization of otoconia, and pathogenesis of benign positional vertigo
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DOI：
10.1016/j.heares.2012.07.003
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发表时间：
2012-10-01
期刊：
影响因子：
2.8
通讯作者：
Thalmann, Ruediger
中科院分区：
文献类型：
--
作者：
Andrade, Leonardo R.;Lins, Ulysses;Thalmann, Ruediger
深度蚀刻技术的实施使耳锥的亚结构元素得到了前所未有的定义，包括内核及其球状附着物的纤细网络。随后测定了主要的可溶性耳锥蛋白Otoconin 90对方解石晶体体外生长的影响，包括成核速度的增加、生长动力学的抑制和显著的形态变化。合理的下一步，耳胶蛋白90的超微结构定位，通过免疫金电子显微镜在年轻成熟的小鼠身上，显示出尽管矿化程度相对较低，但内核仍有高密度的金粒子。在这里，金颗粒通常排列成椭圆形图案，这意味着耳胶90附着在由六角纤维网络组成的支架上，这是耳石的特征。外层皮质矿化程度较高，矿化纤维束平行于地表排列。脱钙后，金颗粒和基质纤维被认为是由耳胶的线性结构表型组成的，它们在相同的方向上排列，这表明它们是引导耳胶蛋白90介导的矿化的支架。在多面尖端，矿化程度最高，尽管金颗粒的密度相对较低，这可能是由于致密矿物相的置换作用所致。透射电子显微镜显示，单个微晶聚集成等向柱状物。柱子排列成平行的片层，转化成矿化的块，分级组装成复杂的耳锥镶嵌体。另一组基于免疫金电子显微镜的幼鼠实验表明，连接耳锥的纤维由短链胶原耳胶组成。到两岁时，小鼠耳锥(相当于中年人类)的表层已经脱矿，导致连接耳锥的纤维锚定减弱或丧失。因此，微小的机械干扰可以将彼此分离的耳石释放到内淋巴间隙中。在人类中，良性位置性眩晕(BPV)被认为是由于耳锥从内淋巴管移位到半圆形管内，使其受体容易受到重力的刺激而导致严重的眩晕发作。这些在人类身上的观察结果，再加上目前的动物实验，为BPV的早期阶段提供了一个初步的致病基础。(C)2012爱思唯尔B.V.保留所有权利。
Implementation of the deep-etch technique enabled unprecedented definition of substructural elements of otoconia, including the fibrillar meshwork of the inner core with its globular attachments. Subsequently the effects of the principal soluble otoconial protein, otoconin 90, upon calcite crystal growth in vitro were determined, including an increased rate of nucleation, inhibition of growth kinetics and significant morphologic changes. The logical next step, ultrastructural localization of otoconin 90, by means of immunogold TEM in young mature mice, demonstrated a high density of gold particles in the inner core in spite of a relatively low level of mineralization. Here gold particles are typically arranged in oval patterns implying that otoconin 90 is attached to a scaffold consisting of the hexagonal fibrillar meshwork, characteristic of otolin. The level of mineralization is much higher in the outer cortex where mineralized fiber bundles are arranged parallel to the surface. Following decalcification, gold particles, as well as matrix fibrils, presumed to consist of a linear structural phenotype of otolin, are aligned in identical direction, suggesting that they serve as scaffold to guide mineralization mediated by otoconin 90. In the faceted tips, the level of mineralization is highest, even though the density of gold particles is relatively low, conceivably due to the displacement by the dense mineral phase. TEM shows that individual crystallites assemble into iso-oriented columns. Columns are arranged in parallel lamellae which convert into mineralized blocks for hierarchical assembly into the complex otoconial mosaic.Another set of experiments based on immunogold TEM in young mice demonstrates that the fibrils interconnecting otoconia consist of the short chain collagen otolin. By two years of age the superficial layer of mouse otoconia (corresponding to mid-life human) has become demineralized resulting in weakening or loss of anchoring of the fibrils interconnecting otoconia. Consequently, otoconia detached from each other may be released into the endolymphatic space by minor mechanical disturbances. In humans, benign positional vertigo (BPV) is believed to result from translocation of otoconia from the endolymphatic space into the semi-circular canals rendering their receptors susceptible to stimulation by gravity causing severe attacks of vertigo. The combinations of these observations in humans, together with the presented animal experiments, provide a tentative pathogenetic basis of the early stage of BPV. (c) 2012 Elsevier B.V. All rights reserved.