Transforming growth factor beta stimulates mammary adenocarcinoma cell invasion and metastatic potential.

Abstract

The experimental metastatic potential of 13762NF mammary adenocarcinoma clone MTLn3 was tested after pretreatment in serum-free medium containing transforming growth factor (TGF) 8,1 at 0-5000 pg/ml. Lung colonies were measured 2 weeks after inoculation in syngeneic F344 rats, and a bell-shaped dose-response curve with 2-to 3-fold increase in number of surface lung metastases was seen. Maximal enhancement occurred at the 50 pg/ml dose level. The effect was specific because addition of neutralizing anti-TGF-18 antibody blocked the stimulatory activity at all levels of TGF-13, pretreatment, but when antibody was given alone, neutralizing anti-TGF-f8 antibody had no effect on untreated cells. Increased metastatic potential appears to be from an increased propensity of cells to extravasate as tested in the membrane invasion culture system. MTLn3 cells penetrated reconstituted basement-membrane barriers 2-to 3.5-fold more than did untreated control cells, depending upon length of TGF-813 exposure. Increased invasive potential is apparently due, in part, to a 2-to 6-fold increase in type IV collagenolytic (gelatinolytic) and a 2.4-fold increase in heparanase activity.TGF-fi1 treatment of MTLn3 cells did not alter their growth rate or morphology in the presence of serum; however, growth was inhibited in serum-free medium. Likewise, adhesion to human umbilical vein endothelial cell monolayers or to immobilized reconstituted basement membrane or fibronectin matrices was unchanged. These results suggest that TGF-1, may modulate metastatic potential of mammary tumor cells by controlling their ability to break down and penetrate basementmembrane barriers.Transforming growth factor (TGF) 3 is a 25-kDa disulfidelinked polypeptide dimer that elicits numerous changes in cellular behavior, including differentiation of epithelial cells (for reviews, see refs. 1-4), modifying proliferation in a wide variety of cell types (1, 4-10), inhibition of angiogenesis (11,12), deposition of extracellular matrix components (11-15), alteration of basement-membrane-degrading enzyme production and/secretion (refs. 11, 16-19 MATERIALS AND METHODSCell Lines and Tissue Culture. 13762NF rat mammary adenocarcinoma clone MTLn3 cells were grown and maintained as described (28,30). Briefly, cells were grown in a-modified Eagle's minimal essential medium (a-MEM) supplemented with 5% fetal bovine serum (FBS) in 100-mm tissue culture dishes (Coming) without antibiotics. Cells were subcultured when the plates became 70-80%o confluent by using 0.25% trypsin solution in Ca2+-and Mg2+-free Dulbecco's phosphate-buffered saline. All cultures were routinely tested and found negative for Mycoplasma sp. contamination.Animals. Pathogen-and virus-free, 6-to 7-week old female Fischer 344/NHSd (F344) rats were obtained from Harlan Sprague-Dawley. Animals were maintained under the guidelines of Glaxo Research Laboratories and the National Institutes of Health. All protocols were approved by Glaxo's Institutional Animal Care and Use Committee. Rats were fed Pur...

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