ヒト発癌過程でのSmad4発現減少を成因とするTGF-β不応性獲得機構の解析

人类癌变过程中Smad4表达降低导致TGF-β难治性获得的机制分析

• 批准号: 12213012
• 负责人: 萩原 弘一
• 金额: $ 2.69万
• 依托单位: Tohoku University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research on Priority Areas (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12213012/
• 关键词: TGF-β; Smad4; 増殖抑制; TGF-βII型受容体; 変異遺伝子; プロモーター; 容量反応曲線; 細胞株

我々は,遺伝子変異と,TGF-βの容量反応曲線を20種類の細胞株において検討した.TGE-βに対して全く反応の無い細胞株と,TGF-βII型受容体またはSmad4に遺伝子変異が認められた細胞株は完全に一致した.他の細胞株は,(1).TGF-βにより増殖を受ける細胞株,(2)TGF-βにより増殖抑制を受ける細胞株の2種に分類できた.(2)TGF-βにより増殖抑制を受ける細胞株では,全ての細胞株において,高濃度(10ng/ml)のTGF-βによっても80%以下の増殖抑制のみが認められた.次に,癌細胞と正常細胞のTGF-βII型受容体,Smad2,3,4,6,7の遺伝子発現をmRNAレベルで比較・検討した.その結果,Smad4に遺伝子変異のない21癌細胞株すべてにおいて,Smad4の発現が正常組織の1/20以下に低下していることを見いだした.Smad4の発現低下は,TGF-βII型受容体が変異を起こしている細胞株でも認められた.さらに,このSmad4の発現低下を引き起こす機構を調べるため,Smad4プロモーターをクローニングし,メチル化によるプロモーターの不活化が存在するか検討した.いずれの細胞株においてもプロモーターのメチル化は認められなかった.western blottingにおいて,一つの癌細胞株からは再現性良く常に同量の′Smad4蛋白が抽出されたが,正常組織からは,再現性が得られなかった.本研究において得られた結果は以下のようである.1.TGF-βII型受容体に変異のある細胞株においては,Smad4の発現減少により,TGF-βシグナル伝達系に内在するligand independentの増殖抑制からのescapeが起こっている.2.Smad4に変異のある細胞株では,TGF-βシグナル伝達系に内在するligand independentの増殖抑制はもともと存在せず,癌細胞はTGF-βシグナル伝達系から完全に開放されている.3.TGF-βII型受容体およびSmad4の両者に異常の無い細胞株においては,Smad4の発現減少により,TGF-βシグナル伝達系に内在するligand independentの増殖抑制からのescapeが起こっている.4.Smad4の発現低下は,プロモーターのメチル化が原因ではない.

We do not know that TGF- β capacity reversal curve is similar to that of TGF- β cell lines. TGE-β capacitor cell lines are completely identical to each other. TGE-β receptor cell lines are completely identical to those of type II acceptors. The colonization of TGF- β-colonization was affected by other cell lines, (1). The colonization inhibition of TGF- β-colonization was subdivided into two subtypes. (2) the colonization inhibition of TGF- β-colonization was affected by cell strain, whole cell strain, and high density (10ng/ml). In the second stage, the normal cells of cancer cells showed TGF- β type II receptor, and Smad2,3,4,6,7 cells showed that mRNA cells were significantly higher than those of normal cells. The results showed that the normal tissue of Smad4 was lower than 20 in normal tissue. Smad4 showed a low level, and TGF- β-type II acceptor showed that the cell line was negative. The results showed that the tumor cell line of cancer cell line was negative, and the normal tissue was lower than 20 in normal tissue. Smad4 showed low level. As a result of the increase in the number of people who are suffering from the disease, the low level of the Smad4 has led to the failure of the machine. The Smad4 has caused a change in the number of people who are not active. The cell line was isolated from the normal tissue, normal tissue and normal tissue. Western blotting. The results of this study showed that the following results were successful. 1. Smad4-β type II recipient cell lines were infected, while those of TGF-β type II recipient cells were reduced, while those of β-receptor cells were found to be intrinsically linked to ligand independent colonization inhibition and escape colonization inhibition. 2.Smad4 immunoglobulin In the TGF- β-phenotypic family, there is an innate cycle of ligand independent colonization inhibition, while the cancer cell line TGF- β has a complete open cycle. 3.TGF-β type II recipients with abortive Smad4 often fail to lose their cell lines in vitro, and Smad4 cells show a low level of DNA loss. TGF- β is caused by ligand independent, colony inhibition, colonization, escape, and so on. 4. Smad4 is low, and the cause is low.