Molecular mechanism of otolith microstructure formation : immunohistochemical localization of two matrix proteins and expressions of tneir mRNAs

耳石微结构形成的分子机制：两种基质蛋白的免疫组织化学定位及其mRNA的表达

• 批准号: 13660178
• 负责人: TAKAGI Yasuaki
• 金额: $ 1.34万
• 依托单位: HOKKAIDO UNIVERSITY (2002)The University of Tokyo (2001)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13660178/
• 关键词: Otolith; Organic matrix; Calcification; Biomineralization; Calcium carbonate; Age determination; Daily age determination; ベイオミネラリゼーション

1. Identification of otolith-matrix-producing cells and localization of matrix proteins in the otolith : Specific antibodies against the major proteins of otolith matrix, OMP-1 and otolin-1, were raised and immunohistochemistry was conducted. The squamous epithelial cells and the transitional epithelial cells of the inner ear sacculus were identified as OMP-1-producing cells, whereas the cells located at the most periferal region of the sensory epithelium were identified as otolin-1-producing cells. The OMP-1 was contained in the otolith only, whereas the otolin-1 was included in both the otolith and the otolith membrane (gelatinous layer). The electron microscopy revealed that the otolith was composed of two types of the organic matrices, a dense sheet-like matrix and a fine meshwork matrix. Both OMP-1 and otolin-1 were detected in the meshwork. Differences in the density of the meshwork matrix makes daily rings, thus it is possible that the daily changes in the rate of production and … More /or deposition of OMP-1 and otolin-1 closely relate to the formation of the daily rings.2. Development of real-time quantitative PCR system for OMP-1 and otolin-1 mRNA expression : We have developed the real-time PCR system to quantify OMP-1 and otolin-1 mRNA expression levels from the left and right sacculi of the individual rainbow trout. The daily changes of the expression levels of OMP-1 and otolin-1 mRNA were now under the investigation.3. Determination of the supersaturation ratio of the inner ear endolymph : Otolith is composed of calcium carbonate poly crystals crystallized from the endolymph. Therefore, the growth rate of the otolith calcium carbonate must be highly dependent on the supersaturation ratio (S_a) of the endolymph. We have measured individually the ionic composition of the endolymph and calculated the S_a. The endolymph S_a was 2.027-4.303, clearly indicating that the endolymph is supersaturated with respect to the aragonite. The S_a was closely related to the pH of the endolymph, thus, pH-regulation of the endolymph is crucial for the otolith growth. Less

1.耳石基质产生细胞的鉴定和耳石中基质蛋白的定位：针对耳石基质的主要蛋白OMP-1和otolin-1的特异性抗体被提出并进行免疫组织化学。内耳球囊的鳞状上皮细胞和移行上皮细胞被鉴定为OMP-1产生细胞，而位于感觉上皮的最外周区域的细胞被鉴定为otolin-1产生细胞。OMP-1仅包含在耳石中，而otolin-1包含在耳石和耳石膜（凝胶层）中。电镜观察表明，耳石由致密的片状基质和细小的网状基质两种有机基质组成。OMP-1和otolin-1均表达于神经网络中。在网络矩阵的密度的差异，使日常环，因此，这是可能的，每天的变化率的生产和 ...更多信息 OMP-1和otolin-1的沉积与日年轮的形成密切相关. OMP-1和otolin-1 mRNA表达的实时定量PCR系统的开发：我们已经开发了实时PCR系统来定量来自个体虹鳟鱼的左右球囊的OMP-1和otolin-1 mRNA表达水平。OMP-1和otolin-1 mRNA表达水平的日变化正在研究中.内耳内淋巴液过饱和度的测定：耳石由内淋巴液结晶的碳酸钙多晶体组成。因此，耳石碳酸钙的生长速率与内淋巴过饱和度（S_a）密切相关。我们分别测定了内淋巴的离子组成，并计算了S_a。内淋巴的S_a值为2.027-4.303，表明内淋巴对文石的过饱和状态。S_a与内淋巴的pH值密切相关，因此，内淋巴pH值的调节对耳石的生长至关重要。少

项目成果

Murayama, E.: "Two matrix proteins from fish otolith. How does the otolith grow?"Biomineralization : formation, diversity, evolution and application, I. Kobayashi and H. Ozawa eds.. in press.

Murayama, E.：“来自鱼耳石的两种基质蛋白。耳石如何生长？”生物矿化：形成、多样性、进化和应用，I. Kobayashi 和 H. Ozawa 编辑。正在出版。

村山英美, 都木靖彰, 長澤寛道: "魚類の耳石はいかにして形成されるか?結晶成長を制御するタンパク質,OMP-1とotolin-1"化学と生物. 41. 11-12 (2003)

Hidemi Murayama、Yasuaki Tsuki、Hiromichi Nagasawa：“鱼中的耳石是如何形成的？控制晶体生长的蛋白质 OMP-1 和 otolin-1”《化学与生物学》41. 11-12 (2003)。

Murayama, E.: "Fish otolith contains a unique structural protein, otolin1"European Journal of Biochemistry. 269. 688-696 (2002)

Murayama, E.：“鱼耳石含有一种独特的结构蛋白，otolin1”欧洲生物化学杂志。

Murayama, E., Takagi, Y.et al.: "Fish otolith contains a unique structural protein, otolin-1"European Journal of Biochemistry. 269. 688-696 (2002)

Murayama, E., Takagi, Y.等人：“鱼耳石含有一种独特的结构蛋白，otolin-1”欧洲生物化学杂志。

Muyayama, E., Takagi, Y. et al.: "Fish otolith contains a unique structural protein, otolin-1"European Journal of Biochemistry. (in press).

Muyayama, E., Takagi, Y. 等人：“鱼耳石含有一种独特的结构蛋白，otolin-1”欧洲生物化学杂志。