Reverse‐zymographic analysis of protease nexin‐II/amyloid β protein precursor of human carcinoma cell lines, with special reference to the grade of differentiation and metastatic phenotype

Trypsin inhibitors in serum‐free conditioned media (SFCM) of various human carcinoma cell lines were analyzed by reverse zymography. Most of the cells secreted high‐molecular‐weight trypsin inhibitors (HMTI) larger than 100 kDa. The cell lines of colorectal carcinoma origin had a tendency to secrete HMTI whose molecular weight was a little higher than that of the other cell lines. Analysis of SFCM of subclones with different histological differentiation and metastatic/invasive potentials derived from a single pancreatic carcinoma cell line SUIT‐2 showed that the HMTI activity in SFCM was correlated to the degree of histological differentiation in vivo and tended to be inversely correlated to their metastatic/invasive capabilities. Immunoblotting analysis revealed that these HMTI were protease nexin‐ll/amyloid β protein precursors (PN‐II/APP). Semi‐quantificative reverse‐transcriptase/polymerase‐chain reaction study for PN‐II/APP mRNAs suggested that the differences in PN‐II/APP activities in SFCM between the subclones might be post‐transcriptional or post‐secretional events. In addition, SFCM of a highly metastatic subclone contained 43‐kDa protein which reacted to anti‐APP monoclonal antibody (MAb) suggesting that the subclone may have APP‐degrading activity. © 1995 Wiley‐Liss, Inc.

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