Rui Yang

Macrophages secrete a variety of pro-inflammatory cytokines in response to pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) but abnormal release of cytokines unfortunately promotes cytokine storms. Dimethyl fumarate (DMF), an FDA-approved drug for multiple sclerosis (MS) treatment, has been found as an effective therapeutic agent for resolution. In this study, the anti-inflammatory effect of DMF was found to correlate to selenoprotein thioredoxin reductase 1 (TXNRD1). DMF irreversibly modified the Sec⁴⁹⁸ residue and C-terminal catalytic cysteine residues of TXNRD1 in a time- and dose-dependent manner. In LPS-stimulated RAW 264.7 cells, cellular TXNRD activity was increased through up-regulation of the protein level and DMF inhibited TXNRD activity and the nitric oxide (NO) production of RAW 264.7 cells. Meanwhile, the inhibition of TXNRD1 by DMF would contribute to the redox regulation of inflammation and promote the nuclear factor erythroid 2-related factor 2 (NRF2) activation. Notably, inhibition of cellular TXNRD1 by auranofin or TRi-1 showed anti-inflammatory effect in RAW 264.7 cells. This finding demonstrated that targeting TXNRD1 is a potential mechanism of using immunometabolites for dousing inflammation in response to pathogens and highlights the potential of TXNRD1 inhibitors in immune regulation.

Piperlongumine, a natural alkaloid substance extracted from the fruit of the long pepper (Piper longum Linn.), is known to inhibit the cytosolic thioredoxin reductase (TXNRD1 or TrxR1) and selectively kill cancer cells. However, the details and mechanism of the inhibition by piperlongumine against TXNRD1 remain unclear. In this study, based on the classical DTNB reducing assay, irreversible inhibition of recombinant TXNRD1 by piperlongumine was found and showed an apparent k_inact value of 0.206 × 10⁻³ µM⁻¹ min⁻¹. Meanwhile, compared with the wild-type TXNRD1 (-GCUG), the UGA-truncated form (-GC) of TXNRD1 was resistant to piperlongumine, suggesting the preferential target of piperlongumine is the selenol (-SeH) at the C-terminal redox motif of the enzyme. Interestingly, the high concentration of piperlongumine-inhibited TXNRD1 showed that its Sec-dependent activity is decayed but its intrinsic NADPH oxidase activity is retained. Furthermore, piperlongumine did not induce ferroptosis in HCT116 cells at 10 µM, whereas significantly promoted erastin-induced lipid oxidation, which could be alleviated by supplying glutathione (GSH) or N-acetyl L-cysteine (NAC). However, restricting GSH synthesis by inhibiting glutaminase (GLS) using the small molecule inhibitor CB-839 only slightly enhanced erastin-induced cell death. Taken together, this study elucidates the molecular mechanism of the antitumor capacity of piperlongumine by targeting TXNRD1 and reveals the potential possibility of inhibiting TXNRD1 to strengthen cancer cells’ ferroptosis.

Food colorants are widely used by humans in food production and preparation; however, their potential toxicity requires an in-depth analysis. In this study, five out of 15 commercial food colorants, namely, lutein, betanin, caramel, crocin and chlorophyll, significantly inhibited wild type selenoprotein thioredoxin reductase 1 (TrxR1, TXNRD1) in vitro. The hyperactive Sec⁴⁹⁸ residue of TrxR1 was targeted by those five colorants, which was confirmed by the site-directed mutagenesis of TrxR1. Furthermore, two colorants, chlorophyll and betanin, triggered the oligomerization of TrxR1. A chlorophyll-derived compound, chlorophyllin, irreversibly inhibited the 5,5′-dithiobis-2-nitrobenzoic acid (DTNB) reducing activity of TrxR1 with K_inact = 6.96 × 10⁻³ ± 0.49 × 10⁻³ µM⁻¹ min⁻¹. Moreover, chlorophyllin reduced the cellular TrxR activity, leading to reactive oxygen species (ROS) accumulation and, subsequently, promoting cancer cell death. In conclusion, this study might contribute to understand the food safety of commercial colorants and provide chemotherapeutic compounds by targeting TrxR1.