Biological Function of Type VIII Collagen

VIII 型胶原蛋白的生物学功能

• 批准号: 06044154
• 负责人: NINOMIYA Yoshifumi
• 金额: $ 1.6万
• 依托单位: Okayama University Medical School
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for international Scientific Research
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 无数据
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06044154/
• 关键词: Type VIII Collagen; Smooth Muscle Cells; Keratinocytes; Transgenic Mice

In order to know which cells or organs produce type VIII collagen, we extracted RNA from various cells and tissues and performed Northern blot analysis using specific cDNAs. Cultured human aortic endothelial cells were found to keep synthesizing alpha1 (VIII) and alpha2 (VIII) chains until tenth passage. These cells synthesize collagen type I,III,and V at the same time. In separate experiment rat aortic smooth muscle cells were tested whether they produce collagens. What we detected was that the cells synthesized 75 kDa bacterial collagenase sensitive materials. Of interest was that production of this materials was enhanced 20 to 30 times by addition of heparin into the culture medium. Currently we are testing whether this bacterial collagenase sensitive material is the alpha-chain of type VIII by Northern blotting analysis. We also demonstrated that mouse keratinocytes produce collagen VIII at the mRNA level using Northern-blotting and in situ hybridization.Several lines of transgenic mice did show strange expression pattern of type VIII collagen gene during development. It seemed that its pattern was strict and developmentally regulated. However, we did not know whether the pattern was artifact or real, since we did not predict the expression pattern ; transient in certain tissues. Direct in situ hybridization was performed using the specific probes for mouse collagen type VIII at the developmental stage and proved that the expression pattern was real.

为了了解哪些细胞或器官产生 VIII 型胶原蛋白，我们从各种细胞和组织中提取 RNA，并使用特定的 cDNA 进行 Northern blot 分析。培养的人主动脉内皮细胞被发现持续合成 alpha1 (VIII) 和 alpha2 (VIII) 链直至第十代。这些细胞同时合成 I、III 和 V 型胶原蛋白。在单独的实验中，测试了大鼠主动脉平滑肌细胞是否产生胶原蛋白。我们检测到细胞合成了 75 kDa 细菌胶原酶敏感材料。有趣的是，通过在培养基中添加肝素，这种材料的产量提高了 20 至 30 倍。目前我们正在通过Northern印迹分析测试这种细菌胶原酶敏感材料是否为VIII型α链。我们还通过 Northern 印迹和原位杂交证明，小鼠角质形成细胞在 mRNA 水平上产生 VIII 型胶原蛋白。几个转基因小鼠品系在发育过程中确实表现出了 VIII 型胶原蛋白基因的奇怪表达模式。看来它的模式是严格的、发展规律的。然而，我们不知道该模式是人为的还是真实的，因为我们没有预测表达模式；在某些组织中短暂存在。使用发育阶段小鼠VIII型胶原蛋白的特异性探针进行直接原位杂交，证明表达模式是真实的。

项目成果

Inoguchi K.,Yoshioka H.,Khaleduzzaman M.,and Ninomiya Y.: "The mRNA for the α(XIX)collagen chain,a new member of FACITs,contain a long unusual 3′ untranslated region and displays many unique splicing variants." J.Biochem.(Tokyo). 117. 137-146 (1995)

Inoguchi K.、Yoshioka H.、Khaleduzzaman M. 和 Ninomiya Y.：“FACIT 的新成员 α(XIX) 胶原链的 mRNA 包含一个长的不寻常的 3 非翻译区，并显示出许多独特的剪接变体。 “ J.Biochem.（东京）。117. 137-146（1995）

Sugimoto M.,Oohashi T.,and Ninomiya Y.: "The two genes,COL4A5 and COL4A6,coding for the two new basement membrane collagen chains,α5(IV) and α6(IV),are located head-to-head in close proximity on chromosome Xq22 and COL4A6 is transcribed from two alternate

Sugimoto M.、Oohashi T. 和 Ninomiya Y.：“编码两个新基底膜胶原链 α5(IV) 和 α6(IV) 的两个基因 COL4A5 和 COL4A6 头对头位于染色体 Xq22 和 COL4A6 上的紧密接近是由两个备用转录的

Inoguchi K., Yoshioka H., Khaleduzzaman M., and Ninomiya Y.: "The mRNA for the alpha1 (XIX) collagen chain, a new member of FACTTs, contain a long unusual 3'untranslated region and displays many unique splicing variants." J.Biochem. (Tokyo). 117. 137-146

Inoguchi K.、Yoshioka H.、Khaleduzzaman M. 和 Ninomiya Y.：“α1 (XIX) 胶原链的 mRNA 是 FACTT 的新成员，包含一个长的不寻常的 3 非翻译区，并显示出许多独特的剪接变体。

Yoshioka H.,Greenwel P.,Inoguchi K.,Truter S.,Inagaki Y.,Ninimiya Y.,and Ranirez F.: "Structural and functional analysis of the promoter of the human α(XI) collagen gene." J.Biol.Chim.270. 418-424 (1995)

Yoshioka H.、Greenwel P.、Inoguchi K.、Truter S.、Inagaki Y.、Ninimiya Y. 和 Ranirez F.：“人类 α(XI) 胶原蛋白基因启动子的结构和功能分析。”生物化学.270。418-424（1995）

Oohashi T., Sugimoto M., Mattei M.-G., and Ninomiya Y.: "Identification of a new collagen IV chain, alpha6 (IV), by cDNA isolation and assignment of the gene to chromosome Xq22, which is the same locus for COL4A5." J Biol Chem. 269. 7520-7526 (1994)

Oohashi T.、Sugimoto M.、Mattei M.-G. 和 Ninomiya Y.：“通过 cDNA 分离和将基因分配到染色体 Xq22 来鉴定新的胶原蛋白 IV 链 alpha6 (IV)，这与染色体 Xq22 相同