Projected t-SNE for batch correction

Motivation: Low-dimensional representations of high-dimensional data are routinely employed in biomedical research to visualize, interpret and communicate results from different pipelines. In this article, we propose a novel procedure to directly estimate t-SNE embeddings that are not driven by batch effects. Without correction, interesting structure in the data can be obscured by batch effects. The proposed algorithm can therefore significantly aid visualization of high-dimensional data. Results: The proposed methods are based on linear algebra and constrained optimization, leading to efficient algorithms and fast computation in many high-dimensional settings. Results on artificial single-cell transcription profiling data show that the proposed procedure successfully removes multiple batch effects from t-SNE embeddings, while retaining fundamental information on cell types. When applied to single-cell gene expression data to investigate mouse medulloblastoma, the proposed method successfully removes batches related with mice identifiers and the date of the experiment, while preserving clusters of oligodendrocytes, astrocytes, and endothelial cells and microglia, which are expected to lie in the stroma within or adjacent to the tumours. Contact: [email protected]

Differential expression, function and response to inflammatory stimuli of 11β-hydroxysteroid dehydrogenase type 1 in human fibroblasts: a mechanism for tissue-specific regulation of inflammation

Stromal cells such as fibroblasts play an important role in defining tissue-specific responses during the resolution of inflammation. We hypothesized that this involves tissue-specific regulation of glucocorticoids, mediated via differential regulation of the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Expression, activity and function of 11β-HSD1 was assessed in matched fibroblasts derived from various tissues (synovium, bone marrow and skin) obtained from patients with rheumatoid arthritis or osteoarthritis. 11β-HSD1 was expressed in fibroblasts from all tissues but mRNA levels and enzyme activity were higher in synovial fibroblasts (2-fold and 13-fold higher mRNA levels in dermal and synovial fibroblasts, respectively, relative to bone marrow). Expression and activity of the enzyme increased in all fibroblasts following treatment with tumour necrosis factor-α or IL-1β (bone marrow: 8-fold and 37-fold, respectively, compared to vehicle; dermal fibroblasts: 4-fold and 14-fold; synovial fibroblasts: 7-fold and 31-fold; all P < 0.01 compared with vehicle). Treatment with IL-4 or interferon-γ was without effect, and there was no difference in 11β-HSD1 expression between fibroblasts (from any site) obtained from patients with rheumatoid arthritis or osteoarthritis. In the presence of 100 nmol/l cortisone, IL-6 production – a characteristic feature of synovial derived fibroblasts – was significantly reduced in synovial but not dermal or bone marrow fibroblasts. This was prevented by co-treatment with an 11β-HSD inhibitor, emphasizing the potential for autocrine activation of glucocorticoids in synovial fibroblasts. These data indicate that differences in fibroblast-derived glucocorticoid production (via the enzyme 11β-HSD1) between cells from distinct anatomical locations may play a key role in the predeliction of certain tissues to develop persistent inflammation

Expression of FcRL4 defines a pro-inflammatory, RANKL-producing B cell subset in rheumatoid arthritis

OBJECTIVES: The success of B cell targeting therapies has highlighted the importance of B cells in rheumatoid arthritis pathogenesis. We have previously shown that B cells in the RA synovium are capable of producing pro-inflammatory and bone-destructive cytokines including RANKL. Here we sought to characterise the nature and functional relevance of the RANKL-producing B cell subset in the RA synovium. METHODS: Synovial fluid and peripheral blood B cells from patients with RA were analysed by flow cytometry for markers of B cell differentiation and activation and for chemokine receptors. FcRL4(+) and FcRL4(−) B cells sorted from synovial fluid were analysed for cytokine expression using Taqman low-density arrays. Synovial tissue biopsies obtained from patients with RA were analysed by immunofluorescence for CD20, RANKL and FcRL4. FCRL4 mRNA expression was determined in synovial tissue of RA patients and non-inflammatory control subjects by real-time PCR. RESULTS: RANKL-producing B cells in RA synovial tissue and fluid were identified as belonging to a distinct subset of B cells defined by expression of the transmembrane protein FcRL4. FcRL4+ B cells express a distinct combination of cytokines and surface proteins indicating a function distinct from that of FcRL4− B cells. Notably, FcRL4+ B cells expressed high levels of TNF-α and RANKL mRNA. CONCLUSIONS: We have identified a novel pro-inflammatory B cell population in the RA synovium which is defined by expression of FcRL4 and responsible for RANKL production. This B cell population expresses high levels of CD20, and its removal by rituximab may contribute to the anti-inflammatory effect of this drug

Treatment of inflammatory arthritis via targeting of tristetraprolin, a master regulator of pro-inflammatory gene expression

Objectives Tristetraprolin (TTP), a negative regulator of many pro-inflammatory genes, is strongly expressed in rheumatoid synovial cells. The mitogen-activated protein kinase (MAPK) p38 pathway mediates the inactivation of TTP via phosphorylation of two serine residues. We wished to test the hypothesis that these phosphorylations contribute to the development of inflammatory arthritis, and that, conversely, joint inflammation may be inhibited by promoting the dephosphorylation and activation of TTP. Methods The expression of TTP and its relationship with MAPK p38 activity were examined in non-inflamed and rheumatoid arthritis (RA) synovial tissue. Experimental arthritis was induced in a genetically modified mouse strain, in which endogenous TTP cannot be phosphorylated and inactivated. In vitro and in vivo experiments were performed to test anti-inflammatory effects of compounds that activate the protein phosphatase 2A (PP2A) and promote dephosphorylation of TTP. Results TTP expression was significantly higher in RA than non-inflamed synovium, detected in macrophages, vascular endothelial cells and some fibroblasts and co-localised with MAPK p38 activation. Substitution of TTP phosphorylation sites conferred dramatic protection against inflammatory arthritis in mice. Two distinct PP2A agonists also reduced inflammation and prevented bone erosion. In vitro anti-inflammatory effects of PP2A agonism were mediated by TTP activation. Conclusions The phosphorylation state of TTP is a critical determinant of inflammatory responses, and a tractable target for novel anti-inflammatory treatments

Base de dados do Herbário IAN da Embrapa Amazônia Oriental.

Base de Dados do Herbário IAN da Embrapa Amazônia Oriental; Introdução; Histórico; Características da base de dados; Desenvolvimento; Segurança das informações; Usuários; Disponibilização na web.bitstream/item/94025/1/DOC-392.pd

Inflammation causes remodeling of mitochondrial cytochrome c oxidase mediated by the bifunctional gene C15orf48

Dysregulated mitochondrial function is a hallmark of immune-mediated inflammatory diseases. Cytochrome c oxidase (CcO), which mediates the rate-limiting step in mitochondrial respiration, is remodeled during development and in response to changes of oxygen availability, but there has been little study of CcO remodeling during inflammation. Here, we describe an elegant molecular switch mediated by the bifunctional transcript C15orf48, which orchestrates the substitution of the CcO subunit NDUFA4 by its paralog C15ORF48 in primary macrophages. Expression of C15orf48 is a conserved response to inflammatory signals and occurs in many immune-related pathologies. In rheumatoid arthritis, C15orf48 mRNA is elevated in peripheral monocytes and proinflammatory synovial tissue macrophages, and its expression positively correlates with disease severity and declines in remission. C15orf48 is also expressed by pathogenic macrophages in severe coronavirus disease 2019 (COVID-19). Study of a rare metabolic disease syndrome provides evidence that loss of the NDUFA4 subunit supports proinflammatory macrophage functions

Rheumatoid synovial fluid interleukin-17-producing CD4 T cells have abundant tumor necrosis factor-alpha co-expression, but little interleukin-22 and interleukin-23R expression

Introduction\ud Th17 cells have been implicated in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to systematically analyse the phenotype, cytokine profile and frequency of interleukin-17 (IL-17) producing CD4-positive T cells in mononuclear cells isolated from peripheral blood, synovial fluid and synovial tissue of RA patients with established disease, and to correlate cell frequencies with disease activity. \ud \ud Methods\ud Flow cytometry was used to analyse the phenotype and cytokine production of mononuclear cells isolated from peripheral blood (PBMC) (n = 44), synovial fluid (SFMC) (n = 14) and synovium (SVMC) (n = 10) of RA patients and PBMC of healthy controls (n = 13). \ud \ud Results\ud The frequency of IL-17-producing CD4 T cells was elevated in RA SFMC compared with RA PBMC (P = 0.04). However, the frequency of this population in RA SVMC was comparable to that in paired RA PBMC. The percentage of IL-17-producing CD4 T cells coexpressing tumor necrosis factor alpha (TNFα) was significantly increased in SFMC (P = 0.0068). The frequency of IFNγ-producing CD4 T cells was also significantly higher in SFMC than paired PBMC (P = 0.042). The majority of IL-17-producing CD4 T cells coexpressed IFNγ. IL-17-producing CD4 T cells in RA PBMC and SFMC exhibited very little IL-22 or IL-23R coexpression. \ud \ud Conclusions\ud These findings demonstrate a modest enrichment of IL-17-producing CD4 T cells in RA SFMC compared to PBMC. Th17 cells in SFMC produce more TNFα than their PBMC counterparts, but are not a significant source of IL-22 and do not express IL-23R. However, the percentage of CD4 T cells which produce IL-17 in the rheumatoid joint is low, suggesting that other cells may be alternative sources of IL-17 within the joints of RA patients. \ud \u

Expression of chemokines CXCL4 and CXCL7 by synovial macrophages defines an early stage of rheumatoid athritis

BACKGROUND AND OBJECTIVES: For our understanding of the pathogenesis of rheumatoid arthritis (RA), it is important to elucidate the mechanisms underlying early stages of synovitis. Here, synovial cytokine production was investigated in patients with very early arthritis. METHODS: Synovial biopsies were obtained from patients with at least one clinically swollen joint within 12 weeks of symptom onset. At an 18-month follow-up visit, patients who went on to develop RA, or whose arthritis spontaneously resolved, were identified. Biopsies were also obtained from patients with RA with longer symptom duration (>12 weeks) and individuals with no clinically apparent inflammation. Synovial mRNA expression of 117 cytokines was quantified using PCR techniques and analysed using standard and novel methods of data analysis. Synovial tissue sections were stained for CXCL4, CXCL7, CD41, CD68 and von Willebrand factor. RESULTS: A machine learning approach identified expression of mRNA for CXCL4 and CXCL7 as potentially important in the classification of early RA versus resolving arthritis. mRNA levels for these chemokines were significantly elevated in patients with early RA compared with uninflamed controls. Significantly increased CXCL4 and CXCL7 protein expression was observed in patients with early RA compared with those with resolving arthritis or longer established disease. CXCL4 and CXCL7 co-localised with blood vessels, platelets and CD68(+) macrophages. Extravascular CXCL7 expression was significantly higher in patients with very early RA compared with longer duration RA or resolving arthritis CONCLUSIONS: Taken together, these observations suggest a transient increase in synovial CXCL4 and CXCL7 levels in early RA

Inflammation Dynamically Regulates Steroid Hormone Metabolism and Action within Macrophages in Rheumatoid Arthritis

Rationale: In inflammatory diseases such as rheumatoid arthritis (RA), steroid metabolism is a central component mediating the actions of immuno-modulatory glucocorticoids and sex steroids. However, the regulation and function of cellular steroid metabolism within key leukocyte populations such as macrophages remain poorly defined. In this study, the inflammatory regulation of global steroid metabolism was assessed in RA macrophages. Methods: Bulk RNA-seq data from RA synovial macrophages was used to assess transcripts encoding key enzymes in steroid metabolism and signalling. Changes in metabolism were assessed in synovial fluids, correlated to measures of disease activity and functionally validated in primary macrophage cultures. Results: RNA-seq revealed a unique pattern of differentially expressed genes, including changes in genes encoding the enzymes 11β-HSD1, SRD5A1, AKR1C2 and AKR1C3. These correlated with disease activity, favouring increased glucocorticoid and androgen levels. Synovial fluid 11β-HSD1 activity correlated with local inflammatory mediators (TNFα, IL-6, IL-17,), whilst 11β-HSD1, SRD5A1 and AKR1C3 activity correlated with systemic measures of disease and patient pain (ESR, DAS28 ESR, global disease activity). Changes in enzyme activity were evident in inflammatory activated macrophages in vitro and revealed a novel androgen activating role for 11β-HSD1. Together, increased glucocorticoids and androgens were able to suppress inflammation in macrophages and fibroblast-like-synoviocytes. Conclusions: This study underscores the significant increase in androgen and glucocorticoid activation within inflammatory polarized macrophages of the synovium, contributing to local suppression of inflammation. The diminished profile of inactive steroid precursors in postmenopausal women may contribute to disturbances in this process, leading to increased disease incidence and severity.<br/