Identification and characterization of microRNAs in oilseed rape (Brassica napus) responsive to infection with the pathogenic fungus Verticillium longisporum using Brassica AA (Brassica rapa) and CC (Brassica oleracea) as reference genomes SummaryVerticillium longisporum, a soil-borne pathogenic fungus, causes vascular disease in oilseed rape (Brassica napus). We proposed that plant microRNAs (miRNAs) are involved in the plant-V. longisporum interaction.To identify oilseed rape miRNAs, we deep-sequenced two small RNA libraries made from V. longisporum infected/noninfected roots and employed Brassica rapa and Brassica oleracea genomes as references for miRNA prediction and characterization.We identified 893 B. napus miRNAs representing 360 conserved and 533 novel miRNAs, and mapped 429 and 464 miRNAs to the AA and CC genomes, respectively. Microsynteny analysis with the conserved miRNAs and their flanking protein coding sequences revealed 137 AA-CC genome syntenic miRNA pairs and 61 AA and 42 CC genome-unique miRNAs. Sixtytwo miRNAs were responsive to the V. longisporum infection. We present data for specific interactions and simultaneously reciprocal changes in the expression levels of the miRNAs and their targets in the infected roots. We demonstrate that miRNAs are involved in the plant-fungus interaction and that miRNA168-Argonaute 1 (AGO1) expression modulation might act as a key regulatory module in a compatible plant-V. longisporum interaction.Our results suggest that V. longisporum may have evolved a virulence mechanism by interference with plant miRNAs to reprogram plant gene expression and achieve infection.
Background Dimethyl fumarate (DMF) is a treatment for moderate-to-severe psoriasis and multiple sclerosis. DMF therapy typically improves skin inflammation within the first 3 months of treatment. DMF is a prodrug that generates the hydroxycarboxylic acid receptor 2 (HCA2) agonist, monomethyl fumarate (MMF). Despite widespread clinical use, DMF's mechanism of action is not fully understood. Objectives We wished to characterize the changes induced by DMF in peripheral neutrophils within the first 3 months of treatment to better understand its early antipsoriatic effects. Methods Flow cytometry was used to assess T-cell and neutrophil frequencies, apoptosis and activation phenotype. In vitro culture of neutrophils with DMF and MMF was used to evaluate apoptosis and HCA2 internalization. Serum levels of neutrophil degranulation products were measured by enzyme-linked immunosorbent assay. Results Patients with psoriasis had significantly higher leucocyte counts at baseline compared with controls, with a large population of pro-inflammatory CD62L lo CD11b bright neutrophils. Analysis revealed that DMF treatment reduced the frequency of CD62L lo CD11b bright neutrophils and serum levels of neutrophil activation markers. This reduction was not linked to increased apoptosis. Conclusions Our results reveal a novel in vivo effect of DMF therapy on proinflammatory neutrophils that likely contributes to this treatment's antipsoriatic efficacy.What is already known about this topic?• Oral dimethyl fumarate (DMF) improves psoriasis within the first 3 months of treatment but its mechanism of action has not been fully elucidated.• It is known that DMF is a prodrug for monomethyl fumarate, which itself is a hydroxycarboxylic acid receptor 2 (HCA2) agonist.• Treatment with oral DMF is linked to the loss of peripheral T cells (which do not express HCA2), while changes in neutrophils (which are HCA2-positive) have not been widely reported.What does this study add?• This study describes that patients with untreated psoriasis have elevated numbers of pro-inflammatory CD62L lo CD11b bright neutrophils.• DMF therapy reduces both the number of pro-inflammatory neutrophils and the serum concentrations of markers of neutrophil activation within the first 3 months of treatment.• DMF treatment does not promote neutrophil apoptosis in patients with psoriasis.
Fumaric acid esters (FAEs) are used as an oral treatment for psoriasis. Dimethylfumarate (DMF) and its metabolite monomethylfumarate (MMF) are regarded as the pharmacologically active moieties. Indoleamine 2,3-dioxygenase (IDO) is the key enzyme for the metabolism of tryptophan. The kynurenine pathway is established as a major regulator of innate and adaptive immunity. Here, we investigated the effect of DMF and MMF on IDO activity and expression in human peripheral blood mononuclear cells (PBMCs). IDO activity was determined by measuring the concentration of kynurenine in the culture medium using a HPLC technique. IDO and kynureninase protein expressions were analysed by Western blot. Our results demonstrated that DMF and MMF dose-dependently reduced the levels of L-kynurenine in PBMCs activated by interferon-γ (IFN-γ). Furthermore, MMF had an inhibitory effect on IDO activity in vitro with an ED of 10 μmol/L, a value within the therapeutic concentration range for this molecule. We also observed that IDO and kynureninase expressions were reduced in PBMCs in a dose-dependent manner by DMF and MMF. The results of our study show that DMF and MMF (in therapeutic concentrations) inhibited IDO and kynureninase activity and expression in a NF-κB-dependent manner in PBMCs while also decreasing the level of L-kynurenine in these cells. As we found that FAEs inhibit both IDO expression and enzymatic activity leading to a modulation of tryptophan degradation, we believe this effect may contribute to the clinical efficacy of this drug in psoriasis by downregulating pro-inflammatory mediators generated by the kynurenine pathway.
Antigen‐specific T lymphocytes are the central regulators of tolerance versus immune pathology against otherwise innocuous antigens and key targets of antigen‐specific immune therapy. Recent advances in the understanding of T cells in tolerance and allergy resulted from improved technologies to directly characterize allergen‐specific T cells by multiparameter flow cytometry or single‐cell sequencing. This unravelled phenotypically and functionally distinct populations, such as Type 2a T helper cells (Th2a), follicular Th cells (Tfh), regulatory T cells (Treg), Type 1 regulatory T cells (Tr1), and follicular T regulatory cells. Here we will discuss the role of the different Th‐cell subsets in the healthy state, during sensitization and development of allergy, and in tolerance induction by allergen immunotherapy (AIT). To date, the mechanisms of AIT as the only causal treatment of allergy are not completely understood. The analyses of allergen‐specific T cells directly ex vivo during AIT support the concept of specific‐Th2(a) cell deletion rather than an expansion of allergen‐specific Tr1 or Treg cells as underlying mechanism.
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