Blocking of JB6 Cell Transformation by Tanshinone IIA: Epigenetic Reactivation of Nrf2 Antioxidative Stress Pathway

Abstract

Abstract. Increasing numbers of natural products have been found to possess anticancer effects. Nuclear factor erythroid-2-related factor-2 (Nrf2) is a master regulator of the antioxidative stress response, and our previous studies found that epigenetic modification of the Nrf2 gene appears to be a critical mechanism. Salvia miltiorrhiza, a Chinese herbal medicine widely used in Asian countries, has been shown to possess anticancer and antioxidant effects. Tanshinone IIA (TIIA), an active component in S. miltiorrhiza, has been reported to activate Nrf2 pathway. The objective of this study was to investigate the epigenetic regulation of Nrf2 by TIIA in mouse skin epidermal JB6 cells and the functional consequences for cell transformation. TIIA was found to induce antioxidant response element-luciferase and upregulate the mRNA and protein levels of Nrf2 and Nrf2 downstream target genes HO-1 and NQO-1. TIIA decreased the colony formation of JB6 cells by approximately 80%. TIIA decreased the protein levels of DNMT1, DNMT3a, DNMT3b, and HDAC3 and inhibited the enzymatic activity of HDACs. Bisulfite genomic sequencing indicated that TIIA demethylated the first five CpGs in the promoter region of the Nrf2 gene. Chromatin immunoprecipitation assays showed that TIIA treatment increased the recruitment of RNA polymerase II at Nrf2 transcription start site but had limited effects on enrichment of Ac-H3 in Nrf2 promoter. Taken together, our results show that TIIA activates the Nrf2 signaling pathway and induces epigenetic demethylation of the CpGs of Nrf2. The epigenetic reactivation of the Nrf2 signaling pathway by TIIA could potentially contribute to the attenuation of JB6 cellular transformation and anticancer effects.

Curcumin inhibits anchorage-independent growth of HT29 human colon cancer cells by targeting epigenetic restoration of the tumor suppressor gene DLEC1

“…The colony-formation assay was performed as described previously with some modifications [34, 35]. The HT29, sh-Mock, and sh-DLEC1 cells (8 × 10 3 / well) were transferred to 1 mL of BME containing 0.33% agar over 3 mL of BME containing 0.5% agar with 10% FBS in 6-well plates.…”

Keap1–Nrf2 regulated redox signaling in utero: Priming of disease susceptibility in offspring

“…MGO ↑ ER stress , with Tm and Tg mimicking MGO to ↑ demethylation enzyme Tet1, Keap1 hypomethylation and Keap1 expression [128] Myocardial biopsies from non-diabetic (NDM) and type-2 diabetic (DM-T2) cardiomyopathy patients ↓ Keap1 promoter methylation, ↑ Keap1 expression and ↓Nrf2-dependent gene expression, [129] Developmental toxicity Mouse embryonic stem cells exposed to embryotoxicant monobutyl phthalate (MBP) Concomittant ↑ in methylation (MAT2A, AHCY) and Nrf2 related (GSTA4, PARK7) gene expression [151] Cancer 10 colorectal cancer cell lines or human tissue specimens ↑ Keap1 methylation in CRC cell lines or biopsies [152] Cancer TRAMP C1 tumor cells and TRAMP C3 non-tumour cells Nrf2 expression ↓ correlates with ↑ Nrf2 promoter methylation [153] Cancer TRAMP-C1 cells Curcumin (2.5µM) ↓ Nrf2 promoter methylation, ↑ NQO1 expression [154] Cancer JB6 cells Tanshinone IIA (~10µM) ↓ Nrf2 promoter methylation, ↑Nrf2-dependent gene expression [155] Cancer TRAMP-C1 cells 3,3"-diindolylmethane (DIM, 5µM) ↓ Nrf2 promoter methylation …”

Current Perspectives on Epigenetic Modifications by Dietary Chemopreventive and Herbal Phytochemicals

“…Another Chinese herb with great promise in altering epigenetic mechanisms is Salvia miltiorrhiza, also known as Danshen. We found that tanshinone IIA, one of the main active components from Danshen, blocks TPA (12-O-tetradecanoylphorbol-13-acetate)-mediated JB6 transformation through epigenetic regulation of the Nrf2 signaling pathway [111]. Treatment with tanshinone IIA reduced the methylation of the Nrf2 promoter, elevated the expression of Nrf2 and downstream targets, suppressed the protein levels of DNMT1, DNMT3a, DNMT3b, and HDAC3, and inhibited HDAC activity [111].…”

“…We found that tanshinone IIA, one of the main active components from Danshen, blocks TPA (12-O-tetradecanoylphorbol-13-acetate)-mediated JB6 transformation through epigenetic regulation of the Nrf2 signaling pathway [111]. Treatment with tanshinone IIA reduced the methylation of the Nrf2 promoter, elevated the expression of Nrf2 and downstream targets, suppressed the protein levels of DNMT1, DNMT3a, DNMT3b, and HDAC3, and inhibited HDAC activity [111]. Another study showed that tanshinone IIA decreases inflammatory responses in LPS-induced macrophages and inhibits the proliferation of inflammation-stimulated colon cancer cells by inhibiting the over-expressed miR-155 in macrophages [112].…”