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Proteinase secretion in serum-independent cell lines established from human cancer cell lines.

由人类癌细胞系建立的不依赖于血清的细胞系中的蛋白酶分泌。

基本信息

批准号：
06672188
负责人：
YASUMITSU Hidetaro
金额：
$ 1.02万
依托单位：
KIHARA INSITUTE FOR BIOLOGICAL RESEARCH/YOKOHAMA CITY UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
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

项目摘要

1.Establishment of serum-independent human cell lines :After selection by culturing human cancer cell lines in the synthetic medium without serum nor cytokines for at least 6 months, 31 serum-independent growing cell lines were established from human cancer cell lines such as stomach, colon and ovarian cell lines. During establishment under serum-free conditions, parental cell lines showed roughly four patterns of growth properties : 1) within 2 weeks, all cells died simultaneiously. 2) by 4 weeks, cell number gradually decreased and disappeared. 3) cells could grow only in mass culture. 4) even from a single cell, it could gradually grow.2.Expression of proteinase and their inhibitors in serum-dependent and serum-independent cell lines. : By the analyzes using zymography, reversezymography and Northern-blotting, stomach cancer cell line established from malignant tumor secreted trypsin-like enzyme. The enzyme was identified to be trypsinogen-1 by its amino acid sequencing. And the enzyme was more expressed in serum-independent cell line than its parental serum-dependent cell line.3.Regulation of trypsinogen-1 mRNA expression by serum and cytokines. : When the effect of cytokines and growth factors on the expression fo trypsinogen-1 gene in human stomach cancer cells were evaluated, serum and inflammatory cytokines were revealed to regulate the expression of trypsinogen-1 gene.mRNA expression of trypsinogen-1 gene in other cell lines. : After screening various human cell lines on the expression of trypsinogen-1 gene, several human cancer cell lines such as colon and stomach cancer cell lines were revealed to express mRNA of the gene. However, only two human stomach cancer cell lines were regulated by serum factors.

1.建立不依赖于血清的人类细胞系：在不含血清和细胞因子的合成培养基中培养人类癌细胞系至少6个月，经过筛选，从人类癌细胞系（例如胃、结肠和卵巢细胞系）中建立了31个不依赖于血清的生长细胞系。在无血清条件下建立期间，亲代细胞系大致表现出四种生长特性模式：1) 2周内，所有细胞同时死亡。 2）到4周时，细胞数量逐渐减少并消失。 3）细胞只能在大量培养中生长。 4)即使是单个细胞，也可以逐渐生长。2.血清依赖性和血清非依赖性细胞系中蛋白酶及其抑制剂的表达。：通过酶谱法、逆酶谱法和Northern印迹分析，从恶性肿瘤分泌的胰蛋白酶样酶建立的胃癌细胞系。通过氨基酸测序，该酶被鉴定为胰蛋白酶原-1。并且该酶在血清非依赖性细胞系中的表达量高于其亲代血清依赖性细胞系。3.血清和细胞因子对胰蛋白酶原1 mRNA表达的调节。：在评估细胞因子和生长因子对人胃癌细胞中胰蛋白酶原-1 基因表达的影响时，发现血清和炎症细胞因子调节胰蛋白酶原-1 基因的表达。其他细胞系中胰蛋白酶原-1 基因的 mRNA 表达。：在对各种人类细胞系的胰蛋白酶原-1基因的表达进行筛选后，发现一些人类癌细胞系，例如结肠癌细胞系和胃癌细胞系，表达该基因的mRNA。然而，只有两种人胃癌细胞系受到血清因子的调节。