Fibronectin fibrils and growth factors stimulate anchorage-independent growth of a murine mammary carcinoma.
Stromal cells are important regulators of mammary carcinoma growth and metastasis. We have previously shown that a 3T3-L1 adipocyte cell line secretes hepatocyte growth factor (HGF), which stimulates proliferation of a murine mammary carcinoma (SP1) in monolayer cultures (DNA Cell Biol. 13, 1189-1897, 1994). We now examine the role of growth factors and the extracellular matrix protein fibronectin in stimulation of anchorage-independent growth of SP1 cells. Purified transforming growth factor-beta (TGF-beta) stimulated significant colony growth in soft agar cultures, whereas HGF had a lesser effect. Analysis by confocal microscopy revealed that carcinoma cell colonies contained extracellular microfibrils composed of fibronectin. Partial depletion of fibronectin from 7% FBS/agar cultures reduced the number of colonies; colony growth could be recovered by adding back exogenous fibronectin. Addition of the 70-kDa N-terminal fragment of fibronectin, which inhibits fibronectin fibril formation, reduced growth of SP1 cell colonies, but an 85-kDa fragment containing the cell binding domain did not inhibit colony growth. These findings indicate that deposition of extracellular fibronectin fibrils is necessary, but not sufficient, for anchorage-independent growth of SP1 mammary carcinoma cells; growth factors are also required. SP1 cells had less fibronectin mRNA and secreted less fibronectin protein under anchorage-independent conditions than under anchorage-dependent conditions, as determined by Northern blotting and immunoprecipitation analysis. Thus, both growth factors (HGF and TGF-beta) and fibronectin may be important regulators of paracrine stimulation by stromal cells of anchorage-independent growth of mammary carcinoma cells.
Saulnier
R
R
Cancer Research Laboratory, Queen's University, Kingston, Ontario, Canada.
Bhardwaj
B
B
Klassen
J
J
Leopold
D
D
Rahimi
N
N
Tremblay
E
E
Mosher
D
D
Elliott
B
B
D016475:3T3 Cells / Q000378:metabolism; D000230:Adenocarcinoma; D017667:Adipocytes / Q000378:metabolism; D000818:Animals; D015152:Blotting, Northern; D002448:Cell Adhesion / Q000502:physiology; D002455:Cell Division / Q000187:drug effects; D005109:Extracellular Matrix / Q000737:chemistry / Q000502:physiology; D005260:Female; D005353:Fibronectins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D017228:Hepatocyte Growth Factor / Q000378:metabolism / Q000502:physiology*; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism*; D051379:Mice; D008808:Mice, Inbred CBA; D014411:Neoplastic Stem Cells; D012333:RNA, Messenger / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D005353:Fibronectins; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D017228:Hepatocyte Growth Factor