Cell experiment:

LNCaP cells are cultured at 37℃ in 5% CO2 in RPMI supplemented with 10% FBS and antibiotics. Cell viability is determined by an MTT assay. Briefly, LNCaP cells (1×105 cells/mL) are plated onto 24 well plates and incubated overnight in serum and serum-free RPMI media. The cells are treated with the indicated concentrations of Mangiferin (100, 200, and 400 μM) for 1 h and then stimulated with TNF-α (20 ng/mL) for 24 h. Then, the cells are incubated with a solution of 0.5 mg/mL MTT and incubation for 45 min at 37℃ and 5% CO2. Supernatant is removed and the formation of formazan is observed by monitoring the signal at 540 nm using a microplate reader[2].

Mangiferin is a Nrf2 activator. Mangiferin suppresses nuclear translocation of the NF-κB subunits p65 and p50.

Mangiferin is glucosylxanthone extracted from plants of the Anacardiaceae and Gentianaceae families. Mangiferin (50 μM) significantly increases Nrf2 protein accumulation in HL-60 cells, particularly in the nucleus. Mangiferin also enhances the binding of Nrf2 to an antioxidant response element (ARE), significantly up-regulates NAD(P)H:quinone oxidoreductase 1 (NQO1) expression and reduces intracellular ROS in HL60 cells. Mangiferin alone dose-dependently inhibits the proliferation of HL-60 cells. In mononuclear cells (MNCs), Mangiferin significantly relieves oxidative stress, but attenuates etoposide-induced cytotoxicity[1]. Mangiferin is a natural phytopolyphenol that exhibits various pharmacological properties. Mangiferin is present in several plant species such as Mangifera indica, Iris unguicularis, and Anemarrhena asphodeloides. Mangiferin downregulates TNF-α-induced MMP-9 mRNA and protein expression by suppressing NF-κB activity, consequently suppressing the invasion of LNCaP cells.To assess whether Mangiferin influences the viability of LNCaP cells, MTT assay is performed 24 h after treatment with different concentrations of Mangiferin in the presence or absence of TNF-α in serum and serum-free conditions. There are no cytotoxic evident in LNCaP cells treated with up to 400 μM of Mangiferin alone under both serum and serum-free conditions. Additionally, in the presence of TNF-α (20 ng/mL), Mangiferin does not affect cell viability. Mangiferin (400 μM) treatment significantly decreases the NF-κB luciferase activity in TNF-α-stimulated LNCaP cells. Pretreatment with Mangiferin (400 μM) for 1 h significantly decreases TNF-α-induced NF-κB activity. The RT-PCR shows that Mangiferin (400 μM) significantly suppresses the TNF-α-induced MMP-9 expression in LNCaP cells[2].

References:
[1]. Zhang BP, et al. Mangiferin activates Nrf2-antioxidant response element signaling without reducing the sensitivity to etoposide of human myeloid leukemia cells in vitro. Acta Pharmacol Sin. 2014 Feb;35(2):257-66.
[2]. Dilshara MG, et al. Mangiferin inhibits tumor necrosis factor-α-induced matrix metalloproteinase-9 expression and cellular invasion by suppressing nuclear factor-κB activity. BMB Rep. 2015 Oct;48(10):559-64.