European Guest Session: Ireland

Thursday, 12.09.2024, 11:30 am

Chair: Barry Moran

Barry Moran

Barry Moran

Trinity Biomedical Sciences Institute, Trinity College Dublin

Complementary cell analysis technologies to elucidate skin disease – No Kuddelmuddel!

Background: Cell analysis technologies can add important information to under-recognized and understudied skin diseases such as chronic spontaneous urticaria (CSU) and hidradenitis suppurativa (HS). CSU is a skin condition characterized by recurrent itchy hives that has a major impact upon quality of life, while HS is a chronic skin disease with painful, oozing lesions and abscesses. Treatment to date for both conditions is inadequate in many patients.
Objectives: Characterising the skin and blood immune cells, their transcriptome and secretome, in these diseases will lead to a better understanding of their pathogenesis, providing a rationale for new or stratified therapeutic strategies.
Methods: In these studies we employed flow and imaging cytometry, microscopy, single cell RNA sequencing, multiplex assays and ELISA to characterise immune cells isolated from patients and healthy donors.
Results: This presentation will discuss the findings of both studies. The CSU study identified an increase in mast cell precursors in patient blood, with high expression of these cells a strong predictor of better clinical response. The HS study identified distinct immune cell subsets and an enhancement of specific genes and pathways associated with Th17 cells, IL-17, IL-1β, and the NLRP3 inflammasome (an inflammatory protein complex) in patient skin. Inhibiting this NLRP3 inflammasome in HS patient skin explants significantly reduced inflammatory markers, providing a rationale for a potential new therapy.
Conclusion: Complementary single cell analysis technologies can provide invaluable insights into skin disease pathology and treatments.

Bioscetch

Dr Barry Moran leads the Flow Cytometry Facility at Trinity College Dublin, supporting over 130 multi-disciplinary research groups. He is also teaching and research active, running his department’s PhD training modules alongside his primary research interest of immune dysfunction in skin disease. He is a Fellow and Executive Committee member of the Royal Microscopical Society (RMS) and the Core Technologies for Life Sciences society (CTLS) and, with Alfonso Blanco, he leads the Cytometry Society of Ireland.

David Finlay

David Finlay

Trinity Biomedical Sciences Institute, Trinity College Dublin, Ireland

Using Click-chemistry to study nutrient uptake in single cells

It is now clear that some of the metabolic discoveries made using in vitro cultured immune cells do not hold true for the metabolic features of these cells in vivo.  The in vivo environment is not homogenous and there are differences in metabolic conditions at the sites of diseases including cancer and infection. The next great frontier for the immunometabolism field is to measure single immune cell metabolism in vivo and at the site of disease. Working towards this goal, we have developed  a new technology to accurately measure nutrient uptake, the first rate limiting step for cellular metabolic pathways, using a biorthogonal chemistry-based (CLICK-chemistry) approach. This involves using a minimally modified nutrient, containing a small CLICK handle, that is taken up by the cell via the normal physiological route. A fluorophore is attached to the CLICK-nutrient after transport has occurred, when inside cell. Using multiple CLICK configurations it is possible to perform 3 sequential CLICK reactions in each immune cell. In this way we have simultaneously measured amino acid uptake, fatty acid uptake and rates of protein synthesis, providing multiple dimensions of metabolic flux analysis in individual cells. When combined with other single cell metabolic stains and assays it is now possible to generate an ex vivo metabolic footprint of each individual immune cell taken from complex immune populations from diverse tissues and disease sites. This approach, what we have called 4D+MetaFlux, is revealing new insights into the heterogeneity of immune cells metabolism within and between tissues.

Biosketch
David Finlay is Professor in Immunometabolism a the Trinity Biomedical Sciences Institute, Trinity College Dublin. He was awarded a bachelor’s degree in Biochemistry in 2001, completed PhD (2006) and postdoctoral training at the University of Dundee in Scotland and was elected to fellowship of Trinity College Dublin (2018). His research expertise lies at the interface of immunology and cellular metabolism, focusing on Natural Killer cells and Dendritic cells. His team is developing novel single cell measures of in vivo immunometabolism using novel biorthogonal chemistry approaches. Dr Finlay’s research has been supported by awards including an  SFI Career Development Award (2014), ERC-CoG (2017) and ERC-PoC (2023), and an Irish Research Council Advanced Laureate award (2023).

Ella Fouhy

Ella Fouhy

School of Biomolecular and Biomedical Science, University College Dublin

Uncovering the early circulating environment that precedes a diagnosis of Pre-eclampisa

Pre-eclampsia (PE) is a serious complication affecting 1 in 12 pregnancies, defined by new-onset hypertension and proteinuria after 20 weeks of gestation. Early detection of PE is key for appropriate clinical decision-making throughout pregnancy. Extracellular vesicles (EVs) are important circulating messengers in the blood regulating a myriad of biological and pathological processes. EVs are highly implicated in pro-inflammatory diseases and their levels are significantly increased following a clinical diagnosis of PE. We hypothesise that EVs circulating in the first trimester are altered in those that subsequently develop PE and may be an early indication of impending disease. Ethical approval was gained to access the Biological Resource Bank in the Coombe Women and Infants University Hospital Dublin for plasma samples taken in early pregnancy (11-14+6 weeks’ gestation) for healthy women, and women who had a confirmed diagnosis of preterm PE or term PE, after 20 weeks of gestation. Multicolour flow cytometry was employed to quantify circulating platelet-derived EVs in plasma and significant alterations in concentration and cell of origin was observed between the groups. Strikingly, we identified distinct circulating EV populations that could differentiate between future incidence of either preterm or term PE. These FCM findings provide novel insights into the underlying aetiology of PE and its distinct associated phenotypes.

Biosketch
Ella Fouhy is a final year graduate PhD student under the supervision of Prof Patricia Maguire in the Conway SPHERE research Group in University College Dublin. ConwaySPHERE is an interdisciplinary research team co-directed by Prof Maguire and Prof Fionnuala Ní Áinle, which brings together clinical, academic and scientific collaborators nationally and internationally, harnessing unique tools to diagnose and understand a host of diseases, including thrombotic disorders and inflammatory diseases. One of their flagship projects is AIPREMie, where they are developing an AI-powered clinical decision support tool for risk stratification in the pregnancy complication pre-eclampsia.