Reconstruction of basement membranes aiming at artificial organs

针对人工器官的基底膜重建

基本信息

批准号：
10557113
负责人：
NINOMIYA Yoshifumi
金额：
$ 8.26万
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：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

Ultimate goal of the research project is to make artificial organs to insert basement membrane materials between epithelial layers and extracellular matrix. In order to make type IV collagen molecules in vitro, we we did the following investigations :* Molecular composition of basement membranes underneath smooth muscle cells varies in different organs, which could be related to specific function of the individual organs.* We first isolated and characterized genomic DNA fragments that cover the 5'flanking sequences of COL4A3 and COL4A4 genes, which provided information to delineate the promoter activity for the tissue-specific expression of the six type IV collagen genes.* Molecular forms of collagen IV in follicle basement membranes are different in development.* We identified interactions between the α2(IV) and ProMMP-9 by immunoprecipitations and blotting.* Basement membrane collagen IV molecules diminish in parallel with malignancy and invasiveness when analyzed in tumor progression.* Although mRNA expression level of α5 decreased in kidney of a canine Alport case, that of α6 resulted in unchanged. But the protein level of both chains cannot be found any. These results suggest a possibility of a molecular form of α5/α6.* Differential expression of collagen IV genes in epithelial basement membranes was analyzed in detail.* Here we define a novel function for soluble non-collagenous (NC1) domains of the α2(IV), α3(IV), and α6(IV) chains of human collagen type IV in the regulation of angiogenesis and tumor growth. These NCI domains were shown to regulate endothelial cell adhesion and migration by distinct αv and β1 integrin-dependent mechanisms.* In Lmx1b(-/-) mice, expression of both α3 and α4 is strongly diminished in GBM, whereas that of α1, α2 and α5 is unchanged. Moreover, LMX1B binds specifically to a putative enhancer in intron 1 of mouse/human COL4A4 and upregulates reporter constructs containing the enhancer-like sequence.

该研究项目的最终目标是制造人造器官，将基底膜材料插入上皮层和细胞外基质之间。为了在体外制造 IV 型胶原分子，我们进行了以下研究：* 平滑肌下方基底膜的分子组成。不同器官中的细胞有所不同，这可能与各个器官的特定功能有关。*我们首先分离并表征了覆盖 COL4A3 和 COL4A4 基因 5' 侧翼序列的基因组 DNA 片段，该片段提供了描述六种 IV 型胶原蛋白基因的组织特异性表达的启动子活性。* 卵泡基底膜中 IV 型胶原蛋白的分子形式在发育过程中是不同的。* 我们通过免疫沉淀和印迹鉴定了 α2(IV) 和 ProMMP-9 之间的相互作用* 在肿瘤进展过程中进行分析时，基底膜 IV 型胶原蛋白分子与恶性肿瘤和侵袭性同时减少。* 尽管犬 Alport 病例肾脏中 α5 的 mRNA 表达水平下降，但 α6 的 mRNA 表达水平没有变化。两条链的蛋白质水平均未发现。这些结果表明可能存在 α5/α6 的分子形式。*详细分析了上皮基底膜中 IV 型胶原蛋白基因的差异表达。*在这里，我们定义了可溶性非胶原蛋白的新功能。 -人 IV 型胶原蛋白的 α2(IV)、α3(IV) 和 α6(IV) 链的胶原 (NC1) 结构域在调节血管生成和肿瘤生长中的作用这些 NCI 结构域显示可调节内皮细胞。通过不同的 αv 和 β1 整合素依赖性机制实现粘附和迁移。*在 Lmx1b(-/-) 小鼠中，GBM 中 α3 和 α4 的表达均大幅减少，而 α1、α2 和 α5 的表达则没有变化。此外，LMX1B 结合。特异性针对小鼠/人 COL4A4 内含子 1 中的推定增强子，并上调含有增强子样序列的报告构建体。