2-Methoxyestradiol, an estradiol metabolite, inhibits neointima formation and smooth muscle cell growth via double blockade of the cell cycle.
2-Methoxyestradiol (2-ME), an endogenous metabolite of estradiol with no affinity for estrogen receptors, is a potent anticarcinogenic agent (in phase II clinical trials) and mediates the inhibitory effects of estradiol on smooth muscle cell (SMC) growth. Here we studied the intracellular mechanisms by which 2-ME inhibits SMC growth and whether 2-ME prevents injury-induced neointima formation. 2-ME concentrations that inhibit proliferation of cycling human aortic SMCs by >or=50% blocked cell-cycle progression in G(0)/G(1) and in G(2)/M phase, as determined by flow cytometry. Consistent with the cell-cycle effects, at a molecular level (Western blots), 2-ME inhibited cyclin D(1) and cyclin B(1) expression; cyclin-dependent kinase (cdk)-1 and cdk-2 activity; and retinoblastoma protein (pRb), extracellular signal-regulated kinase (ERK) 1/2, and Akt phosphorylation. 2-ME also upregulated the Cdk inhibitor p27 and interfered with tubulin polymerization. Moreover, 2-ME augmented COX-2 expression, suggesting that it may also inhibit SMC growth via prostaglandin formation. In rats, treatment with 2-ME abrogated injury-induced neointima formation; decreased proliferating SMCs; downregulated expression of proliferating-cell nuclear antigen (PCNA), c-myc, cyclin D(1), cyclin B(1), phosphorylated Akt, phosphorylated ERK1/2, p21, and pRb; inhibited cdk-1 and cdk-4 activity; and upregulated expression of cyclooxygenase (COX)-2 and p27. Caspase-3 cleavage assay and fluorescence-activated cell-sorting (FACS) analysis showed no evidence of apoptosis in 2-ME-treated SMCs, and TUNEL staining in carotid segments showed no evidence of 2-ME-induced apoptosis in vivo. The antimitotic effects of 2-ME on SMCs are mediated by the inhibition of key cell-cycle regulatory proteins and effects on tubulin polymerization and COX-2 upregulation. These effects of 2-ME most likely contribute to the antivasoocclusive actions of this endogenous compound.
Barchiesi
Federica
F
Department of Obstetrics and Gynecology, Clinic for Endocrinology, University Hospital Zurich, Switzerland.
Jackson
Edwin K
EK
Fingerle
Juergen
J
Gillespie
Delbert G
DG
Odermatt
Bernhard
B
Dubey
Raghvendra K
RK
D000077584:2-Methoxyestradiol; D000818:Animals; D001011:Aorta; D002453:Cell Cycle / Q000187:drug effects*; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors; D049109:Cell Proliferation / Q000187:drug effects*; D051546:Cyclooxygenase 2 / Q000235:genetics; D004958:Estradiol / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use; D006801:Humans; D007399:Interphase / Q000187:drug effects; D032389:Myocytes, Smooth Muscle / Q000166:cytology*; D051381:Rats; D014404:Tubulin / Q000378:metabolism; D017539:Tunica Intima / Q000187:drug effects* / Q000254:growth & development
D018797:Cell Cycle Proteins; D014404:Tubulin; D004958:Estradiol; D000077584:2-Methoxyestradiol; D051546:Cyclooxygenase 2; C489691:Ptgs2 protein, rat