Immunology Highlights
Cutting-edge Irish research
We recently ran a competition among undergraduate students to summarise cutting-edge Irish research papers for a general audience. Read on to discover the winning submissions.
​The TNFSF12/TWEAK Modulates Colonic Inflammatory Fibroblast Differentiation and Promotes Fibroblast-Monocyte Interactions
summarised by Sarah Balfe
Authors: Carlos Matellan, Ciarán Kennedy, Miren Itxaso Santiago-Vela, Johanna Hochegger, Méabh B Ní Chathail, Amanda Wu, Christopher Shannon, Helen M Roche, Seema S Aceves, Catherine Godson, Mario C Manresa
Inflammatory Bowel Disease (IBD) includes both Crohn’s Disease and Ulcerative Colitis and affects millions of people around the world. Unfortunately, there is no cure and the treatments available only work in certain people. Recently, scientists have discovered that fibroblasts, a type of cell involved in maintaining a healthy intestine, becomes inflamed in patients with IBD. This inflamed fibroblast is believed to contribute to the symptoms of the disease. However, we do not know what causes these cells to become inflamed. This study investigated how a certain inflammatory molecule, known as TWEAK (TNF-like Weak Inducer of Apoptosis), can cause intestinal fibroblasts to become inflamed. They also analysed how these inflamed fibroblasts can interact with and activate a type of immune cell called a monocyte. Monocytes are very important cells in the immune system that help to protect the body against infection and illness. However, in certain diseases such as IBD, they can become overactivated causing unwanted inflammation.
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Researchers grew intestinal fibroblasts in the lab and treated them with TWEAK. The results revealed that TWEAK causes intestinal fibroblasts to become inflamed and produce inflammatory molecules. These inflamed fibroblasts closely matched a certain type of fibroblast identified in samples from patients suffering with ulcerative colitis.
The scientists also combined these inflamed fibroblasts with monocytes to test the effects of their interaction. They found that the interaction caused monocytes to become activated. This increased activation resulted in the release of inflammatory signals associated with amplified inflammation. In addition, the study also showed that blocking a particular signalling pathway involved in inflammation reduced the ability of the inflamed fibroblasts to activate monocytes.
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Overall, the results highlight TWEAK as a potential target for the treatment of IBD. It also provides us with more information on the broader role of inflamed fibroblasts in inflammatory diseases such as arthritis and cancer, promoting research into new treatment strategies in this area.
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​Respiratory Syncytial Virus NS1 Inhibits Anti-Viral Interferon-α-Induced JAK/STAT Signaling, by Limiting the Nuclear Translocation of STAT1
summarised by Nour Bouzidi
Authors: Claudia Efstathiou, Yamei Zhang, Shubhangi Kandwal, Darren Fayne, Eleanor J. Molloy, Nigel J. Stevenson
Respiratory syncytial virus (RSV) is the leading cause of respiratory infection in infants, partly through disrupting one of the body’s immune responses called the interferon (IFN) response. The IFN response is one of the earliest responses against viral infection, transmitting signals into the cell to activate proteins known as STAT proteins. STAT proteins then travel to the cell’s nucleus and activate antiviral genes which in turn produce proteins to combat the virus.
This study aimed to identify the pathways in which RSV interferes with immune signalling in a lung cell line, notably via disruption of this signalling pathway. RSV produces 11 proteins in total, including 2 named NS1 and NS2, which are known to disrupt the IFN response.
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To study how RSV-NS1 and RSV-NS2 affects the activity of antiviral genes, cells were infected with either NS1 or NS2 along with another ‘light-producing’ gene. This gene produces light when active, with brighter light indicating greater activation of the gene of interest. The cells were exposed to IFN-É‘ for 18 hours which served to activate the IFN response and its antiviral target genes. Cells infected with RSV-NS1 showed reduced gene activation, as indicated by the lower light signals. Additionally, the proteins produced by these genes were also reduced 24h and 48h after NS1 infection, suggesting that the NS1 protein prevents full activation of antiviral genes.
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The IFN response occurs in multiple steps, one of which includes the activation of STAT proteins. To evaluate the specific step at which STAT signalling is impaired, levels of different STATs were measured, such as pSTAT1. Levels of pSTAT1 increased as expected indicating that the virus does not prevent STAT protein activation and must impact a step found further down the signalling cascade.
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STAT protein transport was then investigated as a potential mechanism behind the signalling disruption. After activation, STAT proteins need to travel to the nucleus, which is the cell region where gene activation occurs. By looking at levels of STAT1 inside to outside the nucleus, NS1 was found to reduce transport of STAT1 into the nucleus compared to cells without NS1. This showed that transport of STAT1 is impaired by NS1 which may explain the reduced antiviral gene activation.
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STAT1 is transported by a protein called KPNA1. KPNA1 levels were found to be unaffected by NS1 infection. However, when looking at how KPNA1 attaches to STAT1, the virus markedly reduced this interaction. Without this attachment, the KPNA1 protein is unable to transport STAT1 to activate antiviral genes, a process essential to combat RSV infection.
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Overall, this study discovered a potential way in which the RSV virus decreases IFN signalling by preventing activation of antiviral genes and disrupting the transport of proteins involved in their activation. These results uncover a potential therapeutic target to fight off RSV infection, which would block one of the mechanisms RSV uses to decrease the immune response.​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​
Characterisation of the Pro-Inflammatory Cytokine Signature in Severe COVID-19
summarised by Mairead Clancy
Authors: Heike C. Hawerkamp, Adam H. Dyer, Neha D. Patil, Matt McElheron, Niamh O'Dowd, Laura O'Doherty, Aisling Ui Mhaonaigh, Angel M. George, Aisling M. O'Halloran, Conor Reddy, Rose Anne Kenny, Mark A. Little, Ignacio Martin-Loeches, Colm Bergin, Sean P. Kennelly, Seamas C. Donnelly, Nollaig M. Bourke, Aideen Long, Jacklyn Sui, Derek G. Doherty, Niall Conlon, Cliona Ni Cheallaigh, Padraic G. Fallon
The SARS-CoV-2 (COVID-19) virus spread rapidly in 2020 resulting in millions of deaths. Hawerkamp et al. (2023) [1] analysed the plasma of 118 unvaccinated patients with COVID-19 to identify a combination of proinflammatory cytokine expression that varies with respect to disease severity. Inflammation is the immune system’s response to a pathogen, releasing proinflammatory cytokines to create a toxic environment killing the pathogen [2]. Cytokines are proteins that regulate the immune and inflammatory response [3]. Severe COVID-19 is associated with a hyper-inflammatory phenotype, in which proinflammatory molecules are excessively produced causing dysregulation of the immune response, tissue damage and/or death.
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The symptoms of COVID-19 vary dramatically from asymptomatic to severe respiratory distress, pneumonia and mortality. Investigating the dysregulation of the immune response with respect to disease severity increases knowledge of pathogenesis and allows for development of personalised treatment of COVID-19 increasing survival rates. Patients in the study were classified into severity categories – mild, moderate, severe and dead. Of the 118 participants, 26 had mild COVID-19, 58 had moderate and 34 had severe. Patients with severe COVID-19 required oxygen therapy via non-invasive ventilation or mechanical ventilation. 20 proinflammatory plasma biomarkers (cytokines) were analysed. An ELISA assay was conducted to detect and quantify soluble proteins (cytokines). Biomarker concentrations were measured and further analysed to identify variability between disease severities.
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Overall, patients with COVID-19 showed increases in 16 of the 20 proinflammatory biomarkers and a decrease in the anti-inflammatory cytokine, TGF-B. Decreased TGF-B results in dysregulation of cell proliferation and antibody expression. The expression of 11 biomarkers varied across disease severity. 8 biomarkers differed between severe and moderate disease states and 9 biomarkers between severe and mild. There were strong correlations seen in six pairs of cytokines which reflect the hyper-inflammatory phenotype of severe COVID-19. The cytokines IL-2, IL-6, IP-10, TNF-a, IL-10, IL-33, C-GSF and IL-1B were shown to be the greatest contributors to hyper-inflammation and are independently associated with severe COVID-19.
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Previous reports have shown the IP-10 cytokine is highly correlated with disease severity and mortality [4]. IP-10 causes systemic inflammation by lowering the proliferation of T-cells. Increased G-CSF increases production of granulocytes - immune cells that release cytotoxic molecules. High G-CSF has been associated with disease severity, increased oxygen usage and mortality in COVID-19 [5]. IL-33 drives production of Th2 associated cytokines to defend against extracellular pathogens and is associated with COVID-19 severity [6]. Previous research reordered increased cytokine levels as a “Cytokine Storm Syndrome” [7] – the rapid release of cytokines into the bloodstream promoting hyperinflammation, resulting in systemic tissue damage, multi-organ failure or death [8]. This study suggests the cytokine response in severe COVID-19 is more moderate and distinct than CSS but more unique than other respiratory diseases.
This study is not without limitations. Almost all patients were treated with dexamethasone, which is known to have potent effects on the immune response to COVID-19. Dexamethasone use may have skewed results. Further research should investigate how the identified cytokine signature contributes to disease progression and severity.
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References
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Hawerkamp et al., Front Immunol 2024. DOI: 10.3389/fimmu.2023.1170012 PUBMED
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AU, A., An overview of inflammation: mechanism and consequences. . Frontiers Biology, 2011. 6(4): p. 274.
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Zhang, J.-M. and J. An, Cytokines, inflammation, and pain. International anesthesiology clinics, 2007. 45(2): p. 27-37.
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Yang, Y., et al., Plasma IP-10 and MCP-3 levels are highly associated with disease severity and predict the progression of COVID-19. Journal of Allergy and Clinical Immunology, 2020. 146(1): p. 119-127. e4.
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Jøntvedt Jørgensen, M., et al., Increased interleukin-6 and macrophage chemoattractant protein-1 are associated with respiratory failure in COVID-19. Scientific reports, 2020. 10(1): p. 21697.
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Makaremi, S., et al., The role of IL-1 family of cytokines and receptors in pathogenesis of COVID-19. Inflammation Research, 2022. 71(7): p. 923-947.
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Furci, F., et al., IL-33 and the cytokine storm in COVID-19: from a potential immunological relationship towards precision medicine. International Journal of Molecular Sciences, 2022. 23(23): p. 14532.
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Fajgenbaum, D.C. and C.H. June, Cytokine storm. New England Journal of Medicine, 2020. 383(23): p. 2255-2273.
Bystander Activation of Bordetella pertussis-Induced Nasal Tissue-Resident Memory (TRM) CD4 T Cells Confers Heterologous Immunity to Klebsiella pneumoniae
summarised by Laura Lane
Authors: Lucy M. Curham, Jenny M. Mannion, Clíodhna M. Daly, Mieszko M. Wilk, Lisa Borkner, Stephen J. Lalor, Rachel M. McLoughlin, Kingston H.G. Mills
TRM cells are a special kind of memory T cells that stay in the body after you get over an infection or after vaccination, ready to quickly attack the same pathogen if it attacks again. These cells are crucial for long-lasting protection against whooping cough, which is caused by the bacteria Bordetella pertussis. Even though the whooping cough vaccine is widely used, the disease is making a comeback, partly because the newer acellular vaccines don't stop the bacteria from settling in the nose like the older whole-cell vaccines did.
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In this study, researchers used female C57BL/6 mice, kept in a very clean environment, to show that B. pertussis-specific CD4 TRM cells can be activated by completely different pathogens like Klebsiella pneumoniae. This happens through something called bystander activation, where certain immune system messengers—cytokines like IL-23, IL-1β, and IL-18—are released by immune cells responding to the other pathogens, not by direct contact with the pathogen itself. Once these TRM cells are activated, they produce another messenger called IL-17A, which recruits white blood cells called neutrophils and other immune cells to the site of infection to help fight off these secondary infections.
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The researchers used various methods to study this process: they used flow cytometry to look at different types of cells and what signals they were sending out, ELISA tests to measure the levels of cytokines in the cell cultures, and RT-PCR to check how much of each cytokine gene was being used by the cells. Their tests showed that TRM cells specific to B. pertussis could be activated by E. coli LPS a component of bacteria in the lab when dendritic cells (a type of immune cell) were also present. This activation led to the production of IL-17A but was specific to TRM cells found in the respiratory system, not those in the lymph nodes. Using a blocker for IL-12p40, a cytokine subunit, lowered the production of IL-17A and IFN-γ, suggesting that cytokines really are behind this activation. These respiratory TRM cells from mice that had recovered from infection also responded to heat killed K. pneumoniae and Staphylococcus aureus by releasing IL-17A, showing that various pathogens can trigger these TRM cells through cytokine signals.
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The study also discovered that just stimulating these cells with IL-23, IL-1β, or IL-18 was enough to get them to produce IL-17A, especially when helped by IL-7, which is known to help memory T cells survive. This shows that respiratory TRM cells can be directly activated by cytokines without needing to recognise the specific pathogen directly.
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In experiments with live mice, spraying LPS or heat killed K. pneumoniae into the noses of mice that had previously had B. pertussis boosted the number of IL-17A-producing TRM cells and brought more neutrophils into the nasal area, strengthening the local immune defence. Vaccinating mice through the nose with the old whole-cell whooping cough vaccine also successfully turned on B. pertussis-specific CD4 TRM cells, which protected against K. pneumoniae. This kind of immunity, fuelled by IL-17A and bringing in neutrophils and other immune cells, suggests that old-style whole-cell vaccines could offer broader protection, which is promising for future vaccine development.
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These findings emphasise how CD4 TRM cells that have been turned on by a Bordetella pertussis infection can be activated by different pathogens through bystander activation. This allows them to provide protection not just against the original pathogen but against other infections too, thanks to cytokines that activate them without needing to directly engage the pathogen. This capability of TRM cells could be key to creating vaccines that protect more broadly, especially against pathogens that have changed to avoid vaccine effects. This means that bystander activation of TRM cells might be an important consideration for designing next-generation vaccines targeting respiratory infections.​​​​​​