p53-independent G1 cell cycle arrest of human colon carcinoma cells HT-29 by sulforaphane is associated with induction of p21CIP1 and inhibition of expression of cyclin D1.
Isothiocyanate sulforaphane (SFN) is a potent cancer chemopreventive agent. We investigated the mechanisms underlying the anti-proliferative effects of SFN in the human colon carcinoma cell line, HT-29. We demonstrate that SFN inhibits the growth of HT-29 cells in a dose- and time-dependent manner. Treatment of serum-stimulated HT-29 cells with SFN suppressed the re-initiation of cell cycle by inducing a G(1) phase cell cycle arrest. At high doses (>25 microM), SFN dramatically induces the expression of p21(CIP1) while significantly inhibits the expression of the G(1) phase cell cycle regulatory genes such as cyclin D1, cyclin A, and c-myc. This regulation can be detected at both the mRNA and protein levels as early as 4 h post-treatment of SFN at 50 microM. Additionally, SFN activates MAPKs pathways, including ERK, JNK and p38. Exposure of HT-29 cells with both SFN and an antioxidant, either NAC or GSH, completely blocked the SFN-mediated activation of these MAPK signaling cascades, regulation of cyclin D1and p21(CIP1) gene expression, and G(1)phase cell cycle arrest. This finding suggests that SFN-induced oxidative stress plays a role in these observed effects. Furthermore, the activation of the ERK and p38 pathways by SFN is involved in the upregulation of p21(CIP1) and cyclin D1, whereas the activation of the JNK pathway plays a contradictory role and may be partially involved in the downregulation of cyclin D1. Because cyclin D1 and p21(CIP1) play opposing roles in G(1) phase cell cycle progression regulation, blocking the activation of each MAPK pathway with specific MAPK inhibitors, is unable to rescue the SFN-induced G(1) phase cell cycle arrest in HT-29 cells.
Shen
Guoxiang
G
Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, 08854, USA.
Xu
Changjiang
C
Chen
Chi
C
Hebbar
Vidya
V
Kong
Ah-Ng Tony
AN
D000111:Acetylcysteine / Q000494:pharmacology; D016588:Anticarcinogenic Agents / Q000494:pharmacology; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D016193:G1 Phase / Q000187:drug effects* / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D005978:Glutathione / Q000494:pharmacology; D019073:HT29 Cells; D006801:Humans; D017879:Isothiocyanates; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013454:Sulfoxides; D013778:Tetrazolium Salts / Q000737:chemistry / Q000378:metabolism; D013861:Thiocyanates / Q000494:pharmacology*; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
D016588:Anticarcinogenic Agents; D050759:Cyclin-Dependent Kinase Inhibitor p21; D017879:Isothiocyanates; D013454:Sulfoxides; D013778:Tetrazolium Salts; D013861:Thiocyanates; D016159:Tumor Suppressor Protein p53; D019938:Cyclin D1; D020928:Mitogen-Activated Protein Kinases; C016766:sulforaphane; D005978:Glutathione; D000111:Acetylcysteine