Induction by 7,12-dimethylbenz(a)anthracene of molecular and biochemical alterations in transformed human mammary epithelial stem cells, and protection by N-acetylcysteine.
Several lines of evidence suggest that stem cells are major targets for carcinogens. A normal human breast epithelial cell type was previously shown to possess stem cell characteristics. Further cell lines were derived following sequential transfection with SV40 large T-antigen (immortal, non-tumorigenic M13SV1 cells), exposure to X-rays (weakly tumorigenic M13SV1R2 cells), and ectopic expression of c-erbB2/neu (highly tumorigenic M13SV1R2N1 cells). We evaluated some characteristics of these cells and their susceptibility to the breast carcinogen 7,12-dimethylbenz(a)anthracene (DMBA). Compared to M13SV1 cells, the two untreated tumorigenic cell lines displayed higher levels of connexin 43 expression and NF-kappaB nuclear translocation, and a higher frequency of fhit loss. The baseline nuclear translocation of AP-1 and pCREB was particularly evident in M13SV1R2N1 cells and was further enhanced by DMBA treatment, indicating an interaction between c-erbB2/neu and DMBA-induced signalling. Treatment with DMBA did neither affect the baseline fhit loss nor p53 mutation, whereas it increased NF-kappaB nuclear translocation, the proportion of apoptotic cells, and the levels of connexin 43, common 4977-bp mitochondrial DNA deletion, and bulky adducts to nuclear DNA. DMBA-treated M13SV1 cells underwent significant oxidative DNA damage and exhibited the highest DNA adduct levels, while they had the lowest apoptotic rate. Co-treatment of cells with N-acetylcysteine (NAC) attenuated DMBA-induced toxicity and DNA alterations, particularly in M13SV1 cells. Thus, the immortal cell line derived from the normal human adult breast stem cell without further tumorigenic progression is the most susceptible both to DMBA-related alterations and to the protective effects of NAC.
De Flora
Silvio
S
Department of Health Sciences, Section of Hygiene and Preventive Medicine, University of Genoa, I-16132 Genoa, Italy. sdf@unige.it
Scarfì
Sonia
S
Izzotti
Alberto
A
D'Agostini
Francesco
F
Chang
Chia-Cheng
CC
Bagnasco
Maria
M
De Flora
Antonio
A
Trosko
James E
JE
D015127:9,10-Dimethyl-1,2-benzanthracene / Q000633:toxicity*; D000111:Acetylcysteine / Q000494:pharmacology*; D017766:Acid Anhydride Hydrolases / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D002273:Carcinogens / Q000633:toxicity*; D002461:Cell Line, Transformed; D002467:Cell Nucleus; D002478:Cells, Cultured; D018031:Connexin 43 / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology*; D006801:Humans; D042361:Mammary Glands, Human / Q000187:drug effects* / Q000378:metabolism; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D021381:Protein Transport; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism; D013234:Stem Cells / Q000187:drug effects* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D000099170:Bis(5'-adenosyl)triphosphatase
D015127:9,10-Dimethyl-1,2-benzanthracene; D000111:Acetylcysteine; D017766:Acid Anhydride Hydrolases; D002273:Carcinogens; D018031:Connexin 43; D016166:Free Radical Scavengers; D016328:NF-kappa B; D009363:Neoplasm Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D016159:Tumor Suppressor Protein p53; D000099170:Bis(5'-adenosyl)triphosphatase