In vitro formation of bilayer composite graft from chondrocytes and bone-marrow-derived stromal cells

软骨细胞和骨髓基质细胞双层复合移植物的体外形成

基本信息

批准号：
15591564
负责人：
WADA Yuichi
金额：
$ 2.11万
依托单位：
Chiba University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

Recently, several strategies using scaffold, such as collagen gel and three-dimensional polymer, have been developed for the production of cartilaginous tissue in vitro. However, some patients desire a new technique of osteochondral implantation without artificial or animal-tissue-derived scaffold. The purpose of this study is to establish in vitro formation of bilayer composite graft from chondrocytes and bone-marrow-derived stromal cells for the use of osteochondral implantation. The alginate-recovered-chondrocyte method(ARC method), which is a novel two-step scaffold-free culture system to produce cartilaginous tissue, was used in this study. Stromal cells from rabbit bone marrow were seeded in high density on the tissue culture insert, followed by three-dimensional chondrocyte culture using the ARC method on the insert. Obtained bilayer tissue, consists of upper cartilage layer and lower bone marrow cell layer, were examnined with both biochemical and histological analysis. When cell number of bone marrow cells was increased, proteoglycan content of the formed tissue was decreased, whereas its collagen content did not change significantly. The composite graft cultured for 4 weeks was hard enough to be held with forceps, and stiffer than regular ARC tissue that was simply made of chondrocytes. The cartilage layer of the tissue was stained well with Safranin-O and anti-type II collagen antibody. The bilayer structure was maintained at 4 weeks after transplantation under the skin of athymic nude mice, while ossification was not observed in the layer of bone marrow cells. This study showed that, for the first time, in vitro formation of osteochondral composite graft was possible without using scaffold. For the clinical application, farther investigation may be needed, especially to achieve proper bone formation in the marrow cell layer.

最近，已经开发了几种使用支架的策略，例如胶原蛋白凝胶和三维聚合物，用于在体外生产软骨组织。但是，一些患者希望一种新的骨软骨植入技术，而没有人工或动物组织衍生的脚手架。这项研究的目的是在体外形成来自软骨细胞和骨髓衍生的基质细胞的双层复合移植物，用于使用骨软骨植入。这项研究使用了藻酸盐恢复的 - 核细胞法（ARC方法），它是一种新型的两步无支架培养系统，用于产生软骨组织。将来自兔骨髓的基质细胞高密度在组织培养物插入物上播种，然后使用插入物上的ARC方法进行三维软骨细胞培养。获得的双层组织，由上软骨层和下骨髓细胞层组成，并通过生化和组织学分析检查。当增加骨髓细胞的细胞数量时，蛋白聚糖的蛋白酶含量减少，而其胶原蛋白含量没有显着变化。与仅由软骨细胞制成的常规弧组织相比，培养4周的复合移植物足够困难，可以用镊子固定和更硬。 safranin-O和抗型II胶原蛋白抗体对组织的软骨层染色很好。双层结构在移植后4周保持在无胸腺裸鼠的皮肤下，而在骨髓细胞层中未观察到骨化。这项研究表明，在不使用脚手架的情况下，可能首次可以在体外形成骨软骨复合材料移植物。对于临床应用，可能需要进行更远的研究，尤其是为了在骨髓细胞层中实现适当的骨形成。

项目成果

期刊论文数量（20）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Regenerative medicine as a treatment of knee injury.

再生医学作为膝关节损伤的治疗方法。

DOI：
发表时间：
2004
期刊：
Bone Joint and Tendon 17
影响因子：
0
作者：
Seto H;Tanaka S.;Nakamura K;et al.;和田佑一;和田佑一;Yuichi Wada
通讯作者：
Yuichi Wada

K Tsuchiya, Y Wada, etc.: "Effects of HAS2 on N-cadherin expression during chondrogenic cellular condensation process in vitro"Osteoarthritis and Cartilage. 11(A). 90-91 (2003)

K Tsuchiya，Y Wada等：“体外软骨细胞凝结过程中HAS2对N-钙粘蛋白表达的影响”骨关节炎和软骨。

DOI：
发表时间：
期刊：
影响因子：
0
作者：
通讯作者：

Chondrocyte implantation for the repair of cartilage defect.

软骨细胞植入用于修复软骨缺损。

DOI：
发表时间：
2003
期刊：
Orthopedic and traumatic surgery 46
影响因子：
0
作者：
Seto H;Tanaka S.;Nakamura K;et al.;和田佑一;和田佑一;Yuichi Wada;Yuichi Wada;和田佑一;和田佑一;和田佑一;Yuichi Wada
通讯作者：
Yuichi Wada

Autologous chondrocyte implantation.