ORCID Profile
0000-0003-2878-476X
Current Organisations
University of Southampton
,
University of SouthamptonSouthampton
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Publisher: Frontiers Media SA
Date: 29-11-2017
Publisher: Springer Science and Business Media LLC
Date: 15-03-2018
Abstract: A major endeavor of systems biology is the construction of graphical and computational models of biological pathways as a means to better understand their structure and function. Here, we present a protocol for a biologist-friendly graphical modeling scheme that facilitates the construction of detailed network diagrams, summarizing the components of a biological pathway (such as proteins and biochemicals) and illustrating how they interact. These diagrams can then be used to simulate activity flow through a pathway, thereby modeling its dynamic behavior. The protocol is ided into four sections: (i) assembly of network diagrams using the modified Edinburgh Pathway Notation (mEPN) scheme and yEd network editing software with pathway information obtained from published literature and databases of molecular interaction data (ii) parameterization of the pathway model within yEd through the placement of 'tokens' on the basis of the known or imputed amount or activity of a component (iii) model testing through visualization and quantitative analysis of the movement of tokens through the pathway, using the network analysis tool Graphia Professional and (iv) optimization of model parameterization and experimentation. This is the first modeling approach that combines a sophisticated notation scheme for depicting biological events at the molecular level with a Petri net-based flow simulation algorithm and a powerful visualization engine with which to observe the dynamics of the system being modeled. Unlike many mathematical approaches to modeling pathways, it does not require the construction of a series of equations or rate constants for model parameterization. Depending on a model's complexity and the availability of information, its construction can take days to months, and, with refinement, possibly years. However, once assembled and parameterized, a simulation run, even on a large model, typically takes only seconds. Models constructed using this approach provide a means of knowledge management, information exchange and, through the computation simulation of their dynamic activity, generation and testing of hypotheses, as well as prediction of a system's behavior when perturbed.
Publisher: American Society for Clinical Investigation
Date: 02-08-2021
DOI: 10.1172/JCI141895
Publisher: Royal Society of Chemistry (RSC)
Date: 2021
DOI: 10.1039/D1LC00292A
Abstract: Pirouette coupling involves Dean flow for cell and reporter bead inertial ordering for efficient co-encapsulation, achieving a throughput of 1 million cells per hour, a 2.5% multiplet rate and a 70% cell capture efficiency.
Publisher: Elsevier BV
Date: 03-2021
Publisher: Oxford University Press (OUP)
Date: 24-11-2020
Abstract: Crohn’s disease [CD] arises through host-environment interaction. Abnormal gene expression results from disturbed pathway activation or response to bacteria. We aimed to determine activated pathways and driving cell types in paediatric CD. We employed contemporary targeted autoimmune RNA sequencing, in parallel to single-cell sequencing, to ileal tissue derived from paediatric CD and controls. Weighted gene co-expression network analysis [WGCNA] was performed and differentially expressed genes [DEGs] were determined. We integrated clinical data to determine co-expression modules associated with outcomes. In all, 27 treatment-naive CD [TN-CD], 26 established CD patients and 17 controls were included. WGCNA revealed a 31-gene signature characterising TN-CD patients, but not established CD, nor controls. The CSF3R gene is a hub within this module and is key in neutrophil expansion and differentiation. Antimicrobial genes, including S100A12 and the calprotectin subunit S100A9, were significantly upregulated in TN CD compared with controls [p = 2.61 x 10-15 and p = 9.13 x 10-14, respectively] and established CD [both p = 0.0055]. Gene-enrichment analysis confirmed upregulation of the IL17-, NOD- and Oncostatin-M-signalling pathways in TN-CD patients, identified in both WGCNA and DEG analyses. An upregulated gene signature was enriched for transcripts promoting Th17-cell differentiation and correlated with prolonged time to relapse [correlation-coefficient-0.36, p = 0.07]. Single-cell sequencing of TN-CD patients identified specialised epithelial cells driving differential expression of S100A9. Cell groups, determined by single-cell gene expression, demonstrated enrichment of IL17-signalling in monocytes and epithelial cells. Ileal tissue from treatment-naïve paediatric patients is significantly upregulated for genes driving IL17-, NOD- and Oncostatin-M-signalling. This signal is driven by a distinct subset of epithelial cells expressing antimicrobial gene transcripts.
Publisher: Cold Spring Harbor Laboratory
Date: 09-04-2021
DOI: 10.1101/2021.04.08.439026
Abstract: The future of single cell ersity screens involves ever-larger s le sizes, dictating the need for higher throughput methods with low analytical noise to accurately describe the nature of the cellular system. Current approaches are limited by the Poisson statistic, requiring dilute cell suspensions and associated losses in throughput. In this contribution, we apply Dean entrainment to both cell and bead inputs, defining different volume packets to effect efficient co-encapsulation. Volume ratio scaling was explored to identify optimal conditions. This enabled the co-encapsulation of single cells with reporter beads at rates of ~1 million cells/hour, while increasing assay signal-to-noise with cell multiplet rates of ~2.5% and capturing ~70% of cells. The method, called Pirouette-seq, extends our capacity to investigate biological systems. Pirouette-seq involves cell and reporter bead inertial ordering for efficient co-encapsulation, achieving a throughput of 1 million cells/hour, a 2.5% multiplet rate and a 70% cell capture efficiency.
Publisher: The American Association of Immunologists
Date: 15-07-2022
Abstract: NK cells are promising cellular therapeutics against hematological and solid malignancies. Immunogenetic studies have identified that various activating killer cell Ig-like receptors (KIRs) are associated with cancer outcomes. Specifically, KIR2DS2 has been associated with reduced incidence of relapse following transplant in hematological malignancies and improved outcomes in solid tumors, but the mechanism remains obscure. Therefore, we investigated how KIR2DS2 expression impacts NK cell function. Using a novel flow cytometry panel, we show that human NK cells with high KIR2DS2 expression have enhanced spontaneous activation against malignant B cell lines, liver cancer cell lines, and primary chronic lymphocytic leukemia cells. Surface expression of CD16 was increased on KIR2DS2high NK cells, and, accordingly, KIR2DS2high NK cells had increased activation against lymphoma cells coated with the clinically relevant anti-CD20 Abs rituximab and obinutuzumab. Bulk RNA sequencing revealed that KIR2DS2high NK cells have upregulation of NK-mediated cytotoxicity, translation, and FCGR gene pathways. We developed a novel single-cell RNA-sequencing technique to identify KIR2DS2+ NK cells, and this confirmed that KIR2DS2 is associated with enhanced NK cell–mediated cytotoxicity. This study provides evidence that KIR2DS2 marks a population of NK cells primed for anticancer activity and indicates that KIR2DS2 is an attractive target for NK-based therapeutic strategies.
Publisher: Frontiers Media SA
Date: 15-06-2021
DOI: 10.3389/FIMMU.2021.665312
Abstract: Langerhans cells (LCs) reside in the epidermis as a dense network of immune system sentinels, coordinating both immunogenic and tolerogenic immune responses. To determine molecular switches directing induction of LC immune activation, we performed mathematical modelling of gene regulatory networks identified by single cell RNA sequencing of LCs exposed to TNF-alpha, a key pro-inflammatory signal produced by the skin. Our approach delineated three programmes of LC phenotypic activation (immunogenic, tolerogenic or ambivalent), and confirmed that TNF-alpha enhanced LC immunogenic programming. Through regulon analysis followed by mutual information modelling, we identified IRF1 as the key transcription factor for the regulation of immunogenicity in LCs. Application of a mathematical toggle switch model, coupling IRF1 with tolerance-inducing transcription factors, determined the key set of transcription factors regulating the switch between tolerance and immunogenicity, and correctly predicted LC behaviour in LCs derived from different body sites. Our findings provide a mechanistic explanation of how combinatorial interactions between different transcription factors can coordinate specific transcriptional programmes in human LCs, interpreting the microenvironmental context of the local tissue microenvironments.
Publisher: Cold Spring Harbor Laboratory
Date: 15-05-2021
DOI: 10.1101/2021.05.12.21257086
Abstract: The worldwide pandemic caused by SARS-CoV-2 has claimed millions of lives and has had a profound effect on global life. Understanding the pathogenicity of the virus and the body’s response to infection is crucial in improving patient management, prognosis, and therapeutic strategies. To address this, we performed functional transcriptomic profiling to better understand the generic and specific effects of SARS-CoV-2 infection. Whole blood RNA sequencing was used to profile a well characterised cohort of patients hospitalised with COVID-19, during the first wave of the pandemic prior to the availability of approved COVID-19 treatments and who went on to survive or die of COVID-19, and patients hospitalised with influenza virus infection between 2017 and 2019. Clinical parameters between patient groups were compared, and several bioinformatic tools were used to assess differences in transcript abundances and cellular composition. The analyses revealed contrasting innate and adaptive immune programmes, with transcripts and cell subsets associated with the innate immune response elevated in patients with influenza, and those involved in the adaptive immune response elevated in patients with COVID-19. Topological analysis identified additional gene signatures that differentiated patients with COVID-19 from patients with influenza, including insulin resistance, mitochondrial oxidative stress and interferon signalling. An efficient adaptive immune response was furthermore associated with patient survival, while an inflammatory response predicted death in patients with COVID-19. A potential prognostic signature was found based on a selection of transcript abundances, associated with circulating immunoglobulins, nucleosome assembly, cytokine production and T cell activation, in the blood transcriptome of COVID-19 patients, upon admission to hospital, which can be used to stratify patients likely to survive or die. The results identified distinct immunological signatures between SARS-CoV-2 and influenza, prognostic of disease progression and indicative of different targeted therapies. The altered transcript abundances associated with COVID-19 survivors can be used to predict more severe outcomes in patients with COVID-19.
Publisher: Springer Science and Business Media LLC
Date: 16-01-2020
DOI: 10.1038/S41467-019-14125-X
Abstract: Langerhans cells (LC) can prime tolerogenic as well as immunogenic responses in skin, but the genomic states and transcription factors (TF) regulating these context-specific responses are unclear. Bulk and single-cell transcriptional profiling demonstrates that human migratory LCs are robustly programmed for MHC-I and MHC-II antigen presentation. Chromatin analysis reveals enrichment of ETS-IRF and AP1-IRF composite regulatory elements in antigen-presentation genes, coinciding with expression of the TFs, PU.1, IRF4 and BATF3 but not IRF8. Migration of LCs from the epidermis is accompanied by upregulation of IRF4, antigen processing components and co-stimulatory molecules. TNF stimulation augments LC cross-presentation while attenuating IRF4 expression. CRISPR-mediated editing reveals IRF4 to positively regulate the LC activation programme, but repress NF2EL2 and NF-kB pathway genes that promote responsiveness to oxidative stress and inflammatory cytokines. Thus, IRF4-dependent genomic programming of human migratory LCs appears to enable LC maturation while attenuating excessive inflammatory and immunogenic responses in the epidermis.
Publisher: Springer Science and Business Media LLC
Date: 19-05-2023
DOI: 10.1038/S41467-023-38588-1
Abstract: Regulation of cutaneous immunity is severely compromised in inflammatory skin disease. To investigate the molecular crosstalk underpinning tolerance versus inflammation in atopic dermatitis, we utilise a human in vivo allergen challenge study, exposing atopic dermatitis patients to house dust mite. Here we analyse transcriptional programmes at the population and single cell levels in parallel with immunophenotyping of cutaneous immunocytes revealed a distinct dichotomy in atopic dermatitis patient responsiveness to house dust mite challenge. Our study shows that reactivity to house dust mite was associated with high basal levels of TNF-expressing cutaneous Th17 T cells, and documents the presence of hub structures where Langerhans cells and T cells co-localised. Mechanistically, we identify expression of metallothioneins and transcriptional programmes encoding antioxidant defences across all skin cell types, that appear to protect against allergen-induced inflammation. Furthermore, single nucleotide polymorphisms in the MTIX gene are associated with patients who did not react to house dust mite, opening up possibilities for therapeutic interventions modulating metallothionein expression in atopic dermatitis.
Publisher: Frontiers Media SA
Date: 15-11-2019
Publisher: Cold Spring Harbor Laboratory
Date: 10-10-2019
DOI: 10.1101/800631
Abstract: Single-cell transcriptomics has sensitivity limits that restrict low abundance transcript identification, affects clustering and introduce artefact. Here, we describe Constellation DropSeq (C-DropSeq), a molecular transcriptome filter that delivers two orders of magnitude sensitivity gains by maximising read utility while reducing sequencing depth and costs. The simple and powerful method is broadly compatible with library preparation routines and was demonstrated by identifying and characterizing the activation of rare dendritic cell sub-populations.
Publisher: Oxford University Press (OUP)
Date: 20-09-2020
DOI: 10.1111/BJD.19431
Publisher: American Society for Clinical Investigation
Date: 21-07-2021
DOI: 10.1172/JCI148136
Publisher: Elsevier BV
Date: 11-2018
Publisher: Elsevier BV
Date: 05-2022
Location: United Kingdom of Great Britain and Northern Ireland
Location: United Kingdom of Great Britain and Northern Ireland
No related grants have been discovered for Marta Ewa Polak.