Effect of a Protease Inhibitor on the Adhesion of Ehrlich Ascites Cells to Host Cells in vivo

Abstract

Summary.-Ehrlich ascites tumours (EAT) were grown in mice by injecting 1 x 106 cells intraperitoneally. In mice which received one or more injections of 30 mg soybean trypsin inhibitor (TI) i.p. during tumour growth, the number of recoverable tumour cells was significantly reduced by up to 92%. Also, the mean size of these cells was significantly smaller.When the rate of labelled thymidine incorporation in vitro was compared in TItreated and control cells, no significant differences were detected. However, when the population doubling time of EAT cells in vivo was calculated, it was apparent that recoverable TI-treated cells were dividing more rapidly than controls. Consequently, the reduced number of cells recovered from TI-treated mice did not result from a reduced growth rate.Viability, assessed by trypan blue dye exclusion and rate of labelled chromium release, was the same in TI-treated and control cell populations. Thus TI was nontoxic to EAT cells and the reduced number of cells from treated tumours was not therefore due to cytotoxicity.Scanning electron microscopy revealed that normal EAT cells did not adhere to internal host surfaces but that after TI treatment they adhered in lar e numbers to produce an appearance which resembled a confluent monolayer. This binding to host tissue accounted for the reduction in the number of cells recovered from TItreated animals. We propose that TI acts as a protease inhibitor to prevent intrinsic proteolytic enzyme activity at the tumour cell surface. This activity would normally destroy the binding sites required for adhesion to host tissue.

A trypsin-like neutral protease on Ehrlich ascites cell surfaces: its role in the activation of tumour-cell zymogen of collagenase

“…in mice exactly as described by Whur et al (1973). The cells were collected in isotonic saline after 8-10 days' incubation and the surrounding ascitic fluid was separated by centrifugation for 5 min at 300 g. The cells were washed x 6 by centrifugation in isotonic saline to remove all traces of ascitic fluid before use.…”

“…STUDIES ON EHRLICH ASCITES CELLS grown in mice (Whur et al, 1973) showed that when these cells were reinjected into fresh mice together with soybean trypsin inhibitor (SBTI) the tumour cells became deposited as a confluent monolayer on the hosts' internal abdominal surfaces. The authors suggested that the reason for this organized deposition was that the SBTI prevented proteolytic activity of an enzyme on the cell surface.…”

Endocytosis of the antiprotease aprotinin by Landschütz ascites carcinoma cells and its effects in vitro and in vivo

“…2 Effect8 of aprotinin on tumour growth G-rowth of the tumour, measured by the number of recoverable ascites cells within both saline and aprotinin-treated animals at various times after injection of 106 viable tumour cells is shown in Fig. 4 (Back and Leblanc, 1977;Kinjo et al, 1963;Verloes et al, 1978;Whur et al, 1973) (Hynes, 1976;Talmadge et al, 1974). In vtio however, such effects are t-likely to be less pronounced, due to more rapid catabolism of aprotinin and its widespread binding to ubiquitous sialyl moieties; factors which probably account for its relative inefficiency as a therapeutic agent in the tumour-bearing host, and which are likely to be exaggerated by rapid cell replication and copious ascitic fluid production.…”

“…Also, in view of reports that a natural protease inhibitor (soybean trypsin inhibitor) impairs growth of ascites tumours (Whur et al, 1973;Verloes et al, 1978) (Watson, 1958) and lead citrate (Reynolds, 1963) prior to examination in an AEI EM6B transmission electron microscope.…”

β-Naphthylamidase activity of the cell surface of Ehrlich ascites cells. Reversible control of enzyme activity by metal ions and thiols

“…in mice, as described by Whur et al (1973). The cells were collected in isotonic saline after 8-10 days' incubation and the surrounding ascitic fluid was separated by centrifugation for 5 min at 300 g. The cells were washed x 6 by centrifugation in isotonic saline before use.…”