Project description:Patients with myelodysplastic syndromes (MDSs) display severe anemia but the mechanisms underlying this phenotype are incompletely understood. Right open-reading-frame kinase 2 (RIOK2) encodes a protein kinase located at 5q15, a region frequently lost in patients with MDS del(5q). Here we show that hematopoietic cell-specific haploinsufficient deletion of Riok2 (Riok2f/+Vav1cre) led to reduced erythroid precursor frequency leading to anemia. Proteomic analysis of Riok2f/+Vav1cre erythroid precursors suggested immune system activation, and transcriptomic analysis revealed an increase in p53-dependent interleukin (IL)-22 in Riok2f/+Vav1cre CD4+ T cells (TH22). Further, we discovered that the IL-22 receptor, IL-22RA1, was unexpectedly present on erythroid precursors. Blockade of IL-22 signaling alleviated anemia not only in Riok2f/+Vav1cre mice but also in wild-type mice. Serum concentrations of IL-22 were increased in the subset of patients with del(5q) MDS as well as patients with anemia secondary to chronic kidney disease. This work reveals a possible therapeutic opportunity for reversing many stress-induced anemias by targeting IL-22 signaling.
Project description:- tryptic digests of CD34+ FAC-sorted cells dilution series acquired in DIA mode on Orbitrap Lumos - tryptic digests of HSC, MEP, GMP, CMP FAC-sorted cells acquired in DIA mode on Orbitrap Lumos
Project description:Microarray experiments were performed using FAC-sorted young photoreceptors to analyze their transcriptome in comparison to remaining retinal cells at same developmental stage and retinal progenitors.
Project description:Microarray experiments were performed using FAC-sorted young photoreceptors to analyze their transcriptome in comparison to remaining retinal cells at same developmental stage and retinal progenitors. For each replicate, retinae of 6 to 8 postnatal day 0 pNestin-GFP or postnatal day 4 rhoEGFP mice were dissected and FAC-sorted based on GFP expression. RNA of fractions was isolated and subsequently analyzed with microarray experiment.
Project description:We achieved local recombination of striatal astrocytes and silenced Notch signalling activity through Rbpj-K depletion. Recombined astrocytes and neural progenitors generated from the striaral glia were FAC sorted and their molecular profile investigated in a scRNAseq experiment performed 5 wpi.
Project description:Rapid expansion of stress erythroid progenitors is a key response to acute anemia during stress erythropoiesis. Besides the rapidly amplifying progenitors, a small population of stem-cell like erythroid progenitors undergoes limited number of cell divisions and maintains their stemness. In this study, we addressed the differences in expression profiles of regulatory genes between two stress erythroid progenitor populations that were identified by proliferation capacity and physiological status. We used microarrays to detail the gene expression profiles of stem-cell like stress erythroid progenitors and rapidly amplifying stress erythroid progenitors during stress erythropoiesis.
Project description:Psychological stress has been previously reported to worsen symptoms of inflammatory bowel disease (IBD). Similarly, intestinal tertiary lymphoid organs (TLOs) are associated with more severe inflammation. While there is active debate about the role of TLOs and stress in IBD pathogenesis, there are no studies investigating TLO formation in the context of psychological stress. Our mouse model of Crohn's disease-like ileitis, the SAMP1/YitFc (SAMP) mouse, was subjected to 56 consecutive days of restraint stress (RS). Stressed mice had significantly increased colonic TLO formation. However, stress did not significantly increase small or large intestinal inflammation in the SAMP mice. Additionally, 16S analysis of the stressed SAMP microbiome revealed no genus-level changes. Fecal microbiome transplantation into germ-free SAMP mice using stool from unstressed and stressed mice replicated the behavioral phenotype seen in donor mice. However, there was no difference in TLO formation between recipient mice. Stress increased the TLO formation cytokines interleukin-23 (IL-23) and IL-22 followed by up-regulation of antimicrobial peptides. SAMP × IL-23r-/- (knockout [KO]) mice subjected to chronic RS did not have increased TLO formation. Furthermore, IL-23, but not IL-22, production was increased in KO mice, and administration of recombinant IL-22 rescued TLO formation. Following secondary colonic insult with dextran sodium sulfate, stressed mice had reduced colitis on both histology and colonoscopy. Our findings demonstrate that psychological stress induces colonic TLOs through intrinsic alterations in IL-23 signaling, not through extrinsic influence from the microbiome. Furthermore, chronic stress is protective against secondary insult from colitis, suggesting that TLOs may function to improve the mucosal barrier.
Project description:Characterision of the bioactive peptide content of M-cells. M-cells derived from organoids were sorted using FAC sorting techniques and were lysed and analysed by LC-MS/MS using a peptidomics approach.
Project description:IL-22 is a member of the IL-10 cytokine family involved in host protection against extracellular pathogens, by promoting epithelial cell regeneration and barrier functions. Dysregulation of IL-22 production has also frequently been observed in acute respiratory distress syndrome (ARDS) and several chronic inflammatory and autoimmune diseases. We have previously described that human CD28, a crucial co-stimulatory receptor necessary for full T cell activation, is also able to act as a TCR independent signaling receptor and to induce the expression of IL-17A and inflammatory cytokines related to Th17 cells, which together with Th22 cells represent the main cellular source of IL-22. Here we characterized the role of CD28 autonomous signaling in regulating IL-22 expression in human CD4+ T cells. We show that CD28 stimulation in the absence of TCR strongly up-regulates IL-22 gene expression and secretion. As recently observed for IL-17A, we also found that CD28-mediated regulation of IL-22 transcription requires the cooperative activities of both IL-6-activated STAT3 and RelA/NF-κB transcription factors. CD28-mediated IL-22 production also promotes the barrier functions of epithelial cells by inducing mucin and metalloproteases expression. Finally, by using specific inhibitory drugs, we also identified CD28-associated class 1A phosphatidylinositol 3-kinase (PI3K) as a pivotal mediator of CD28-mediated IL-22 expression and IL-22-dependent epithelial cell barrier functions.