|Budget Amount *help
¥1,600,000 (Direct Cost : ¥1,600,000)
Fiscal Year 1995 : ¥700,000 (Direct Cost : ¥700,000)
Fiscal Year 1994 : ¥900,000 (Direct Cost : ¥900,000)
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.