Search

BackgroundThe decrease of IL-2 level is believed to play an important role in the disease occurrence and development of SLE, but the relevant mechanisms have not been fully clarified. Many studies have found that the level of soluble interleukin 2 receptor α (sIL-2Rα) in SLE patients is significantly increased. Considering the fact that sIL-2Rα has the ability to bind IL-2, we want to know whether the increased sIL-2Rα has some impact on the level and function of IL-2 in SLE patients.MethodsNew onset SLE patients, treated SLE patients and healthy volunteers were recruited. The levels of serum IL-2, IL-2 mRNA in CD3+ T cells and serum sIL-2Rα were detected and compared in these subjects. Two mixed solid-phase sandwich ELISA system were designed to measure exclusively the heterodimers complex of sIL-2Rα/IL-2. The sera from SLE patients were pretreated with or without immune complex dissociation solution and detected for IL-2 levels. IL-2 standard or serum from HCs were used to co-incubate with recombinant sIL-2Rα or serum samples with high levels of sIL-2Rα and detected for IL-2 levels by ELISA. The inhibitory effect of sIL-2Rα on IL-2 biological activity was investigated by CTLL-2 cell proliferation assay. The frequencies and absolute counts of Treg cells were detected by flow cytometry before and after the addition of recombinant sIL-2Rα.ResultsThe levels of serum IL-2 in SLE patients were significantly decreased and negatively correlated with SLEDAI. However, there was no significant difference in IL-2 mRNA levels in CD3+ T cells between SLE patients and healthy controls. The levels of serum sIL-2Rα in SLE patients were significantly increased, positively correlated with the SLEDAI and negatively correlated with the levels of serum IL-2. sIL-2Rα was shown to bind to IL-2 to form immune complex, resulting in false reduction in the detection level of serum IL-2 and significant decrease in biological activity of IL-2. The increase of sIL-2Rα was demonstrated to be one of the important mechanisms for the obstruction of Treg cells differentiation in SLE patients.ConclusionIncreased serum sIL-2Rα can bind to IL-2, leading to obstruction of IL-2 activity and Treg cells differentiation.

BackgroundT-SPOT.TB (T-SPOT) is widely used for the detection of Mycobacterium tuberculosis infection by detecting interferon-gamma (IFN-γ) release in T lymphocytes. This assay is performed on peripheral blood mononuclear cells (PBMCs) separated by Ficoll density gradient centrifugation, which often contain some residual platelets. Here, we investigated the impact of platelets on T-SPOT assay and related mechanisms.MethodsThe correlation between platelet count, platelet-to-lymphocyte ratio (PLR), and the IFN-γ secreting T cells (ISCs) in positive control wells of T-SPOT assay were retrospectively analyzed. T-SPOT assay was performed with un-treated PBMCs, platelets-removed PBMCs, and platelets-enriched PBMCs to confirm the impact of platelets on T-SPOT assay. The activation of platelets and their impact on IFN-γ production in T cells were detected by flow cytometry (FCM). Platelets and T cells were cultured in a mixed culture system and co-culture system respectively, followed by detection of the frequencies of IFN-γ-producing T cells and the levels of intracellular IFN-γ in T cells by FCM. Moreover, the effect of platelet releasate on the T-SPOT assay was evaluated.ResultsThe ISCs in positive control wells of the T-SPOT assay showed a significant decrease with the increase in platelet count. The PLR of the peripheral blood were negatively correlated with the ISCs in positive control wells of the T-SPOT assay. Removal or enrichment of platelets significantly increased or decreased the ISCs and the positive rate of T-SPOT. Inhibition of platelet activation significantly increased the ISCs of T-SPOT. The frequencies of IFN-γ-producing T cells in PBMCs and the levels of intracellular IFN-γ were significantly reduced by the addition of platelets, both in the mixed culture system and the co-culture system. Platelet releasate upon thrombin activation significantly decreased the ISCs of T-SPOT.ConclusionsPlatelets correlate with negative T-SPOT results by inhibiting IFN-γ production in T cells via degranulation.

The increasing prevalence and transmission of the carbapenem resistance gene blaNDM–5 has led to a severe threat to public health. So far, blaNDM–5 has been widely detected in various species of Enterobacterales and different hosts across various cities. However, there is no report on the blaNDM–5– harboring Morganella morganii. In January 2016, the first NDM-5-producing Morganella morganii L241 was found in a stool sample of a patient diagnosed as recurrence of liver cancer in China. Identification of the species was performed using 16S rRNA gene sequencing. Carbapenemase genes were identified through both PCR and sequencing. To investigate the characteristics and complete genome sequence of the blaNDM–5-harboring clinical isolate, antimicrobial susceptibility testing, S1 nuclease pulsed field gel electrophoresis, Southern blotting, transconjugation experiment, complete genome sequencing, and comparative genomic analysis were performed. M. morganii L241 was found to be resistant to broad-spectrum cephalosporins and carbapenems. The complete genome of L241 is made up from both a 3,850,444 bp circular chromosome and a 46,161 bp self-transmissible IncX3 plasmid encoding blaNDM–5, which shared a conserved genetic context of blaNDM–5 (ΔIS3000-ΔISAba125-IS5-blaNDM–5-ble-trpF-dsbC-IS26). BLASTn analysis showed that IncX3 plasmids harboring blaNDM genes have been found in 15 species among Enterobacterales from 13 different countries around the world thus far. In addition, comparative genomic analysis showed that M. morganii L241 exhibits a close relationship to M. morganii subsp. morganii KT with 107 SNPs. Our research demonstrated that IncX3 is a key element in the worldwide dissemination of blaNDM-5 among various species. Further research will be necessary to control and prevent the spread of such plasmids.