Cathepsins D, B, and L in transformed human breast epithelial cells.
To investigate the regulation of lysosomal enzymes during carcinogenesis, we measured cathepsins (Cats) D, B, and L in MCF-10F, which is a human breast epithelial cell line, and cells evolved after treatment with carcinogen and transfected with c-Ha-ras oncogene. The clones used in this study, MCF-10FTras, D3, D3-1, and D3-1Tras, expressed no estrogen receptors and gradually increased invasive potential, while oncogene-transfected lines were also tumorigenic in SCID mice [16, 19]. Cats D, B, and L were determined in the cells and in cell media using enzyme-linked immunosorbent assay (ELISA), specific enzyme activity measurements, and immunocytochemistry. The major intra- and extracellular lysosomal proteinase in these cells was Cat D (30-180 pm/mg), followed by Cat B (2-10 pm/mg) and Cat L (1-5 pm/mg). An inverse relationship between intracellular Cat D levels and invasive potential of carcinogen-treated and c-Ha-ras oncogene-transfected cell lines was observed. No significant changes in extracellular concentration of Cat D precursor in this series of cell lines was observed. Intracellular levels of Cats B and L were unchanged or slightly lower in carcinogen-treated D3 and D3-1 cells, as well as in MCF-10FTras. On the other hand, in D3-1Tras cell line, evolving from c-Ha-ras transfected D3-1 line, 3.5 fold and 4.4 fold increases in Cat B and Cat L, respectively, but a 2 fold decrease in Cat D, were observed compared to the parental cell line. Immunocytochemical staining showed a granular, polarized perinuclear and cytoplasmic staining of cathepsins in all cell lines. Cysteine proteinases stained more frequently and more intensely in D3-1Tras compared to other lines, confirming the immunochemical assays. We hypothesize that several molecular events, caused by a carcinogen and an oncogene such as c-Ha-ras, are needed to increase Cat B and Cat L, but not Cat D, expression. Therefore, the cysteine and aspartic lysosomal proteinases are differentially expressed in the breast cell lines with more invasive phenotype.
Lah
T T
TT
Department of Pathology and Laboratory Medicine, Albert Einstein Medical Center, Philadelphia, Pennsylvania, USA.
Calaf
G
G
Kalman
E
E
Shinde
B G
BG
Somers
R
R
Estrada
S
S
Salero
E
E
Russo
J
J
Daskal
I
I
D001940:Breast / Q000166:cytology / Q000378:metabolism*; D002403:Cathepsins / Q000096:biosynthesis*; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic / Q000378:metabolism*; D002999:Clone Cells; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D010641:Phenotype; D014162:Transfection / Q000235:genetics
D002403:Cathepsins