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Development of the endometrial spheroid using tissue-engineering technique

利用组织工程技术开发子宫内膜球体

基本信息

批准号：
16580232
负责人：
YAMAUCHI Nobuhiko
金额：
$ 2.3万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

The purpose of the present study is to develop the spheroid composed of rat endometrial stromal cells (RES) as an in vitro model to analyze functions of rat endometrium. Spheroids were generated using salmon aterocollagen (SAC). SAC has a low denaturation temperature and need to be cross-linked before being used as a scaffold. In the present study, SAC was cross-linded by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride or dehydrothermal treatment. It seems that fish collagen has lower risk for transmission of infectious deseases.The RES were separated and purified from rat endometrium at day 5 of pregnancy. The RES were cultured up to the confluent state on SAC. Then digested SAC used collagenase. Then RES cell sheet cultured in agarose coated plate to generate a spheroid. After collagenase treatment, the floating cell-sheet shrank and became an aggregated cell mass in a few days ; it finally formed a round-shaped hetero-spheroid composed of RES. When cell viability was examined with TUNEL (terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling), apoptotic cells were not found in the spheroid for 15 days. Immunofluorescent observations used the proliferating cell nuclear antigen (PCNA) antibody showed that proliferating cells were not find in the spheroids. The production of proMMP-2 in the spheroid was similar to that produced by endometrial tissue in vivo.These results indicate that rat endometrial stromal cells are capable of being regenerated as a multicellular spheroid using SAC in vitro. As the present spheroid displays an endometrium-mimic feature in structural and functional similarities, it seems to be a useful in vitro model of rat endometrium. The present method being simple and convenient to form multicellular spheroid from RES, it provides a new sight in research of endometrial stromal functions and implantation.

本研究的目的是建立大鼠子宫内膜基质细胞（RES）组成的球体，作为分析大鼠子宫内膜功能的体外模型。使用鲑鱼缺角胶原（SAC）产生球体。SAC具有低变性温度，并且在用作支架之前需要交联。本研究采用1-乙基-3-（3-二甲氨基丙基）碳二亚胺盐酸盐或去水热处理对SAC进行交联。从妊娠第5天的大鼠子宫内膜中分离纯化了RES。将RES在SAC上培养至汇合状态。然后用胶原酶消化SAC。然后将RES细胞片在琼脂糖包被的平板中培养以产生球状体。胶原酶处理后，漂浮的细胞片收缩，并在几天内成为一个聚集的细胞团，它最终形成了一个圆形的异质球体组成的RES。当细胞活力与TUNEL（末端脱氧核苷酸转移酶介导的dUTP-生物素缺口末端标记）检查，凋亡细胞没有发现在球体中15天。增殖细胞核抗原（PCNA）抗体免疫荧光观察显示，球状体内未见增殖细胞。结果表明，大鼠子宫内膜间质细胞在SAC的作用下，可在体外形成多细胞球状体，其proMMP-2的表达量与体内子宫内膜组织相似。由于本球体在结构和功能上显示出类似于子宫内膜的特征，它似乎是一种有用的大鼠子宫内膜体外模型。本方法简便易行，可使RES形成多细胞球体，为研究子宫内膜间质功能和种植提供了新的视角。

项目成果

期刊论文数量（42）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Development of multidrug resistance type 1 P-glycoprotein function during in vitro maturation of porcine oocyte.

猪卵母细胞体外成熟过程中多药耐药1型P-糖蛋白功能的发展。

DOI：
发表时间：
2006
期刊：
Reproductive Toxicology 21
影响因子：
0
作者：
Arai M;Yamauchi N;Fukuda H;Soh T;Hattori M-A
通讯作者：
Hattori M-A

Evaluation of RNA interference in developing porcine granulosa cells using fluorescence reporter genes

DOI：
10.1262/jrd.50.599
发表时间：
2004-10-01
期刊：
JOURNAL OF REPRODUCTION AND DEVELOPMENT
影响因子：
1.8
作者：
Hirano, T;Yamauchi, N;Hattori, M
通讯作者：
Hattori, M

CHANGES OF NITRIC OXIDE SYNTHASES EXPRESSION IN RAT UTERUS CORRELATED WITH ESTROUS CEYCLE AND PREGNANCY

大鼠子宫中一氧化氮合酶表达的变化与动情周期和妊娠相关

DOI：
发表时间：
2005
期刊：
Reproduction (発表予定)
影响因子：
0
作者：
T.Honda;T.Nomura;M.Mukai;Shinya Aramaki et al.;T.Nomura;Toru Gamo;T.Nomura;Kyohei Nishimura
通讯作者：
Kyohei Nishimura

Evaluation of RNA interference in developing porcine granulos cells using fluorescence reporter genes.

使用荧光报告基因评估猪颗粒细胞发育过程中的 RNA 干扰。

DOI：
发表时间：
2004
期刊：
Journal of Reproduction and Development 50
影响因子：
0
作者：
T.Honda;T.Nomura;M.Mukai;Shinya Aramaki et al.;T.Nomura;Toru Gamo;T.Nomura;Kyohei Nishimura;T.Nomura;Noriko Shibata;T.Nomura;T.Nomura;Takurou Hirano et al.
通讯作者：
Takurou Hirano et al.

Tumor necrosis factor-α stimulates proliferation activity and prostaglandin production of rat endometrial stromal cells in vitro.

肿瘤坏死因子-α 在体外刺激大鼠子宫内膜基质细胞的增殖活性和前列腺素产生。