RNA-seq Transcriptomic Analysis of 4-octyl itaconate Repressing Myogenic Differentiation
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ABSTRACT: The 4-octyl itaconate O1_is a type of cell-permeable itaconate derivative. Studies have shown that with an anti-fibrotic effect in systemic sclerosis, the OI also affects osteoclast differentiation. The aim of this study was to explore the molecular mechanisms underlying the effects of OI on myoblast differentiation by RNA-seq analysis.
Project description:Activated macrophages switch from oxidative phosphorylation to aerobic glycolysis, similar to the Warburg effect, presenting a potential therapeutic target in inflammatory disease. The endogenous metabolite itaconate has recently been reported to regulate macrophage function, but its precise mechanism is not fully understood. Here, we showed that 4-octyl itaconate (OI, a cell-permeable itaconate derivative) directly alkylated cysteine residue 22 on the glycolytic enzyme GAPDH and decreased its enzyme activity. Glycolytic flux analysis by U13C‐glucose tracing also provided evidence that OI generated a blockade of glycolytic flux at GAPDH. OI thereby downregulates aerobic glycolysis in activated macrophages, which is required for its anti-inflammatory effects. The anti-inflammatory effects of OI were replicated by heptelidic acid, 2-DG and reversed by increasing wild-type but not C22A mutant GAPDH expression. OI protected against lipopolysaccharide-induced lethality in vivo and significantly inhibited cytokine release. These findings showed that itaconate is a critical immunometabolite that exerts anti-inflammatory effects by targeting GAPDH to decrease aerobic glycolysis in macrophages.
Project description:<p>The Krebs cycle-derived metabolite itaconate, whose production is catalyzed by immune response gene 1 (IRG1), has excellent potential to link immunity and metabolism in activated macrophages through alkylation or competitive inhibition of target proteins. In support of this, our previous study demonstrated that the stimulator of interferon genes (STING) signaling platform functions as a hub in macrophage immunity and has a profound impact on the prognosis of sepsis. Interestingly, we found that itaconate, an endogenous immunomodulator, can significantly inhibit the activation of STING signaling. Moreover, 4-octyl itaconate (4-OI), which is a permeable itaconate derivative, could alkylate cysteine sites 65, 71, 88, and 147 of STING, thereby inhibiting its phosphorylation and downregulating the production of related inflammatory factors. Furthermore, itaconate and 4-OI inhibited the production of inflammatory factors in sepsis models. Our results have broadened the role of the IRG1-Itaconate axis in immunomodulation and highlighted itaconate and its derivatives as potential therapeutic agents in sepsis.</p>
Project description:Here, we report that macrophage-derived itaconate exerts a significant suppressive effect on dendritic cell (DC) function. To delve deeper into the impact of itaconate on DCs, we treated bone marrow-derived dendritic cells with 4-OI, a derivative of itaconate, and conducted RNA-seq analysis. Gene-set enrichment analysis (GSEA) of 4-OI-treated DCs showed a general downregulation of innate immunity and immune-response pathways.
Project description:4 octyl itaconate (4-OI) is a known anti-inflammatory chemical that activates Nrf2 signaling. Here, we are exploring the capacities of 4-OI to promote the spread of oncolytic Vesicular Stomatitis Virus (VSVd51M) in pLenti control kidney adenocarcinoma 786-O cell line and in Nrf2 knock out 786-O.
Project description:To investigate the role of Irg1-itacaonate on Nrf2-transcripting genes in macrophages, RAW264.7 cells were cultured and were treated 120 μM 4-octyl itaconate (4-OI) or Vehicle for 8 h. We then performed ChIP-seq analysis from 4-OI or vehicle treated RAW264.7 cells using Nrf2 or IgG antibodies.
Project description:To investigate the role of Irg1-itaconate or Hemin in macrophages, bone marrow-derived macrophages (BMDMs) from WT mice and Irg1 KO mice were extracted and cultured. After 7 days of cells culture, the BMDMs from WT mice were treated 120 μM 4-octyl itaconate (4-OI) for 15 h, 15 μM Hemin for 12 h, and the BMDMs from Irg1 KO mice and WT mice were treated Vehicle for 12 h. We then performed gene expression profiling analysis using data obtained from RNA-seq of 4-OI, Hemin or vehicle treated WT BMDMs and Irg1 KO BMDMs.
Project description:Analysis of gene expression in WT and ISG15 KO endothelial cells, stimulated with IFNa and treated with Itaconic acid (IA), Dimethyl itaconate (DI), 4-octyl itaconate (4OI) and ruxolitinib (RUX) to investigate the effects of treatments on transcriptional response in the cells.
Project description:WT and ISG15 KO macrophages were stimulated with interferon-a and treated with itaconic acid and 4-octyl itaconate as they are known for anti-inflammatory effects. In this experiment, we further explored how itaconic acid and 4-octyl itaconate can play its role keeping ruxolitinib as a positive control in order to inhibit JAK/STAT pathway
Project description:Enhanced osteoclast-mediated bone erosion is a prominent hallmark of rheumatoid arthritis. Emerging evidence suggests that this process is facilitated by metabolic dysregulation of osteoclasts. The mitochondrial enzyme aconitate decarboxylase 1 (Acod1, also known as immune responsive gene 1 [Irg1]) serves as a mediator between the metabolic condition and the functional state of different types of cells. Acod1-deficient mice are characterized by enhanced osteoclast differentiation and bone erosion in an inflammatory arthritis model, while therapeutic treatment with the itaconate-derivative 4-octyl-itaconate (4-OI) alleviates the disease phenotype in experimental arthritis. To ascertain the influence of the Acod1-itaconate axis on the genomic transcriptional network of osteoclasts, we performed a whole transcriptome RNA sequencing analysis with fully differentiated osteoclasts from WT and Acod1-deficient mice that were cultured in the presence or absence of 4-OI.