9 research outputs found
The role of lipid laden macrophages in airways disease
Asthma is a chronic inflammatory disease of the airway and is often associated with Gastro-Oesophageal Reflux (GOR). Lipid laden macrophages (LLM) in the airway have previously been reported indicative of aspiration secondary to GOR. We hypothesised that lipid droplets from undigested or partially digested food may be aspirated into the airway and accumulate in scavenging macrophages thus generating an activated population that interact with other immune cells to induce an asthma-like state of disease.Using a lipid laden alveolar macrophage index, we collected 29 bronchoalveolarlavage (BAL) samples from patients with a variety of respiratory disorders, todetermine the use of this test in GOR diagnosis. We investigated the mode of uptake of unmodified lipid and other alternative sources of lipid. We attempted to generate polarised activated macrophages and to determine the activation status of LLM. We also characterised a new macrophage cell line, and compared these to PMA differentiated THP-1 macrophages.We have shown that the LLAMI is incapable of diagnosing GOR. We showed that macrophages are capable of direct dietary lipid uptake, independently of actin or CD36. Lipid accumulation also results from the phagocytic clearance of apoptotic epithelial cells, which may account for the lack of LLAMI specificity for GOR. Our attempts to generate and distinguish polarised macrophages were unsuccessful and as a result, we were not able to determine the activation status of LLM. We demonstrated heterogeneity among primary macrophage cells isolated from BAL fluid collected from patients with varying pathologies. Particularly interesting was the finding that macrophages isolated from a patient with GOR showed distinctly greater CD36 expression. Daisy cells are novel cells which appear to be macrophage-like morphologically, functionally and phenotypically. Daisy cells have potential utility with the ability to spontaneously replicate and differentiate an advantage over THP-1 cells which require PMA stimulation
Characterisation of a New Human Alveolar Macrophage-Like Cell Line (Daisy)
Purpose: There is currently no true macrophage cell line and in vitro experiments requiring these cells currently require mitogenic stimulation of a macrophage precursor cell line (THP-1) or ex vivo maturation of circulating primary monocytes. In this study, we characterise a human macrophage cell line, derived from THP-1 cells, and compare its phenotype to the THP-1 cells. Methods: THP-1 cells with and without mitogenic stimulation were compared to the newly derived macrophage-like cell line (Daisy) using microscopy, flow cytometry, phagocytosis assays, antigen binding assays and gene microarrays. Results: We show that the cell line grows predominantly in an adherent monolayer. A panel of antibodies were chosen to investigate the cell surface phenotype of these cells using flow cytometry. Daisy cells expressed more CD11c, CD80, CD163, CD169 and CD206, but less CD14 and CD11b compared with mitogen-stimulated THP-1 cells. Unlike stimulated THP-1 cells which were barely able to bind immune complexes, Daisy cells showed large amounts of immune complex binding. Finally, although not statistically significant, the phagocytic ability of Daisy cells was greater than mitogen-stimulated THP-1 cells, suggesting that the cell line is more similar to mature macrophages. Conclusions: The observed phenotype suggests that Daisy cells are a good model of human macrophages with a phenotype similar to human alveolar macrophages
GWAS meta-analysis of intrahepatic cholestasis of pregnancy implicates multiple hepatic genes and regulatory elements
Intrahepatic cholestasis of pregnancy (ICP) is a pregnancy-specific liver disorder affecting 0.5–2% of pregnancies. The majority of cases present in the third trimester with pruritus, elevated serum bile acids and abnormal serum liver tests. ICP is associated with an increased risk of adverse outcomes, including spontaneous preterm birth and stillbirth. Whilst rare mutations affecting hepatobiliary transporters contribute to the aetiology of ICP, the role of common genetic variation in ICP has not been systematically characterised to date. Here, we perform genome-wide association studies (GWAS) and meta-analyses for ICP across three studies including 1138 cases and 153,642 controls. Eleven loci achieve genome-wide significance and have been further investigated and fine-mapped using functional genomics approaches. Our results pinpoint common sequence variation in liver-enriched genes and liver-specific cis-regulatory elements as contributing mechanisms to ICP susceptibility
Increased inducible heat shock protein 72 expression associated with PBMC isolated from patients with haematological tumours
Background. Heat shock protein 72 (Hsp72) is a highly inducible stress protein and molecular chaperone. Cancers have been shown to be associated with increased Hsp72 expression within the tumour itself and this may lead to resistance to apoptosis. Methods. Peripheral blood mononuclear cells (PBMC) were isolated from patients diagnosed with chronic lymphocytic leukaemia (CLL) (n 27) and chronic myelomonocytic leukaemia (CMML) (n 16) and Hsp72 expression was characterized on both the cell surface and intracellularly by flow cytometry. To allow for comparison PBMC from breast cancer patients (n 25) and healthy volunteers (n 19) were included. Results. Both lymphocytes and monocytes from CLL and CMML patients showed high levels of total Hsp72 expression (46 fold increase) in comparison to breast cancer and healthy subjects. The majority of Hsp72 in these tumours was determined to be cell-surface expressed (6493% of cell total Hsp72). Conclusions. A correlation was observed between lymphocyte and monocyte total Hsp72 expression (p < 0.001) suggesting a common stress response pathway may exist in these blood cells and there are stress conditions present within the circulation. Hsp72 expression was not found to be related to white blood cell count. © 2012 Informa Healthcare
Whole-genome sequencing reveals host factors underlying critical COVID-19
Altres ajuts: Department of Health and Social Care (DHSC); Illumina; LifeArc; Medical Research Council (MRC); UKRI; Sepsis Research (the Fiona Elizabeth Agnew Trust); the Intensive Care Society, Wellcome Trust Senior Research Fellowship (223164/Z/21/Z); BBSRC Institute Program Support Grant to the Roslin Institute (BBS/E/D/20002172, BBS/E/D/10002070, BBS/E/D/30002275); UKRI grants (MC_PC_20004, MC_PC_19025, MC_PC_1905, MRNO2995X/1); UK Research and Innovation (MC_PC_20029); the Wellcome PhD training fellowship for clinicians (204979/Z/16/Z); the Edinburgh Clinical Academic Track (ECAT) programme; the National Institute for Health Research, the Wellcome Trust; the MRC; Cancer Research UK; the DHSC; NHS England; the Smilow family; the National Center for Advancing Translational Sciences of the National Institutes of Health (CTSA award number UL1TR001878); the Perelman School of Medicine at the University of Pennsylvania; National Institute on Aging (NIA U01AG009740); the National Institute on Aging (RC2 AG036495, RC4 AG039029); the Common Fund of the Office of the Director of the National Institutes of Health; NCI; NHGRI; NHLBI; NIDA; NIMH; NINDS.Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care or hospitalization after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes-including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)-in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease
Whole-genome sequencing of patients with rare diseases in a national health system
Most patients with rare diseases do not receive a molecular diagnosis and the aetiological variants and causative genes for more than half such disorders remain to be discovered1. Here we used whole-genome sequencing (WGS) in a national health system to streamline diagnosis and to discover unknown aetiological variants in the coding and non-coding regions of the genome. We generated WGS data for 13,037 participants, of whom 9,802 had a rare disease, and provided a genetic diagnosis to 1,138 of the 7,065 extensively phenotyped participants. We identified 95 Mendelian associations between genes and rare diseases, of which 11 have been discovered since 2015 and at least 79 are confirmed to be aetiological. By generating WGS data of UK Biobank participants2, we found that rare alleles can explain the presence of some individuals in the tails of a quantitative trait for red blood cells. Finally, we identified four novel non-coding variants that cause disease through the disruption of transcription of ARPC1B, GATA1, LRBA and MPL. Our study demonstrates a synergy by using WGS for diagnosis and aetiological discovery in routine healthcare
Publisher Correction: Whole-genome sequencing of a sporadic primary immunodeficiency cohort (Nature, (2020), 583, 7814, (90-95), 10.1038/s41586-020-2265-1)
An amendment to this paper has been published and can be accessed via a link at the top of the paper
Genetic determinants of risk in pulmonary arterial hypertension: international genome-wide association studies and meta-analysis.
BACKGROUND: Rare genetic variants cause pulmonary arterial hypertension, but the contribution of common genetic variation to disease risk and natural history is poorly characterised. We tested for genome-wide association for pulmonary arterial hypertension in large international cohorts and assessed the contribution of associated regions to outcomes. METHODS: We did two separate genome-wide association studies (GWAS) and a meta-analysis of pulmonary arterial hypertension. These GWAS used data from four international case-control studies across 11 744 individuals with European ancestry (including 2085 patients). One GWAS used genotypes from 5895 whole-genome sequences and the other GWAS used genotyping array data from an additional 5849 individuals. Cross-validation of loci reaching genome-wide significance was sought by meta-analysis. Conditional analysis corrected for the most significant variants at each locus was used to resolve signals for multiple associations. We functionally annotated associated variants and tested associations with duration of survival. All-cause mortality was the primary endpoint in survival analyses. FINDINGS: A locus near SOX17 (rs10103692, odds ratio 1·80 [95% CI 1·55-2·08], p=5·13 × 10-15) and a second locus in HLA-DPA1 and HLA-DPB1 (collectively referred to as HLA-DPA1/DPB1 here; rs2856830, 1·56 [1·42-1·71], p=7·65 × 10-20) within the class II MHC region were associated with pulmonary arterial hypertension. The SOX17 locus had two independent signals associated with pulmonary arterial hypertension (rs13266183, 1·36 [1·25-1·48], p=1·69 × 10-12; and rs10103692). Functional and epigenomic data indicate that the risk variants near SOX17 alter gene regulation via an enhancer active in endothelial cells. Pulmonary arterial hypertension risk variants determined haplotype-specific enhancer activity, and CRISPR-mediated inhibition of the enhancer reduced SOX17 expression. The HLA-DPA1/DPB1 rs2856830 genotype was strongly associated with survival. Median survival from diagnosis in patients with pulmonary arterial hypertension with the C/C homozygous genotype was double (13·50 years [95% CI 12·07 to >13·50]) that of those with the T/T genotype (6·97 years [6·02-8·05]), despite similar baseline disease severity. INTERPRETATION: This is the first study to report that common genetic variation at loci in an enhancer near SOX17 and in HLA-DPA1/DPB1 is associated with pulmonary arterial hypertension. Impairment of SOX17 function might be more common in pulmonary arterial hypertension than suggested by rare mutations in SOX17. Further studies are needed to confirm the association between HLA typing or rs2856830 genotyping and survival, and to determine whether HLA typing or rs2856830 genotyping improves risk stratification in clinical practice or trials. FUNDING: UK NIHR, BHF, UK MRC, Dinosaur Trust, NIH/NHLBI, ERS, EMBO, Wellcome Trust, EU, AHA, ACClinPharm, Netherlands CVRI, Dutch Heart Foundation, Dutch Federation of UMC, Netherlands OHRD and RNAS, German DFG, German BMBF, APH Paris, INSERM, Université Paris-Sud, and French ANR