Epigenetic modifications of conjugate linoleic acid (CLA) on tumor suppressor gene, protein tyrosine phosphatase γ (PTPγ), in MCF-7 cells

Abstract

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Conjugated linoleic acid (CLA) are naturally occurring compounds found in ruminant products such as cheese and beef and has been shown to possess anti-cancer activities in in vivo and in vitro. It is widely accepted that epigenetic events, DNA methylation and histone modification affect the status of gene expression. Epigenetic aberrations plays a role in tumorigenesis and are involved in the silencing of tumor suppressor genes. Protein tyrosine phosphatase γ (PTPγ) has been implicated as an estrogen (E₂)-regulated candidate tumor suppressor gene in breast, ovarian and lung tumors. Our lab has previously demonstrated that CLA up-regulated PTPγ in human breast cells. This study is to examine whether CLA-mediated up-regulation of PTPγ expression is regulated by epigenetic modification in MCF-7 cells.

One of the most potent CLA isomer, t10, c12-CLA (10-40 μM), was used to examine CLA’s anti-proliferative activities by MTS and soft agar assays. Cell proliferation and colony formation were suppressed by 3 and 10 days of t10,c12-CLA (10-40 μM) treatment by 20-30% and 35-50%, correspondingly. t10, c12-CLA (10-40 μM, 3 days) significantly decreased DNA methyltransferase (DNMT) and histone deacetylase (HDAC) protein expression by 20-30% and 40-50%, respectively. Furthermore, the combinative effects of t10,c12-CLA (10-40 μM), 5-Azacytidine (5-Aza-CR, 1-10 μM, DNMT inhibitor), and Trichostatin A (TSA, 0.1-1 μM, HDAC inhibitor) on PTPγ protein expression were examined by western blot analysis. Our results showed that PTPγ protein expression was up-regulated by t10, c12-CLA (1.5-1.8 fold), 5-Aza-CR (1.2-1.5 fold) and TSA (1.6-2 fold). The combination of t10, c12-CLA (40 μM) and 5-Aza-CR (1-10 μM), or t10,c12-CLA (40 μM) and TSA (0.1-1 μM) further increased PTPγ protein expression by 2.5-3 or 2-2.5 fold, respectively.

Our experimental data suggest that epigenetic modification of PTPγ expression induced by CLA alone or synergistically enhanced by the combined treatment of 5-Aza-CR and TSA are practical important and clinical relevance. It is noteworthy that a natural food component produced from the fermentation of rumen bacterium in food-producing ruminants such as beef and dairy cattle has health significance to the consumers at large and also for chemoprevention purposes. The beneficial effects of low level and long-term daily consumption of CLA-enriched beef and dairy products may serve as potent natural demethylation compounds. Further basic research targeting the elucidation of regulatory mechanisms of epigenetic modification of chemopreventive food component, such as CLA is on-going in our lab. (Supported by NIH grants CA95915, ES0521-01 and IAMS Research Fund).