Publications by authors named "Josephine Lum"

50 Publications

Single-cell analysis of human skin identifies CD14+ type 3 dendritic cells co-producing IL1B and IL23A in psoriasis.

J Exp Med 2021 Sep 19;218(9). Epub 2021 Jul 19.

Singapore Immunology Network, Agency for Science, Technology and Research, Singapore.

Inflammatory skin diseases including atopic dermatitis (AD) and psoriasis (PSO) are underpinned by dendritic cell (DC)-mediated T cell responses. Currently, the heterogeneous human cutaneous DC population is incompletely characterized, and its contribution to these diseases remains unclear. Here, we performed index-sorted single-cell flow cytometry and RNA sequencing of lesional and nonlesional AD and PSO skin to identify macrophages and all DC subsets, including the newly described mature LAMP3+BIRC3+ DCs enriched in immunoregulatory molecules (mregDC) and CD14+ DC3. By integrating our indexed data with published skin datasets, we generated a myeloid cell universe of DC and macrophage subsets in healthy and diseased skin. Importantly, we found that CD14+ DC3s increased in PSO lesional skin and co-produced IL1B and IL23A, which are pathological in PSO. Our study comprehensively describes the molecular characteristics of macrophages and DC subsets in AD and PSO at single-cell resolution, and identifies CD14+ DC3s as potential promoters of inflammation in PSO.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1084/jem.20202345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8292131PMC
September 2021

An epithelial pathway exacerbates intestinal inflammation by supplementing latent RelA dimers to the canonical NF-κB module.

Proc Natl Acad Sci U S A 2021 Jun;118(25)

Systems Immunology Laboratory, National Institute of Immunology, New Delhi 110067, India;

Aberrant inflammation, such as that associated with inflammatory bowel disease (IBD), is fueled by the inordinate activity of RelA/NF-κB factors. As such, the canonical NF-κB module mediates controlled nuclear activation of RelA dimers from the latent cytoplasmic complexes. What provokes pathological RelA activity in the colitogenic gut remains unclear. The noncanonical NF-κB pathway typically promotes immune organogenesis involving gene products. Because NF-κB pathways are intertwined, we asked whether noncanonical signaling aggravated inflammatory RelA activity. Our investigation revealed frequent engagement of the noncanonical pathway in human IBD. In a mouse model of experimental colitis, we established that -mediated regulations escalated the RelA-driven proinflammatory gene response in intestinal epithelial cells, exacerbating the infiltration of inflammatory cells and colon pathologies. Our mechanistic studies clarified that cell-autonomous signaling supplemented latent NF-κB dimers, leading to a hyperactive canonical RelA response in the inflamed colon. In sum, the regulation of latent NF-κB dimers appears to link noncanonical signaling to RelA-driven inflammatory pathologies and may provide for therapeutic targets.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2024828118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8237674PMC
June 2021

Doxycycline host-directed therapy in human pulmonary tuberculosis.

J Clin Invest 2021 Aug;131(15)

Infectious Diseases Translational Research Programme, Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

BACKGROUNDMatrix metalloproteinases (MMPs) are key regulators of tissue destruction in tuberculosis (TB) and may be targets for host-directed therapy. We conducted a phase II double-blind, randomized, controlled trial investigating doxycycline, a licensed broad-spectrum MMP inhibitor, in patients with pulmonary TB.METHODSThirty patients with pulmonary TB were enrolled within 7 days of initiating anti-TB treatment and randomly assigned to receive either 100 mg doxycycline or placebo twice a day for 14 days, in addition to standard care.RESULTSWhole blood RNA-sequencing demonstrated that doxycycline accelerated restoration of dysregulated gene expression in TB towards normality, rapidly down-regulating type I and II interferon and innate immune response genes, and up-regulating B-cell modules relative to placebo. The effects persisted for 6 weeks after doxycycline discontinuation, concurrent with suppressed plasma MMP-1. Doxycycline significantly reduced sputum MMP-1, -8, -9, -12 and -13, suppressed type I collagen and elastin destruction, reduced pulmonary cavity volume without altering sputum mycobacterial loads, and was safe.CONCLUSIONAdjunctive doxycycline with standard anti-TB treatment suppressed pathological MMPs in PTB patients. Larger studies on adjunctive doxycycline to limit TB immunopathology are merited.TRIAL REGISTRATIONClinicalTrials.gov NCT02774993.FUNDINGSingapore National Medical Research Council (NMRC/CNIG/1120/2014, NMRC/Seedfunding/0010/2014, NMRC/CISSP/2015/009a); the Singapore Infectious Diseases Initiative (SIDI/2013/013); National University Health System (PFFR-28 January 14, NUHSRO/2014/039/BSL3-SeedFunding/Jul/01); the Singapore Immunology Network Immunomonitoring platform (BMRC/IAF/311006, H16/99/b0/011, NRF2017_SISFP09); an ExxonMobil Research Fellowship, NUHS Clinician Scientist Program (NMRC/TA/0042/2015, CSAINV17nov014); the UK Medical Research Council (MR/P023754/1, MR/N006631/1); a NUS Postdoctoral Fellowship (NUHSRO/2017/073/PDF/03); The Royal Society Challenge Grant (CHG\R1\170084); the Sir Henry Dale Fellowship, Wellcome Trust (109377/Z/15/Z); and A*STAR.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/JCI141895DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8321570PMC
August 2021

CD27CD38 plasmablasts are activated B cells of mixed origin with distinct function.

iScience 2021 May 29;24(5):102482. Epub 2021 Apr 29.

Singapore Immunology Network, Agency for Science, Technology and Research, Singapore 138648, Singapore.

Clinically important broadly reactive B cells evolve during multiple infections, with B cells re-activated after secondary infection differing from B cells activated after a primary infection. Here we studied CD27CD38 plasmablasts from patients with a primary or secondary dengue virus infection. Three transcriptionally and functionally distinct clusters were identified. The largest cluster 0/1 was plasma cell-related, with cells coding for serotype cross-reactive antibodies of the IgG1 isotype, consistent with memory B cell activation during an extrafollicular response. Cells in clusters 2 and 3 expressed low levels of antibody genes and high levels of genes associated with oxidative phosphorylation, EIF2 pathway, and mitochondrial dysfunction. Clusters 2 and 3 showed a transcriptional footprint of T cell help, in line with activation from naive B cells or memory B cells. Our results contribute to the understanding of the parallel B cell activation events that occur in humans after natural primary and secondary infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.isci.2021.102482DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8169951PMC
May 2021

High-fat diet induces a predisposition to follicular hyperkeratosis and neutrophilic folliculitis in mice.

J Allergy Clin Immunol 2021 Mar 10. Epub 2021 Mar 10.

Singapore Immunology Network, Agency for Science, Technology and Research, Biopolis, Singapore; Skin Research Institute of Singapore, Agency for Science, Technology and Research, Biopolis, Singapore; Department of Dermatology, Kyoto University Graduate School of Medicine, Kyoto, Japan. Electronic address:

Background: Neutrophilic folliculitis is an inflammatory condition of hair follicles. In some neutrophilic folliculitis, such as in patients with acne and hidradenitis suppurativa, follicular hyperkeratosis is also observed. Neutrophilic folliculitis is often induced and/or exacerbated by a high-fat diet (HFD). However, the molecular mechanisms by which an HFD affects neutrophilic folliculitis are not fully understood.

Objective: Our aim was to elucidate how an HFD promotes the development of neutrophilic folliculitis.

Methods: Mice were fed an HFD, and their skin was subjected to histologic, RNA sequencing, and imaging mass spectrometry analyses. To examine the effect of an HFD on neutrophil accumulation around the hair follicles, phorbol 12-myristate 13-acetate (PMA) was used as an irritant to the skin.

Results: Histologic analysis revealed follicular hyperkeratosis in the skin of HFD-fed mice. RNA sequencing analysis showed that genes related to keratinization, especially in upper hair follicular keratinocytes, were significantly upregulated in HFD-fed mice. Application of PMA to the skin induced neutrophilic folliculitis in HFD-fed mice but not in mice fed a normal diet. Accumulation of neutrophils in the skin and around hair follicles was dependent on CXCR2 signaling, and CXCL1 (a CXCR2 ligand) was produced mainly by hair follicular keratinocytes. Imaging mass spectrometry analysis revealed an increase in fatty acids in the skin of HFD-fed mice. Application of these fatty acids to the skin induced follicular hyperkeratosis and caused PMA-induced neutrophilic folliculitis even in mice fed a normal diet.

Conclusion: An HFD can facilitate the development of neutrophilic folliculitis with the induction of hyperkeratosis of hair follicles and increased neutrophil infiltration around the hair follicles via CXCR2 signaling.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jaci.2021.02.032DOI Listing
March 2021

Role of VapBC12 Toxin-Antitoxin Locus in Cholesterol-Induced Mycobacterial Persistence.

mSystems 2020 Dec 15;5(6). Epub 2020 Dec 15.

Mycobacterial Pathogenesis Laboratory, Translational Health Science and Technology Institute, Faridabad, Haryana, India

The worldwide increase in the frequency of multidrug-resistant and extensively drug-resistant cases of tuberculosis is mainly due to therapeutic noncompliance associated with a lengthy treatment regimen. Depending on the drug susceptibility profile, the treatment duration can extend from 6 months to 2 years. This protracted regimen is attributed to a supposedly nonreplicating and metabolically inert subset of the population, called "persisters." The mechanism underlying stochastic generation and enrichment of persisters is not fully known. We have previously reported that the utilization of host cholesterol is essential for mycobacterial persistence. In this study, we have demonstrated that cholesterol-induced activation of a RNase toxin (VapC12) inhibits translation by targeting proT tRNA in This results in cholesterol-specific growth modulation that increases the frequency of generation of the persisters in a heterogeneous population. Also, a null mutant strain of this toxin (Δ) demonstrated an enhanced growth phenotype in a guinea pig model of infection, depicting its role in disease persistence. Thus, we have identified a novel strategy through which cholesterol-specific activation of a toxin-antitoxin module in enhances persister formation during infection. The current findings provide an opportunity to target persisters, a new paradigm facilitating tuberculosis drug development. The current TB treatment regimen involves a combination of drugs administered for an extended duration that could last for 6 months to 2 years. This could lead to noncompliance and the emergence of newer drug resistance strains. It is widely perceived that the major culprits are the so-called nonreplicating and metabolically inactive "persister" bacteria. The importance of cholesterol utilization during the persistence stage of infection and its potential role in the generation of persisters is very intriguing. We explored the mechanism involved in the cholesterol-mediated generation of persisters in mycobacteria. In this study, we have identified a toxin-antitoxin (TA) system essential for the generation of persisters during infection. This study verified that strain devoid of the VapBC12 TA system failed to persist and showed a hypervirulent phenotype in a guinea pig infection model. Our studies indicate that the VapBC12 TA system acts as a molecular switch regulating persister generation during infection. VapBC12 TA system as a drug target offers opportunities to develop shorter and more effective treatment regimens against tuberculosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mSystems.00855-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7771538PMC
December 2020

Metformin enhances anti-mycobacterial responses by educating CD8+ T-cell immunometabolic circuits.

Nat Commun 2020 10 16;11(1):5225. Epub 2020 Oct 16.

Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A*STAR), Singapore, 138648, Singapore.

Patients with type 2 diabetes (T2D) have a lower risk of Mycobacterium tuberculosis infection, progression from infection to tuberculosis (TB) disease, TB morality and TB recurrence, when being treated with metformin. However, a detailed mechanistic understanding of these protective effects is lacking. Here, we use mass cytometry to show that metformin treatment expands a population of memory-like antigen-inexperienced CD8CXCR3 T cells in naive mice, and in healthy individuals and patients with T2D. Metformin-educated CD8 T cells have increased (i) mitochondrial mass, oxidative phosphorylation, and fatty acid oxidation; (ii) survival capacity; and (iii) anti-mycobacterial properties. CD8 T cells from Cxcr3 mice do not exhibit this metformin-mediated metabolic programming. In BCG-vaccinated mice and guinea pigs, metformin enhances immunogenicity and protective efficacy against M. tuberculosis challenge. Collectively, these results demonstrate an important function of CD8 T cells in metformin-derived host metabolic-fitness towards M. tuberculosis infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-19095-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7567856PMC
October 2020

Macrophage polarisation associated with atherosclerosis differentially affects their capacity to handle lipids.

Atherosclerosis 2020 07 10;305:10-18. Epub 2020 Jun 10.

Department of Infection, Immunity & Cardiovascular Disease, University of Sheffield, Beech Hill Road, Sheffield, S10 2RX, UK. Electronic address:

Background And Aims: Lipid-rich foam cell macrophages drive atherosclerosis via several mechanisms, including inflammation, lipid uptake, lipid deposition and plaque vulnerability. The atheroma environment shapes macrophage function and phenotype; anti-inflammatory macrophages improve plaque stability while pro-inflammatory macrophages promote rupture. Current evidence suggests a variety of macrophage phenotypes occur in atherosclerotic plaques with local lipids, cytokines, oxidised phospholipids and pathogenic stimuli altering their phenotype. In this study, we addressed differential functioning of macrophage phenotypes via a systematic analysis of in vitro polarised, human monocyte-derived macrophage phenotypes, focussing on molecular events that regulate foam-cell formation.

Methods: We examined transcriptomes, protein levels and functionally determined lipid handling and foam cell formation capacity in macrophages polarised with IFNγ+LPS, IL-4, IL-10, oxPAPC and CXCL4.

Results: RNA sequencing of differentially polarised macrophages revealed distinct gene expression changes, with enrichment in atherosclerosis and lipid-associated pathways. Analysis of lipid processing activity showed IL-4 and IL-10 macrophages have higher lipid uptake and foam cell formation activities, while inflammatory and oxPAPC macrophages displayed lower foam cell formation. Inflammatory macrophages showed low lipid uptake, while higher lipid uptake in oxPAPC macrophages was matched by increased lipid efflux capacity.

Conclusions: Atherosclerosis-associated macrophage polarisation dramatically affects lipid handling capacity underpinned by major transcriptomic changes and altered protein levels in lipid-handling gene expression. This leads to phenotype-specific differences in LDL uptake, cellular cholesterol levels and cholesterol efflux, informing how the plaque environment influences atherosclerosis progression by influencing macrophage phenotypes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.atherosclerosis.2020.05.003DOI Listing
July 2020

Immunological history governs human stem cell memory CD4 heterogeneity via the Wnt signaling pathway.

Nat Commun 2020 02 10;11(1):821. Epub 2020 Feb 10.

Singapore Immunology Network (SIgN), Agency for Science Technology and Research (A*STAR), Immunos Building, 8A Biomedical Grove, Biopolis, Republic of Singapore.

The diversity of the naïve T cell repertoire drives the replenishment potential and capacity of memory T cells to respond to immune challenges. Attrition of the immune system is associated with an increased prevalence of pathologies in aged individuals, but whether stem cell memory T lymphocytes (T) contribute to such attrition is still unclear. Using single cells RNA sequencing and high-dimensional flow cytometry, we demonstrate that T heterogeneity results from differential engagement of Wnt signaling. In humans, aging is associated with the coupled loss of Wnt/β-catenin signature in CD4 T and systemic increase in the levels of Dickkopf-related protein 1, a natural inhibitor of the Wnt/β-catenin pathway. Functional assays support recent thymic emigrants as the precursors of CD4 T. Our data thus hint that reversing T defects by metabolic targeting of the Wnt/β-catenin pathway may be a viable approach to restore and preserve immune homeostasis in the context of immunological history.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-14442-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7010798PMC
February 2020

Single-Cell Analysis of Human Mononuclear Phagocytes Reveals Subset-Defining Markers and Identifies Circulating Inflammatory Dendritic Cells.

Immunity 2019 09 29;51(3):573-589.e8. Epub 2019 Aug 29.

Singapore Immunology Network, A(∗)STAR, 8A Biomedical Grove, Immunos Building, Singapore 138648, Singapore; Translational Immunology Institute, Singhealth/Duke-NUS Academic Medical Centre, the Academia, 20 College Road, Discovery Tower Level 8, Singapore 169856, Singapore. Electronic address:

Human mononuclear phagocytes comprise phenotypically and functionally overlapping subsets of dendritic cells (DCs) and monocytes, but the extent of their heterogeneity and distinct markers for subset identification remains elusive. By integrating high-dimensional single-cell protein and RNA expression data, we identified distinct markers to delineate monocytes from conventional DC2 (cDC2s). Using CD88 and CD89 for monocytes and HLA-DQ and FcεRIα for cDC2s allowed for their specific identification in blood and tissues. We also showed that cDC2s could be subdivided into phenotypically and functionally distinct subsets based on CD5, CD163, and CD14 expression, including a distinct subset of circulating inflammatory CD5CD163CD14 cells related to previously defined DC3s. These inflammatory DC3s were expanded in systemic lupus erythematosus patients and correlated with disease activity. These findings further unravel the heterogeneity of DC subpopulations in health and disease and may pave the way for the identification of specific DC subset-targeting therapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.immuni.2019.08.008DOI Listing
September 2019

RNA Sequencing of H3N2 Influenza Virus-Infected Human Nasal Epithelial Cells from Multiple Subjects Reveals Molecular Pathways Associated with Tissue Injury and Complications.

Cells 2019 08 27;8(9). Epub 2019 Aug 27.

Department of Otolaryngology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore.

The human nasal epithelium is the primary site of exposure to influenza virus, the initiator of host responses to influenza and the resultant pathologies. Influenza virus may cause serious respiratory infection resulting in major complications, as well as severe impairment of the airways. Here, we elucidated the global transcriptomic changes during H3N2 infection of human nasal epithelial cells from multiple individuals. Using RNA sequencing, we characterized the differentially-expressed genes and pathways associated with changes occurring at the nasal epithelium following infection. We used in vitro differentiated human nasal epithelial cell culture model derived from seven different donors who had no concurrent history of viral infections. Statistical analysis highlighted strong transcriptomic signatures significantly associated with 24 and 48 h after infection, but not at the earlier 8-h time point. In particular, we found that the influenza infection induced in the nasal epithelium early and altered responses in interferon gamma signaling, B-cell signaling, apoptosis, necrosis, smooth muscle proliferation, and metabolic alterations. These molecular events initiated at the infected nasal epithelium may potentially adversely impact the airway, and thus the genes we identified could serve as potential diagnostic biomarkers or therapeutic targets for influenza infection and associated disease management.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/cells8090986DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6770044PMC
August 2019

Whole-transcriptome sequencing reveals heightened inflammation and defective host defence responses in chronic rhinosinusitis with nasal polyps.

Eur Respir J 2019 11 14;54(5). Epub 2019 Nov 14.

Dept of Otolaryngology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.

Introduction: The pathways underlying chronic rhinosinusitis with nasal polyps (CRSwNP) are unclear. We conducted genome-wide gene expression analysis to determine pathways and candidate gene sets associated with CRSwNP.

Methods: We performed whole-transcriptome RNA sequencing on 42 polyp (CRSwNP-NP) and 33 paired nonpolyp inferior turbinate (CRSwNP-IT) tissues from patients with CRSwNP and 28 inferior turbinate samples from non-CRS controls (CS-IT). We analysed the differentially expressed genes (DEGs) and the gene sets that were enriched in functional pathways.

Results: Principal component-informed analysis revealed cilium function and immune regulation as the two main Gene Ontology (GO) categories differentiating CRSwNP patients from controls. We detected 6182 and 1592 DEGs between CRSwNP-NP CS-IT and between CRSwNP-NP CRSwNP-IT tissues, respectively. Atopy status did not have a major impact on gene expression in various tissues. GO analysis on these DEGs implicated extracellular matrix (ECM) disassembly, -glycan processing, angiogenesis and host viral response in CRSwNP pathogenesis. Ingenuity Pathway Analysis identified significant enrichment of type 1 interferon signalling and axonal guidance canonical pathways, angiogenesis, and collagen and fibrotic changes in CRSwNP (CRSwNP-NP and CRSwNP-IT) tissues compared with CS-IT. Finally, gene set enrichment analysis implicated sets of genes co-regulated in processes associated with inflammatory response and aberrant cell differentiation in polyp formation.

Conclusions: Gene signatures involved in defective host defences (including cilia dysfunction and immune dysregulation), inflammation and abnormal metabolism of ECM are implicated in CRSwNP. Functional validation of these gene expression patterns will open opportunities for CRSwNP therapeutic interventions such as biologics and immunomodulators.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1183/13993003.00732-2019DOI Listing
November 2019

mTORC2/Akt activation in adipocytes is required for adipose tissue inflammation in tuberculosis.

EBioMedicine 2019 Jul 4;45:314-327. Epub 2019 Jul 4.

Department of Medicine, University of Massachusetts Medical School, Worcester, MA, USA. Electronic address:

Background: Mycobacterium tuberculosis has co-evolved with the human host, adapting to exploit the immune system for persistence and transmission. While immunity to tuberculosis (TB) has been intensively studied in the lung and lymphoid system, little is known about the participation of adipose tissues and non-immune cells in the host-pathogen interaction during this systemic disease.

Methods: C57BL/6J mice were aerosol infected with M. tuberculosis Erdman and presence of the bacteria and the fitness of the white and brown adipose tissues, liver and skeletal muscle were studied compared to uninfected mice.

Findings: M. tuberculosis infection in mice stimulated immune cell infiltration in visceral, and brown adipose tissue. Despite the absence of detectable bacterial dissemination to fat tissues, adipocytes produced localized pro-inflammatory signals that disrupted adipocyte lipid metabolism, resulting in adipocyte hypertrophy. Paradoxically, this resulted in increased insulin sensitivity and systemic glucose tolerance. Adipose tissue inflammation and enhanced glucose tolerance also developed in obese mice after aerosol M. tuberculosis infection. We found that infection induced adipose tissue Akt signaling, while inhibition of the Akt activator mTORC2 in adipocytes reversed TB-associated adipose tissue inflammation and cell hypertrophy.

Interpretation: Our study reveals a systemic response to aerosol M. tuberculosis infection that regulates adipose tissue lipid homeostasis through mTORC2/Akt signaling in adipocytes. Adipose tissue inflammation in TB is not simply a passive infiltration with leukocytes but requires the mechanistic participation of adipocyte signals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ebiom.2019.06.052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6642333PMC
July 2019

Plasmacytoid dendritic cells develop from Ly6D lymphoid progenitors distinct from the myeloid lineage.

Nat Immunol 2019 07 18;20(7):852-864. Epub 2019 Jun 18.

Singapore Immunology Network, Agency for Science Technology and Research, Singapore, Singapore.

Dendritic cells (DC) are currently classified as conventional DCs (cDCs) and plasmacytoid DCs (pDCs). Through a combination of single-cell transcriptomic analysis, mass cytometry, in vivo fate mapping and in vitro clonal assays, here we show that, at the single-cell level, the priming of mouse hematopoietic progenitor cells toward the pDC lineage occurs at the common lymphoid progenitor stage, indicative of early divergence of the pDC and cDC lineages. We found the transcriptional signature of a pDC precursor stage, defined here, in the IL-7Rα common lymphoid progenitor population and identified Ly6D, IL-7Rα, CD81 and CD2 as key markers of pDC differentiation, which distinguish pDC precursors from cDC precursors. In conclusion, pDCs developed in the bone marrow from a Ly6DCD2 lymphoid progenitor cell and differentiated independently of the myeloid cDC lineage.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41590-019-0420-3DOI Listing
July 2019

RIG-I Activation by a Designer Short RNA Ligand Protects Human Immune Cells against Dengue Virus Infection without Causing Cytotoxicity.

J Virol 2019 07 28;93(14). Epub 2019 Jun 28.

Singapore Immunology Network, Agency for Science, Technology and Research, Singapore

Virus-derived double-stranded RNA (dsRNA) molecules containing a triphosphate group at the 5' end are natural ligands of retinoic acid-inducible gene I (RIG-I). The cellular pathways and proteins induced by RIG-I are an essential part of the innate immune response against viral infections. Starting from a previously published RNA scaffold (3p10L), we characterized an optimized small dsRNA hairpin (called 3p10LG9, 25 nucleotides [nt] in length) as a highly efficient RIG-I activator. Dengue virus (DENV) infection in cell lines and primary human skin cells could be prevented and restricted through 3p10LG9-mediated activation of RIG-I. This antiviral effect was RIG-I and interferon signal dependent. The effect was temporary and was reversed above a saturating concentration of RIG-I ligand. This finding revealed an effective feedback loop that controls potentially damaging inflammatory effects of the RIG-I response, at least in immune cells. Our results show that the small RIG-I activator 3p10LG9 can confer short-term protection against DENV and can be further explored as an antiviral treatment in humans. Short hairpin RNA ligands that activate RIG-I induce antiviral responses in infected cells and prevent or control viral infections. Here, we characterized a new short hairpin RNA molecule with high efficacy in antiviral gene activation and showed that this molecule is able to control dengue virus infection. We demonstrate how structural modifications of minimal RNA ligands can lead to increased potency and a wider window of RIG-I-activating concentrations before regulatory mechanisms kick in at high concentrations. We also show that minimal RNA ligands induce an effective antiviral response in human skin dendritic cells and macrophages, which are the target cells of initial infection after the mosquito releases virus into the skin. Using short hairpin RNA as RIG-I ligands could therefore be explored as antiviral therapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00102-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6600207PMC
July 2019

A Subset of Type I Conventional Dendritic Cells Controls Cutaneous Bacterial Infections through VEGFα-Mediated Recruitment of Neutrophils.

Immunity 2019 04 27;50(4):1069-1083.e8. Epub 2019 Mar 27.

Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A(∗)STAR), 8A Biomedical Grove, Biopolis, Singapore 138648, Singapore; Skin Research Institute of Singapore (SRIS), Agency for Science, Technology and Research (A(∗)STAR), 11 Mandalay Rd., Singapore 308232, Singapore. Electronic address:

Skin conventional dendritic cells (cDCs) exist as two distinct subsets, cDC1s and cDC2s, which maintain the balance of immunity to pathogens and tolerance to self and microbiota. Here, we examined the roles of dermal cDC1s and cDC2s during bacterial infection, notably Propionibacterium acnes (P. acnes). cDC1s, but not cDC2s, regulated the magnitude of the immune response to P. acnes in the murine dermis by controlling neutrophil recruitment to the inflamed site and survival and function therein. Single-cell mRNA sequencing revealed that this regulation relied on secretion of the cytokine vascular endothelial growth factor α (VEGF-α) by a minor subset of activated EpCAMCD59Ly-6D cDC1s. Neutrophil recruitment by dermal cDC1s was also observed during S. aureus, bacillus Calmette-Guérin (BCG), or E. coli infection, as well as in a model of bacterial insult in human skin. Thus, skin cDC1s are essential regulators of the innate response in cutaneous immunity and have roles beyond classical antigen presentation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.immuni.2019.03.001DOI Listing
April 2019

Two distinct interstitial macrophage populations coexist across tissues in specific subtissular niches.

Science 2019 03;363(6432)

Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore.

Macrophages are a heterogeneous cell population involved in tissue homeostasis, inflammation, and various pathologies. Although the major tissue-resident macrophage populations have been extensively studied, interstitial macrophages (IMs) residing within the tissue parenchyma remain poorly defined. Here we studied IMs from murine lung, fat, heart, and dermis. We identified two independent IM subpopulations that are conserved across tissues: Lyve1MHCIICX3CR1 (Lyve1MHCII) and Lyve1MHCIICX3CR1 (Lyve1MHCII) monocyte-derived IMs, with distinct gene expression profiles, phenotypes, functions, and localizations. Using a new mouse model of inducible macrophage depletion ( ), we found that the absence of Lyve1MHCII IMs exacerbated experimental lung fibrosis. Thus, we demonstrate that two independent populations of IMs coexist across tissues and exhibit conserved niche-dependent functional programming.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.aau0964DOI Listing
March 2019

Corrigendum: A Single-Cell Sequencing Guide for Immunologists.

Front Immunol 2019 26;10:278. Epub 2019 Feb 26.

Singapore Immunology Network, Agency for Science, Technology and Research, Singapore, Singapore.

[This corrects the article DOI: 10.3389/fimmu.2018.02425.].
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2019.00278DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6400142PMC
February 2019

Mapping of γ/δ T cells reveals Vδ2+ T cells resistance to senescence.

EBioMedicine 2019 Jan 7;39:44-58. Epub 2018 Dec 7.

Singapore Immunology Network (SIgN), Agency for Science Technology and Research (A*STAR), Immunos Building, Singapore 138648, Singapore; School of Biological Sciences, Nanyang Technological University, Singapore, Singapore; Department of Microbiology, National University of Singapore, Singapore, Singapore; Department of Biology, Faculty of Science, University Tunis El Manar, Tunis, Tunisia. Electronic address:

Background: Immune adaptation with aging is a major of health outcomes. Studies in humans have mainly focus on αβ T cells while γδ T cells have been neglected despite their role in immunosurveillance. We investigated the impact of aging on γδ T cell subsets phenotypes, functions, senescence and their molecular response to stress.

Methods: Peripheral blood of young and old donors in Singapore have been used to assess the phenotype, functional capacity, proliferation capacity and gene expression of the various γδ T cell subsets. Peripheral blood mononuclear cells from apheresis cones and young donors have been used to characterize the telomere length, epigenetics profile and DNA damage response of the various γδ T cell subsets phenotype.

Findings: Our data shows that peripheral Vδ2+ phenotype, functional capacity (cytokines, cytotoxicity, proliferation) and gene expression profile are specific when compared against all other αβ and γδ T cells in aging. Hallmarks of senescence including telomere length, epigenetic profile and DNA damage response of Vδ2+ also differs against all other αβ and γδ T cells.

Interpretation: Our results highlight the differential impact of lifelong stress on γδ T cells subsets, and highlight possible mechanisms that enable Vδ2+ to be resistant to cellular aging. The new findings reinforce the concept that Vδ2+ have an "innate-like" behavior and are more resilient to the environment as compared to "adaptive-like" Vδ1+ T cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ebiom.2018.11.053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6354624PMC
January 2019

A Single-Cell Sequencing Guide for Immunologists.

Front Immunol 2018 23;9:2425. Epub 2018 Oct 23.

Singapore Immunology Network, Agency for Science, Technology and Research, Singapore, Singapore.

In recent years there has been a rapid increase in the use of single-cell sequencing (scRNA-seq) approaches in the field of immunology. With the wide range of technologies available, it is becoming harder for users to select the best scRNA-seq protocol/platform to address their biological questions of interest. Here, we compared the advantages and limitations of four commonly used scRNA-seq platforms in order to clarify their suitability for different experimental applications. We also address how the datasets generated by different scRNA-seq platforms can be integrated, and how to identify unknown populations of single cells using unbiased bioinformatics methods.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2018.02425DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6205970PMC
September 2019

Hyaluronan Receptor LYVE-1-Expressing Macrophages Maintain Arterial Tone through Hyaluronan-Mediated Regulation of Smooth Muscle Cell Collagen.

Immunity 2018 08 24;49(2):326-341.e7. Epub 2018 Jul 24.

School of Biological Sciences, Nanyang Technological University, Nanyang, Singapore 637551, Singapore; Institute of Molecular and Cell Biology, A(∗)STAR, Singapore 138673, Singapore; KK Women's and Children Hospital, Singapore 229899, Singapore.

The maintenance of appropriate arterial tone is critically important for normal physiological arterial function. However, the cellular and molecular mechanisms remain poorly defined. Here, we have shown that in the mouse aorta, resident macrophages prevented arterial stiffness and collagen deposition in the steady state. Using phenotyping, transcriptional profiling, and targeted deletion of Csf1r, we have demonstrated that these macrophages-which are a feature of blood vessels invested with smooth muscle cells (SMCs) in both mouse and human tissues-expressed the hyaluronan (HA) receptor LYVE-l. Furthermore, we have shown they possessed the unique ability to modulate collagen expression in SMCs by matrix metalloproteinase MMP-9-dependent proteolysis through engagement of LYVE-1 with the HA pericellular matrix of SMCs. Our study has unveiled a hitherto unknown homeostatic contribution of arterial LYVE-1 macrophages through the control of collagen production by SMCs and has identified a function of LYVE-1 in leukocytes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.immuni.2018.06.008DOI Listing
August 2018

Publisher Correction: IgG1 memory B cells keep the memory of IgE responses.

Nat Commun 2018 03 1;9(1):968. Epub 2018 Mar 1.

Singapore Immunology Network (SIgN), 8A Biomedical Grove, Singapore, 138648, Singapore.

The originally published version of this Article contained errors in Fig. 4 that were introduced during the production process. In panel c, the two uppermost labels 'IgE spleen' and 'IgE BM' incorrectly read 'IgG1 spleen' and 'IgE1 BM', respectively. These errors have now been corrected in both the PDF and HTML versions of the Article.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-018-03364-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5832859PMC
March 2018

The tumour microenvironment creates a niche for the self-renewal of tumour-promoting macrophages in colon adenoma.

Nat Commun 2018 02 8;9(1):582. Epub 2018 Feb 8.

School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551, Singapore.

Circulating CCR2 monocytes are crucial for maintaining the adult tissue-resident F4/80MHCII macrophage pool in the intestinal lamina propria. Here we show that a subpopulation of CCR2-independent F4/80MHCII macrophages, which are the most abundant F4/80 cells in neonates, gradually decline in number in adulthood; these macrophages likely represent the fetal contribution to F4/80 cells. In colon adenomas of Apc mice, F4/80MHCII macrophages are not only preserved, but become the dominant subpopulation among tumour-resident macrophages during tumour progression. Furthermore, these pro-tumoural F4/80MHCII and F4/80MHCII macrophages can self-renew in the tumour and maintain their numbers mostly independent from bone marrow contribution. Analyses of colon adenomas indicate that CSF1 may be a key facilitator of macrophage self-renewal. In summary, the tumour microenvironment creates an isolated niche for tissue-resident macrophages that favours macrophage survival and self-renewal.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-018-02834-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5805689PMC
February 2018

Microbiome Influences Prenatal and Adult Microglia in a Sex-Specific Manner.

Cell 2018 01 21;172(3):500-516.e16. Epub 2017 Dec 21.

Institut de Biologie de l'Ecole normale supérieure (IBENS), Ecole Normale Supérieure, CNRS, INSERM, PSL Research University, 75005 Paris, France. Electronic address:

Microglia are embryonically seeded macrophages that contribute to brain development, homeostasis, and pathologies. It is thus essential to decipher how microglial properties are temporally regulated by intrinsic and extrinsic factors, such as sexual identity and the microbiome. Here, we found that microglia undergo differentiation phases, discernable by transcriptomic signatures and chromatin accessibility landscapes, which can diverge in adult males and females. Remarkably, the absence of microbiome in germ-free mice had a time and sexually dimorphic impact both prenatally and postnatally: microglia were more profoundly perturbed in male embryos and female adults. Antibiotic treatment of adult mice triggered sexually biased microglial responses revealing both acute and long-term effects of microbiota depletion. Finally, human fetal microglia exhibited significant overlap with the murine transcriptomic signature. Our study shows that microglia respond to environmental challenges in a sex- and time-dependent manner from prenatal stages, with major implications for our understanding of microglial contributions to health and disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cell.2017.11.042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5786503PMC
January 2018

IgG1 memory B cells keep the memory of IgE responses.

Nat Commun 2017 09 21;8(1):641. Epub 2017 Sep 21.

Singapore Immunology Network (SIgN), 8A Biomedical Grove, Singapore, 138648, Singapore.

The unique differentiation of IgE cells suggests unconventional mechanisms of IgE memory. IgE germinal centre cells are transient, most IgE cells are plasma cells, and high affinity IgE is produced by the switching of IgG1 cells to IgE. Here we investigate the function of subsets of IgG1 memory B cells in IgE production and find that two subsets of IgG1 memory B cells, CD80CD73 and CD80CD73, contribute distinctively to the repertoires of high affinity pathogenic IgE and low affinity non-pathogenic IgE. Furthermore, repertoire analysis indicates that high affinity IgE and IgG1 plasma cells differentiate from rare CD80CD73 high affinity memory clones without undergoing further mutagenesis. By identifying the cellular origin of high affinity IgE and the clonal selection of high affinity memory B cells into the plasma cell fate, our findings provide fundamental insights into the pathogenesis of allergies, and on the mechanisms of antibody production in memory B cell responses.IgE is an important mediator of protective immunity as well as allergic reaction, but how high affinity IgE antibodies are produced in memory responses is not clear. Here the authors show that IgE can be generated via class-switch recombination in IgG1 memory B cells without additional somatic hypermutation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-017-00723-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5608722PMC
September 2017

The Transcriptional Response of to Weak Organic Acids, Carbon Source, and Inactivation Unveils a Role for in Mediating the Fungistatic Effect of Acetic Acid.

G3 (Bethesda) 2017 11 6;7(11):3597-3604. Epub 2017 Nov 6.

Singapore Immunology Network (SIgN), Agency for Science, Technology and Research (A*STAR), Singapore 138648, Singapore

is a resident fungus of the human intestinal microflora. Commonly isolated at low abundance in healthy people, outcompetes local microbiota during candidiasis episodes. Under normal conditions, members of the human gastrointestinal (GI) microbiota were shown to keep colonization under control. By releasing weak organic acids (WOAs), bacteria are able to moderate yeast growth. This mechanism displays a synergistic effect with the absence of glucose in medium of culture, which underlines the complex interactions that faces in its natural environment. Inactivation of the transcriptional regulator in results in a lack of sensitivity to this synergistic outcome. To decipher transcriptional responses to glucose, WOAs, and the role of , we performed RNA sequencing (RNA-seq) on four biological replicates exposed to combinations of these three parameters. We were able to characterize the (i) glucose response, (ii) response to acetic and butyric acid, (iii) regulation of , and (iv) genes responsible for WOA resistance. We identified a group of six genes linked to WOA sensitivity in a glucose--dependent manner and inactivated one of these genes, the putative glucose transporter , in a SC5314 wild-type background. As expected, the mutant displayed a partial complementation to WOA resistance in the absence of glucose. This result points toward a mechanism of WOA sensitivity in involving membrane transporters, which could be exploited to control yeast colonization in human body niches.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.117.300238DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5677169PMC
November 2017
-->