Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine produced by monocytes/macrophage that plays a pathological role in rheumatoid arthritis (RA). In this study, we investigate the effect of thymoquinone (TQ), a phytochemical found in Nigella sativa, in regulating TNF-α-induced RA synovial fibroblast (RA-FLS) activation. Treatment with TQ (1–5 μM) had no marked effect on the viability of human RA-FLS. Pre-treatment of TQ inhibited TNF-α-induced interleukin-6 (IL-6) and IL-8 production and ICAM-1, VCAM-1, and cadherin-11 (Cad-11) expression in RA-FLS (p<0.01). Evaluation of the signaling events showed that TQ inhibited TNF-α-induced phospho-p38 and phospho-JNK expression, but had no inhibitory effect on NF-κB pathway, in RA-FLS (p<0.05; n=4). Interestingly, we observed that selective down-regulation of TNF-α-induced phospho-p38 and phospho-JNK activation by TQ is elicited through inhibition of apoptosis-regulated signaling kinase 1 (ASK1). Furthermore, TNF-α selectively induced phosphorylation of ASK1 at Thr845 residue in RA-FLS, which was inhibited by TQ pretreatment in a dose dependent manner (p<0.01). Pre-treatment of RA-FLS with ASK1 inhibitor (TC ASK10), blocked TNF-α induced expression of ICAM-1, VCAM-1, and Cad-11. Our results suggest that TNF-α-induced ASK1-p38/JNK pathway is an important mediator of cytokine synthesis and enhanced expression of adhesion molecule in RA-FLS and TQ, by selectively inhibiting this pathway, may have a potential therapeutic value in regulating tissue destruction observed in RA.

While microRNAs (miRNAs) regulate the vast majority of protein-encoding transcripts, little is known about how miRNAs themselves are degraded. We recently described Tudor-staphylococcal/micrococcal-like nuclease (TSN)-mediated miRNA decay (TumiD) as a cellular pathway in which the nuclease TSN promotes the decay of miRNAs that contain CA and/or UA dinucleotides. While TSN-mediated degradation of either protein-free or AGO2-loaded miRNAs does not require the ATP-dependent RNA helicase UPF1 in vitro, we report here that cellular TumiD requires UPF1. Results from experiments using AGO2-loaded miRNAs in duplex with target mRNAs indicate that UPF1 can dissociate miRNAs from their mRNA targets, making the miRNAs susceptible to TumiD. miR-seq (deep sequencing of miRNAs) data reveal that the degradation of ∼50% of candidate TumiD targets in T24 human urinary bladder cancer cells is augmented by UPF1. We illustrate the physiological relevance by demonstrating that UPF1-augmented TumiD promotes the invasion of T24 cells in part by degrading anti-invasive miRNAs so as to up-regulate the expression of proinvasive proteins.

Aberrant expression of mitochondrial proteins impairs cardiac function and causes heart disease. The mechanism of regulation of mitochondria encoded protein expression during cardiac disease, however, remains underexplored. Here, we show that multiple pathogenic cardiac stressors induce the expression of miR‐574 guide and passenger strands (miR‐574‐5p/3p) in both humans and mice. miR‐574 knockout mice exhibit severe cardiac disorder under different pathogenic cardiac stresses while miR‐574‐5p/3p mimics that are delivered systematically using nanoparticles reduce cardiac pathogenesis under disease insults. Transcriptomic analysis of miR‐574‐null hearts uncovers family with sequence similarity 210 member A (FAM210A) as a common target mRNA of miR‐574‐5p and miR‐574‐3p. The interactome capture analysis suggests that FAM210A interacts with mitochondrial translation elongation factor EF‐Tu. Manipulating miR‐574‐5p/3p or FAM210A expression changes the protein expression of mitochondrial‐encoded electron transport chain (ETC) genes but not nuclear‐encoded mitochondrial ETC genes in both human AC16 cardiomyocyte cells and miR‐574‐null murine hearts. Together, we discovered that miR‐574 regulates FAM210A expression and modulates mitochondrial‐encoded protein expression, which may influence cardiac remodeling in heart failure.

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In the face of increasing resistance to the existing antibiotics, oxazolidinones (exemplified by linezolid) have been developed as promising antibacterial agents, but may have other useful actions. In the present study, a series of 5‑(1H‑1,2,3‑triazoly) l‑methyl‑, 5‑acetamidomethyl‑morpholino and N‑substituted‑piperazino oxazolidinone derivatives were investigated to determine whether they are active against eukaryotic cells. An MTT assay, validated by cell counting, was used to assess the effect of nine oxazolidinone derivatives (concentrations 100 nM‑10 µM) on the proliferation of MCF7 human breast cancer cells. The three most active compounds were then tested on MDA231 breast cancer cells. Cytotoxicity of the selected derivatives was determined by assessing the extent of apoptosis by flow cytometry. The antimetastatic potential of these compounds was assessed on MDA231 cells using wound healing and agarose invasion assays. The 5‑triazolylmethyl piperazino‑oxazolidinone derivatives containing 4‑N‑(2‑chlorocinnamoyl), 4‑N‑(4‑nitrobenzoyl) and 4‑N‑methylsulfonyl moieties exhibited the most potent cytostatic activity against cancer, inhibiting proliferation by up to 70%, in the same order as their reported antibacterial activity against Staphylococcus aureus, but at higher concentrations. Unexpectedly, several derivatives stimulated proliferation at 100 nM, well below their antibacterial minimum inhibitory concentrations. Certain compounds also retarded the motility and invasion of MDA231 cells. Three of the tested derivatives had no effect on the eukaryotic cell lines, demonstrating their preferential activity against bacteria. Two compounds actually stimulated eukaryotic cell proliferation. The remaining three exhibited potent cytostatic activity against and cancer cells, displaying differences in response at low and high concentrations, which may suggest multiple targets on eukaryotic cells. These latter compounds may be useful as anticancer agents.