Virtual bone reflecting microstructure ofreal bone

虚拟骨骼反映真实骨骼的微观结构

• 批准号: 18592234
• 负责人: KAMIOKA Hiroshi
• 金额: $ 2.48万
• 依托单位: Okayama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18592234/
• 关键词: Osteocvte; Ultra high voltage electron microscopy; tomography; Three dimensional reconstruction; Virtual bone; simulation; hone

Aim: We applied electron tomography to the osteocytes in bone in order to make virtual bone.Materials & Methods: Chick calvariae were desected from 16-day old chick embryo. The calvariae were treated by 1 mM collagenase and 5 mM EDTA and trimmed in 3x3-mm for further use. These sections were fixed with 3% paraformaldehyde for 24h, stained with 2% protargol solution containing copper beads for 48h as silver-staining, and embedded in paraffin. After these treatment, 2 μm sections were placed on fromvar coated cupper grids. For tomographic data collection, sections were dipped in a solution of colloidal gold for 1-2 min. Electron tomographic data collection was carried out with H-3000 ultra-high voltage EM (Hitachi) at 2 MV equipped with a 360 °-tilt specimen holder. Images were recorded on a Tietz Tem-Cam 4k x 4k-pixel F415S slow scan CCD camera (TVIPS) binned to 2k x 2k pixels. Alignment of the tilt series and reconstruction of 3D volumes from aligned images was performed by the weighted back-projection method by using a tomography software package IMOD.Results: Stereo-pair image of osteocyte was taken by ultrahigh-voltage electron microscopy with tilt +/-7 °. Osteocyte clearly appeared with numerous processes and brunches. The tomograms were generated from two +1-60 ° tilt series of a 2.0 μm section of chick calvaria and the microgram were taken 2 ° intervals. We could observe the series with 0.3 μm interval in depth of osteocyte tomography. The final pixel size on the specimen was 8 nm. We could distinguish the osteocyte cell body and their processes with high-density dots of silver on the surface of cell membrane.Conclusions: It is possible to observe silver-stained osteocytes in thick section (2 μm) by ultrahigh-voltage electron microscopy. In addition, high-resolution (8 nm) electron tomography was performed to the osteocytes in chick calvariae.

目的：对骨中的骨细胞进行电子断层扫描，以制作虚拟骨。材料与方法：从16日龄鸡胚中取出鸡颅骨。颅骨用 1 mM 胶原酶和 5 mM EDTA 处理，并修剪成 3x3 毫米以供进一步使用。将这些切片用3%多聚甲醛固定24小时，用含有铜珠的2%普他戈溶液银染48小时，并包埋在石蜡中。这些处理后，将 2 μm 切片放置在涂有弗瓦尔涂层的铜网上。为了收集断层扫描数据，将切片浸入胶体金溶液中 1-2 分钟。电子断层扫描数据采集是使用 H-3000 超高压 EM（日立）在 2 MV 下进行的，配备有 360° 倾斜样品架。图像记录在 Tietz Tem-Cam 4k x 4k 像素 F415S 慢扫描 CCD 相机 (TVIPS) 上，并合并为 2k x 2k 像素。使用断层扫描软件包 IMOD，通过加权反投影方法进行倾斜系列的对准和从对准图像重建 3D 体积。结果：通过倾斜 +/-7° 的超高压电子显微镜拍摄骨细胞的立体对图像。骨细胞清楚地出现，具有许多突起和分支。断层扫描图像是由小鸡颅骨 2.0 μm 切片的两个 +1-60° 倾斜系列生成的，微照片以 2° 间隔拍摄。我们可以观察骨细胞断层扫描深度为0.3μm间隔的系列。样本上的最终像素尺寸为 8 nm。通过细胞膜表面高密度的银点可以区分骨细胞的胞体及其突起。结论：超高压电镜可以在厚切片（2μm）上观察到银染的骨细胞。此外，对小鸡颅骨中的骨细胞进行了高分辨率（8 nm）电子断层扫描。

项目成果

Interface Oral Health Science 2007 Less response of osteocyte than osteoblasts to mechanical force: Implication of different focal adhesion formation.

Interface Oral Health Science 2007 骨细胞对机械力的反应小于成骨细胞：不同粘着斑形成的含义。

• 发表时间: 2007
• 作者: Takano-Yamamoto T;Kamioka H;Sugawara Y
• 通讯作者: Sugawara Y

Mechanosensitivity of A Single Osteocyte : Difference in Cell Process and Cell Body

单个骨细胞的机械敏感性：细胞突起和细胞体的差异

• 发表时间: 2007
• 作者: AONUMA YUKI;ADACHI TAIJI;TANAKA MOTOTSUGU;HOJO MASAKI;TAKANO-YAMAMOTO TERUKO;KAMIOKA HIROSHI
• 通讯作者: KAMIOKA HIROSHI

Regulator of cell-cell communication among osteocytes in bone

骨中骨细胞之间细胞间通讯的调节剂

• 发表时间: 2007
• 作者: Ishihara;Y.;Kamioka;H.;Yamamoto;T.;Yamashiro;T
• 通讯作者: T

Mechanosensitivity of A Single Osteocyte: Difference in Cell Process and Cell Body

单个骨细胞的机械敏感性：细胞过程和细胞体的差异

• 发表时间: 2007
• 期刊: Journal of Biomechanical Science and Engineering Vol.2, Supp1.1
• 作者: AONUMA YUKI;ADACHI TAIJI;TANAKA MOTOTSUGU;HOJO MASAKI;TAKANO-YAMAMOTO TERUKO;KAMIOKA HIROSHI
• 通讯作者: KAMIOKA HIROSHI

Primary cultures of chick osteocytes retain functional gap junctions between osteocytes and between osteocytes and osteoblasts

• DOI: 10.1017/s143192760707016x
• 发表时间: 2007-04-01
• 期刊: MICROSCOPY AND MICROANALYSIS
• 影响因子: 2.8
• 作者: Kamioka, Hiroshi;Ishihara, Yoshihito;Lim, Soo-Siang
• 通讯作者: Lim, Soo-Siang