Altered expression of autoimmune regulator in infant down syndrome thymus, a possible contributor to an autoimmune phenotype.

To access publisher's full text version of this article click on the hyperlink at the bottom of the pageDown syndrome (DS), caused by trisomy of chromosome 21, is associated with immunological dysfunctions such as increased frequency of infections and autoimmune diseases. Patients with DS share clinical features, such as autoimmune manifestations and specific autoantibodies, with patients affected by autoimmune polyendocrine syndrome type 1. Autoimmune polyendocrine syndrome type 1 is caused by mutations in the autoimmune regulator (AIRE) gene, located on chromosome 21, which regulates the expression of tissue-restricted Ags (TRAs) in thymic epithelial cells. We investigated the expression of AIRE and TRAs in DS and control thymic tissue using quantitative PCR. AIRE mRNA levels were elevated in thymic tissue from DS patients, and trends toward increased expression of the AIRE-controlled genes INSULIN and CHRNA1 were found. Immunohistochemical stainings showed altered cell composition and architecture of the thymic medulla in DS individuals with increased frequencies of AIRE-positive medullary epithelial cells and CD11c-positive dendritic cells as well as enlarged Hassall's corpuscles. In addition, we evaluated the proteomic profile of thymic exosomes in DS individuals and controls. DS exosomes carried a broader protein pool and also a larger pool of unique TRAs compared with control exosomes. In conclusion, the increased AIRE gene dose in DS could contribute to an autoimmune phenotype through multiple AIRE-mediated effects on homeostasis and function of thymic epithelial cells that affect thymic selection processes.Swedish Research Council 80409601 Marianne and Marcus Wallenberg Foundation Region Vastra Gotaland ALFGBG-771712 Arbetsmarknadens Forsakringsaktiebolag 100258 IngaBritt and Arne Lundbergs Research Foundation AnnMari and Per Ahlqvists Foundation Gothenburg Medical Society Wilhelm and Martina Lundgrens Research Foundatio

3D Printning – möjligheter och begränsningar

I examensarbetet redogörs för de olika möjligheter samt begränsningar inom 3D printning. 3D printning är en additiv tillverkningsmetod, vilket betyder att material läggs till istället för att det tas bort. I arbetet presenteras själva printningsprocessen med olika delsteg. I arbetet beskrivs de vanligaste teknikerna inom 3DP, vilka är FDM, SLS, SLA, Z-Corporation, DMLS och Vaxmetoden. Inom 3DP finns det många möjligheter då det gäller materialval. De vanligaste materialen som lämpar sig för 3D printning presenteras i detta examensarbete. Olika användningsområden för 3D printade föremål och 3D printningens framtid beskrivs också. Fokus ligger både på 3D printning för företag samt för privatpersoner. Syftet med detta arbete är att redogöra dagens teknik och möjligheter inom 3DP. Arbetet innehåller även en praktisk del där det printades ut en bult och en mutter. Printningen skedde i Sello bibliotek. Sello är ett köpcentrum beläget i Alberga i Esbo. Biblioteket har fyra 3D printers som man får använda gratis. Resultatet av printningen blev rätt så bra, gängorna på bulten och muttern fungerar men på grund av ett litet fel i kalibreringen så är föremålen inte kosmetiskt sett så vackra.Opinnäytetyössä kerrotaan 3D-tulostuksesta sekä sen mahdollisuuksista ja rajoituksista. 3D-tulostus on additiivinen tuotantomenetelmä, eli materiaalia lisätään poistamisen sijasta. Tässä työssä kerrotaan tulostusprosessista alivaiheineen. Opinnäytetyössä kerrotaan tavallisimmista 3D-tulostus tekniikoista, jotka ovat FDM, SLS, SLA, Z-Corporation, DMLS ja Vahametodi. 3D-tulostuksessa voi valita laajasta materiaalivalikoimasta, ja tavallisimmat materiaalit esitetään tässä työssä. Tulostetun kappaleen eri käyttötarkoituksista ja 3D-tulostamisen tulevaisuudesta kerrotaan. Painopiste on sekä yritysten että yksityishenkilöiden 3D-tulostamisessa. Tämän työn tarkoituksena on esittää 3D-tulostamisen uusimmat tekniikat sekä mahdollisuudet. Työhön kuuluu myös käytännön osuus, jossa kerrotaan 3D- tulostamiskokemuksesta, jossa tulostettiin kolmiuloitteinen pultti ja mutteri. Tulostus suoritettiin Sellon kirjastossa. Sello on kauppakeskus joka sijaitsee Espoon Leppävaarassa. Kirjastossa on neljä 3D tulostinta joita saa käyttää maksutta. Tulostuskokeilun tulos on melko hyvä. Pultin ja mutterin kierteet toimivat mutta kalibrointivirheen takia osat eivät ole kovin kauniita.In this thesis 3D printing and its possibilities and restrictions are explained. 3D printing is an additive manufacturing method which means that material is added instead of subtracted. The printing process including its sub-steps is explained. The most common techniques, also explained in this thesis, are FDM, SLS, SLA, Z-Corporation and the Wax method. In 3D printing one can choose from many different materials. The most common materials are explained in this thesis. The area of use for the printed parts, and the future of 3DP are described. The focus is on 3D printing for both companies and individuals. The purpose of this thesis is to account for the newest technologies within 3D printing and its opportunities. The practical section refers to a 3D printing experience of a bolt and a nut. The printing happened in Sello library. Sello is a shopping center in Leppävaara in Espoo. The library have four 3D printers which one may use free of charge. The result of the printing was quite good. The threads on the bolt and the nut are working as intended, but due to a fault in the calibration the surfaces of the bolt and the nut aren’t very smooth

Deep learning and conformal prediction for hierarchical analysis of large-scale whole-slide tissue images

With the increasing amount of image data collected from biomedical experiments there is an urgent need for smarter and more effective analysis methods. Many scientific questions require analysis of image subregions related to some specific biology. Finding such regions of interest (ROIs) at low resolution and limiting the data subjected to final quantification at high resolution can reduce computational requirements and save time. In this paper we propose a three-step pipeline: First, bounding boxes for ROIs are located at low resolution. Next, ROIs are subjected to semantic segmentation into sub-regions at mid-resolution. We also estimate the confidence of the segmented sub-regions. Finally, quantitative measurements are extracted at high resolution. We use deep learning for the first two steps in the pipeline and conformal prediction for confidence assessment. We show that limiting final quantitative analysis to sub regions with high confidence reduces noise and increases separability of observed biological effects

Premature thymic involution is independent of structural plasticity of the thymic stroma

The thymus is the organ devoted to T-cell production. The thymus undergoes multiple rounds of atrophy and redevelopment before degenerating with age in a process known as involution. This process is poorly understood, despite the influence the phenomenon has on peripheral T-cell numbers. Here we have investigated the FVB/N mouse strain, which displays premature thymic involution. We find multiple architectural and cellular features that precede thymic involution, including disruption of the epithelial-endothelial relationship and a progressive loss of pro-T cells. The architectural features, reminiscent of the human thymus, are intrinsic to the nonhematopoietic compartment and are neither necessary nor sufficient for thymic involution. By contrast, the loss of pro-T cells is intrinsic to the hematopoietic compartment, and is sufficient to drive premature involution. These results identify pro-T-cell loss as the main driver of premature thymic involution, and highlight the plasticity of the thymic stroma, capable of maintaining function across diverse interstrain architectures.status: publishe

Characterization of Human Thymic Exosomes

Exosomes are nanosized membrane-bound vesicles that are released by various cell types and are capable of carrying proteins, lipids and RNAs which can be delivered to recipient cells. Exosomes play a role in intercellular communication and have been described to mediate immunologic information. In this article we report the first isolation and characterization of exosomes from human thymic tissue. Using electron microscopy, particle size determination, density gradient measurement, flow cytometry, proteomic analysis and microRNA profiling we describe the morphology, size, density, protein composition and microRNA content of human thymic exosomes. The thymic exosomes share characteristics with previously described exosomes such as antigen presentation molecules, but they also exhibit thymus specific features regarding surface markers, protein content and microRNA profile. Interestingly, thymic exosomes carry proteins that have a tissue restricted expression in the periphery which may suggest a role in T cell selection and the induction of central tolerance. We speculate that thymic exosomes may provide the means for intercellular information exchange necessary for negative selection and regulatory T cell formation of the developing thymocytes within the human thymic medulla.Funding Agencies|Swedish Research Council|80409601|Region Vastra Gotaland|ALFGBG-771712|AFA Forsakring|100258|IngaBritt and Arne Lundbergs Research Foundation||Gothenburg Medical Society||</p

Human lung conventional dendritic cells orchestrate lymphoid neogenesis during chronic obstructive pulmonary disease

Rationale: Emerging evidence supports a crucial role for tertiary lymphoid organs (TLOs) in chronic obstructive pulmonary disease (COPD) progression. However, mechanisms of immune cell activation leading to TLOs in COPD remain to be defined. Objectives: To examine the role of lung dendritic cells (DCs) in T follicular helper (Tfh)-cell induction, a T-cell subset critically implicated in lymphoid organ formation, in COPD. Methods: Myeloid cell heterogeneity and phenotype were studied in an unbiased manner via single-cell RNA sequencing on HLA-DR+ cells sorted from human lungs. We measured the in vitro capability of control and COPD lung DC subsets, sorted using a fluorescence-activated cell sorter, to polarize IL-21(+)CXCL13(+)(IL-21-positive and C-X-C chemokine ligand type 13-positive) Tfh-like cells. In situ imaging analysis was performed on Global Initiative for Chronic Obstructive Lung Disease stage IV COPD lungs with TLOs. Measurements and Main Results: Single-cell RNA-sequencing analysis revealed a high degree of heterogeneity among human lung myeloid cells. Among these, conventional dendritic type 2 cells (cDC2s) showed increased induction of IL-21(+)CXCL13(+) Tfh-like cells. Importantly, the capacity to induce IL-21(+)Tfh-like cells was higher in cDC2s from patients with COPD than in those from control patients. Increased Tfh-cell induction by COPD cDC2s correlated with increased presence of Tfh-like cells in COPD lungs as compared with those in control lungs, and cDC2s colocalized with Tfh-like cells in TLOs of COPD lungs. Mechanistically, cDC2s exhibited a unique migratory signature and (transcriptional) expression of several pathways and genes related to DC-induced Tfh-cell priming. Importantly, blocking the costimulatory OX4OL (OX4O ligand)-OX4O axis reduced Tfh-cell induction by control lung cDC2s. Conclusions: In COPD lungs, we found lung EBI2(+) (Epstein-Barr virus-induced gene 2-positive) OX-40L-expressing cDC2s that induced IL-21(+) Tfh-like cells, suggesting an involvement of these cells in TLO formation

miRNAs from exosomes of different origin compared in a Venn-diagram.

<p>Green circle: shared miRNAs isolated from human thymic EVs (n = 4). Red circle: miRNAs isolated from exosomes descending from Raji B cells. Blue circle: miRNAs isolated from exosomes descending from Jurkat T cells. 40 miRNAs were shared between all three sources while 38 miRNAs were exclusively found in EVs from human thymic tissue.</p

Morphological characteristics of human thymic EVs.

<p>(A) EVs from human thymic cultures visualized with electron microscopy. Arrow heads point toward EVs with a typical exosomal cup-shaped morphology and a size range of 50–100 nm. Samples from 3 individuals were analyzed with electron microscopy. (B) Size distribution of isolated EVs observed in a NanoSight LM10. The data was analyzed with Nanoparticle tracking analysis software, with a minimum expected particle size setting of 30 nm and the number of tracks analyzed for each sample exceeding 200. In agreement with exosome characteristics, most of the isolated EVs have a size of less than 100 nm. Data is presented as mean ± SEM as a result of 5 analyzed samples. (C) Density profile of isolated EVs. EVs were layered on top of a D₂O/sucrose gradient and centrifuged at 100,000 g for 14 hours. Fractions were collected and their density and protein concentration was measured. The density of the isolated EVs peaked at 1.18–1.19 g/ml, which is within the density range of exosomes. Data is presented as mean ± SEM as a result of 5 density gradients.</p