Cloning, Expression and Identification of Specific Genes related Osseointegration to Titanium.

钛骨整合相关特定基因的克隆、表达和鉴定。

• 批准号: 15390592
• 负责人: KANYAMA Manabu
• 金额: $ 9.47万
• 依托单位: OKAYAMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-15390592/
• 关键词: bone marrow cells; Osteoblasts; Titanium; Osseointegration; xab-2; sod-1; galection-1; デンタルインプラント; チタン特異的遺伝子; 骨芽細胞; 骨組織再生; 幹葉系幹細胞

The aim of this study is to clarify the cell behavior induced by titanium and to isolate specific genes that promote the titanium implant-bone integration. Especially, to identify the master key genes related osseointegration to titanium, the osteoblastic cell line, MC3T3-E1 cells were cultured on titanium coated glasses and the gene expression were analyzed.1)Titanium and chrome coated glassesTi and Cr were coated to polished glass surfaces (33 mm in diameter by 1.5 mm in thickness) in a high vacuum condition. Ti and Cr coated glass disks were placed in a 6 well plate and fixed by silicone rings. Non-coated glass disks were served as controls.2)Cell attachment, proliferation and differentiation assessments.Initial cell attachment and proliferation of MC3T3-E1 cells were estimated by MTS-assay (CellTiter 96【○!R】 AQueous One Solution, Promega, USA), and cell differentiation was evaluated by alkaline phosphatase activity assay. The mean relative amount of the attached cells on the Ti-coa … More ted glass was significantly higher than those on non-coated and Cr-coated glass at 3 hours after seeding. On the same way the mean cell number on Ti-coated glass at 3 days after seeding was significantly higher than those of other conditions. The mean alkaline phosphatase(ALP) activity of the seeded osteoblasts also increased with time in these three conditions, while the ALP activity level on the Ti-coated glass at 14 days after seeding was significantly higher than those of other conditions. These results suggest that, Ti-coated glass plate accelerated the cell attachment, proliferation and differentiation of the MC3T3-E1 cells, compared to the non- and Cr-coated glass plates3)Effect of gene expression of osteoblast on titaniumOsteoblasts were cultured on Ti coated, Cr coated and non-coated glass plates. And then differentially expressed genes were identified by cDNA subtractive hybridization. Twenty independent clones were isolated and by nucleotide sequencing of these clones, seven clones were identified including EST genes ; xab-2,sod-1,galectin-1,actin related protein 2/3 mRNA, RIKEN cDNA 2210013021 gene, EST 601086505F1, and EST 01439. And galectin-1,xab-2,sod-1 gene expression on Ti-coated glass were higher than non-coated, Cr-coated glass.These results suggest that Ti-coating is more advantageous for osteoblastic cell attachment, proliferation and differentiation than Cr or non-coating and galectin-1,xab-2 and sod-1 up-regulation could be related to the Ti-bone integration. Less

本研究的目的是阐明钛诱导的细胞行为，并分离促进钛种植体-骨整合的特异性基因。特别地，为了鉴定与钛的骨整合相关的主要关键基因，成骨细胞系MC 3 T3-E1细胞在钛涂层玻璃上培养并分析基因表达。1）钛和Chrome涂层玻璃在高真空条件下将Ti和Cr涂覆到抛光的玻璃表面（直径33 mm ×厚度1.5 mm）。将Ti和Cr涂覆的玻璃盘置于6孔板中并通过硅酮环固定。2）细胞贴壁、增殖和分化的评价：采用MTS-测定法（CellTiter 96[○！R] AQueous One Solution，Promega，USA），并通过碱性磷酸酶活性测定评价细胞分化。钛板上贴壁细胞的平均相对数量 ...更多信息 在接种后3小时，涂覆的玻璃上的那些显著高于未涂覆和涂覆Cr的玻璃上的那些。同样，接种后3天，钛涂层玻璃上的平均细胞数显著高于其他条件下的平均细胞数。在这三种条件下，成骨细胞的平均碱性磷酸酶（ALP）活性也随着时间的推移而增加，而在钛涂层玻璃上的ALP活性水平在14天后显着高于其他条件。3）钛对成骨细胞基因表达的影响将成骨细胞分别培养在钛涂层、铬涂层和非涂层玻璃板上。然后用cDNA消减杂交技术鉴定差异表达基因。分离出20个独立的克隆，并通过这些克隆的核苷酸测序，鉴定出7个克隆，包括EST基因; xab-2、sod-1、半乳糖凝集素-1、肌动蛋白相关蛋白2/3 mRNA、RIKEN cDNA 2210013021基因、EST 601086505 F1和EST 01439。galectin-1、xab-2、sod-1基因在镀钛玻璃上的表达高于未镀钛玻璃和镀铬玻璃，表明镀钛玻璃比镀铬玻璃或未镀钛玻璃更有利于成骨细胞的附着、增殖和分化，galectin-1、xab-2和sod-1基因表达上调可能与钛-骨结合有关。少

项目成果

骨芽細胞様細胞株(MC3T3E1)細胞の細胞接着・増殖・分化および遺伝子発現に対するチタンの影響

钛对成骨样细胞系（MC3T3E1）细胞粘附、增殖、分化和基因表达的影响

• 期刊: 岡山歯学会雑誌 (印刷中)
• 作者: Wei Hua;Tomotake Yoritoki;Nagao Kan;Oguri Takafumi;Nagao Daisuke;Ichikawa Tetsuo;秋山謙太郎
• 通讯作者: 秋山謙太郎

再生医療と歯科補綴学の接点 補綴治療における再生医療のニーズと現時点での研究動向 -確実かつ質の高いインプラント治療を目指して-

再生医学与口腔修复学的交叉点 再生医学在修复治疗中的需求及当前研究趋势 - 追求可靠、高质量的种植治疗 -

• 发表时间: 2005
• 期刊: 日本補綴歯科学会雑誌 49・4
• 作者: 窪木拓男;窪木拓男
• 通讯作者: 窪木拓男

Effect of Titanium on the Attachment, Proliferation, Differentiation, and Gene Expression of Osteoblastic Cell Line, MC3T3-E1 cells.

钛对成骨细胞系 MC3T3-E1 细胞附着、增殖、分化和基因表达的影响。

• 发表时间: 2006
• 期刊: Academy of Osseointegration 21^<st> Annual Meeting Program Guide 80
• 作者: K.Akiyama;F.Myokai;T.Fujisawa;Y.Yosida;M.Ono;M.Kanyama;S.Takashiba;K.Suzuki;T.Kuboki
• 通讯作者: T.Kuboki

Initial cell attachment, proliferation and differentiation of osteoblast like (MC3T3-Ei) cells to titanium and chrome

成骨细胞样 (MC3T3-Ei) 细胞对钛和铬的初始细胞附着、增殖和分化

• 发表时间: 2005
• 期刊: The 1^<st> Congress of Asian Academy of Osseointegration Program and Abstract
• 作者: Akiyama K;Fujisawa T;Ono M;Yoshida Y;Kanyama M;Suzuki K;Kuboki T
• 通讯作者: Kuboki T