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Analysis of Phenotypic Variation of Mast Cells by Using Messenger RNA Expression as a Marker

以信使 RNA 表达为标志物分析肥大细胞表型变异

基本信息

批准号：
06454193
负责人：
KITAMURA Yukihiko
金额：
$ 4.48万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06454193/
关键词：
mast cell protease in situ hybridization cytokine cytokine receptor transcription factor tryptase chymase in sith ハイブリダイゼーションマスト細胞プロテアーゼ in situ ハイブリダイゼーション寄生虫感染結合組織マスト細胞粘膜型マスト細胞

项目摘要

Basophilic granules of mase cells contain various mast cell-specific proteases. Many genes encoding mast cell-specific proteases have been cloned in mice. Mouse mast cells proteases (MMCP) 1 to 5 are chymases, and MMCP-6 and -7 are tryptases. We investigated the factors which influenced the expression of mast cell-specific proteases by using in situ hybridization. The pattern of MMCP expression was affected by mouse atrains, tissues in which mast cells differentiated and signals from cytokine receptors. In 1994, we obtained cultured mast cells (CMCs) by cultureing bone marrow cells of mice in the presecnce of T cell-derived cytokines. Total RNA was extracted from CMCs, and cDNAs of MMCP-2, MMCP-4, MMCP-5, MMCP-6 and mast cell carboxypeptidase were obtained. CMCs were centrifuged and embedded in paraffin, and in situ hybridiazation using digoxigenin-labeled RNA probes was carried out. Since the technical problems were resolved in 1994, we studid the change in mRNA expression patters of mast cell-specific proteases under various culture condition in 1995. We found the gain-of-function mutations of c-kit receptor tyrosine kinase, which is the receptor for stem cell factor, the most important growth factor of mast cells. We investifated the effect of signals through the c-kit receptor on the expression of mast cell-specific proteases. Introduction of the constitutively activated c-kit cDNA into the IC-2 mast cell line resulted in the enhanced expression of MMCP-6.

肥大细胞嗜碱性颗粒含有多种肥大细胞特异性蛋白酶。许多编码肥大细胞特异性蛋白酶的基因已在小鼠中被克隆。小鼠肥大细胞蛋白酶（MMCP）1至5是糜酶，并且MMCP-6和-7是胰蛋白酶。我们用原位杂交技术研究了影响肥大细胞特异性蛋白酶表达的因素。MMCP的表达模式受小鼠软骨、肥大细胞分化的组织和来自细胞因子受体的信号的影响。1994年，我们在T细胞源性细胞因子存在下，通过培养小鼠骨髓细胞获得了培养的肥大细胞（CMCs）。从CMC中提取总RNA，获得MMCP-2、MMCP-4、MMCP-5、MMCP-6和肥大细胞羧肽酶的cDNA。将CMC离心并包埋在石蜡中，并使用地高辛标记的RNA探针进行原位杂交。自1994年技术问题解决后，我们于1995年研究了肥大细胞特异性蛋白酶在不同培养条件下mRNA表达模式的变化。我们发现了c-kit受体酪氨酸激酶的功能获得性突变，c-kit受体酪氨酸激酶是干细胞因子的受体，而干细胞因子是肥大细胞最重要的生长因子。我们研究了通过c-kit受体的信号对肥大细胞特异性蛋白酶表达的影响。将组成型激活的c-kit cDNA导入IC-2肥大细胞系导致MMCP-6的表达增强。