Linked expression of Ah receptor, ARNT, CYP1A1, and CYP1B1 in rat mammary epithelia, in vitro, is each substantially elevated by specific extracellular matrix interactions that precede branching morphogenesis.
Cytochrome P4501B1 (CYP1B1), the major constitutively expressed CYP in the rat mammary gland, is induced by Ah-receptor (AhR) ligands, while CYP1A1 is predominantly expressed only after induction. These CYPs contribute to carcinogenic activation of polycyclic aromatic hydrocarbons (PAHs). AhR, ARNT, and CYP1B1 were only weakly expressed, even after 2,3,7,8-tetrachlorodibenzo-p-dioxin induction, when rat mammary epithelial cells (RMEC) were cultured on plastic. RMEC cultured on the extracellular matrix (ECM), Matrigel, or on a floating gel of collagen I demonstrated branching morphogenesis and substantially increased basal CYP1B1 and induced CYP1A1 expression, in parallel with large increases in AhR and ARNT expression. Branching was more pronounced in the Wistar Kyoto than in the Wistar Furth rat strain. Although EGF enhanced branching, neither strain nor growth factor treatment substantially impacted CYP expression. Increased AhR and ARNT expression is observed within 24 h of dispersal on Matrigel, substantially prior to branch formation. Culture on thin layers of collagen I, collagen IV, and laminin, respectively, failed to reproduce the branching morphogenesis or increases in AhR, ARNT, or CYP expression. However, adherent, gelled collagen I recapitulated the increased protein expression, without supporting branching. This increased protein expression was closely paralleled by enhanced expression of beta-catenin and E-cadherin, components of cell-cell adhesion complexes. A synthetic peptide that selectively antagonizes integrin-ECM interactions reduced branch formation, without diminishing AhR, ARNT, and CYP expression. These data demonstrate that early ECM surface adhesion interactions mediate AhR and ARNT expression, which enhances CYP expression, independent of branching morphogenesis.
Department of Pharmacology and the Environmental Toxicology Center, University of Wisconsin Medical School, 1300 University Avenue, Madison, WI 53706, USA.
Michele Campaigne
MC
Larsen
Paul B
PB
Brake
Richard S
RS
Pollenz
Colin R
CR
Jefcoate
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000096:biosynthesis*; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis*; D065633:Cytochrome P-450 CYP1B1; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D004338:Drug Combinations; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005260:Female; D007797:Laminin / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D009024:Morphogenesis / Q000502:physiology*; D011509:Proteoglycans / Q000378:metabolism; D051381:Rats; D011920:Rats, Inbred WF; D011921:Rats, Inbred WKY; D018336:Receptors, Aryl Hydrocarbon / Q000096:biosynthesis*; D013045:Species Specificity; D014157:Transcription Factors / Q000096:biosynthesis*
C497398:ARNT protein, rat; D004268:DNA-Binding Proteins; D004338:Drug Combinations; D016326:Extracellular Matrix Proteins; D007797:Laminin; D011509:Proteoglycans; D018336:Receptors, Aryl Hydrocarbon; D014157:Transcription Factors; C059485:matrigel; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D003094:Collagen; D001189:Aryl Hydrocarbon Hydroxylases; C585499:Cyp1b1 protein, rat; D019363:Cytochrome P-450 CYP1A1; D065633:Cytochrome P-450 CYP1B1