Reduced interleukin-4 receptor alpha expression on CD8(+) T cells correlates with higher quality anti-viral immunity

Published January 31, 2013With the hope of understanding how interleukin (IL)-4 and IL-13 modulated quality of anti-viral CD8⁺ T cells, we evaluated the expression of receptors for these cytokines following a range of viral infections (e.g. pox viruses and influenza virus). Results clearly indicated that unlike other IL-4/IL-13 receptor subunits, IL-4 receptor α (IL-4Rα) was significantly down-regulated on anti-viral CD8⁺ T cells in a cognate antigen dependent manner. The infection of gene knockout mice and wild-type (WT) mice with vaccinia virus (VV) or VV expressing IL-4 confirmed that IL-4, IL-13 and signal transducer and activator of transcription 6 (STAT6) were required to increase IL-4Rα expression on CD8⁺ T cells, but not interferon (IFN)-γ. STAT6 dependent elevation of IL-4Rα expression on CD8⁺ T cells was a feature of poor quality anti-viral CD8⁺ T cell immunity as measured by the production of IFN-γ and tumor necrosis factor α (TNF-α) in response to VV antigen stimulation in vitro. We propose that down-regulation of IL-4Rα, but not the other IL-4/IL-13 receptor subunits, is a mechanism by which CD8⁺ T cells reduce responsiveness to IL-4 and IL-13. This can improve the quality of anti-viral CD8⁺ T cell immunity. Our findings have important implications in understanding anti-viral CD8⁺ T cell immunity and designing effective vaccines against chronic viral infections.Danushka K. Wijesundara, David C. Tscharke, Ronald J. Jackson, Charani Ranasingh

Bacillus cereus non-haemolytic enterotoxin activates the NLRP3 inflammasome

Inflammasomes are important for host defence against pathogens and homeostasis with commensal microbes. Here, we show non-haemolytic enterotoxin (NHE) from the neglected human foodborne pathogen Bacillus cereus is an activator of the NLRP3 inflammasome and pyroptosis. NHE is a non-redundant toxin to haemolysin BL (HBL) despite having a similar mechanism of action. Via a putative transmembrane region, subunit C of NHE initiates binding to the plasma membrane, leading to the recruitment of subunit B and subunit A, thus forming a tripartite lytic pore that is permissive to efflux of potassium. NHE mediates killing of cells from multiple lineages and hosts, highlighting a versatile functional repertoire in different host species. These data indicate that NHE and HBL operate synergistically to induce inflammation and show that multiple virulence factors from the same pathogen with conserved function and mechanism of action can be exploited for sensing by a single inflammasome

Expression and Cellular Immunogenicity of a Transgenic Antigen Driven by Endogenous Poxviral Early Promoters at Their Authentic Loci in MVA

CD8+ T cell responses to vaccinia virus are directed almost exclusively against early gene products. The attenuated strain modified vaccinia virus Ankara (MVA) is under evaluation in clinical trials of new vaccines designed to elicit cellular immune responses against pathogens including Plasmodium spp., M. tuberculosis and HIV-1. All of these recombinant MVAs (rMVA) utilize the well-established method of linking the gene of interest to a cloned poxviral promoter prior to insertion into the viral genome at a suitable locus by homologous recombination in infected cells. Using BAC recombineering, we show that potent early promoters that drive expression of non-functional or non-essential MVA open reading frames (ORFs) can be harnessed for immunogenic expression of recombinant antigen. Precise replacement of the MVA orthologs of C11R, F11L, A44L and B8R with a model antigen positioned to use the same translation initiation codon allowed early transgene expression similar to or slightly greater than that achieved by the commonly-used p7.5 or short synthetic promoters. The frequency of antigen-specific CD8+ T cells induced in mice by single shot or adenovirus-prime, rMVA-boost vaccination were similarly equal or marginally enhanced using endogenous promoters at their authentic genomic loci compared to the traditional constructs. The enhancement in immunogenicity observed using the C11R or F11L promoters compared with p7.5 was similar to that obtained with the mH5 promoter compared with p7.5. Furthermore, the growth rates of the viruses were unimpaired and the insertions were genetically stable. Insertion of a transgenic ORF in place of a viral ORF by BAC recombineering can thus provide not only a potent promoter, but also, concomitantly, a suitable insertion site, potentially facilitating development of MVA vaccines expressing multiple recombinant antigens

The antigen-specific CD8+ T cell repertoire in unimmunized mice includes memory phenotype cells bearing markers of homeostatic expansion

Memory T cells exhibit superior responses to pathogens and tumors compared with their naive counterparts. Memory is typically generated via an immune response to a foreign antigen, but functional memory T cells can also be produced from naive cells by homeostatic mechanisms. Using a recently developed method, we studied CD8 T cells, which are specific for model (ovalbumin) and viral (HSV, vaccinia) antigens, in unimmunized mice and found a subpopulation bearing markers of memory cells. Based on their phenotypic markers and by their presence in germ-free mice, these preexisting memory-like CD44hi CD8 T cells are likely to arise via physiological homeostatic proliferation rather than a response to environmental microbes. These antigen-inexperienced memory phenotype CD8 T cells display several functions that distinguish them from their CD44lo counterparts, including a rapid initiation of proliferation after T cell stimulation and rapid IFN-γ production after exposure to proinflammatory cytokines. Collectively, these data indicate that the unprimed antigen-specific CD8 T cell repertoire contains antigen-inexperienced cells that display phenotypic and functional traits of memory cells

Statement in Support of: "Virology under the Microscope-a Call for Rational Discourse"

Letter to the Editor. Published 25 April 2023Peter Speck, Jason Mackenzie, Rowena A. Bull, Barry Slobedman, Heidi Drummer, Johanna Fraser, Lara Herrero, Karla Helbig, Sarah Londrigan, Gregory Moseley, Natalie Prow, Grant Hansman, Robert Edwards, Chantelle Ahlenstiel, Allison Abendroth, David Tscharke, Jody Hobson-Peters, Robson Kriiger-Loterio, Rhys Parry, Glenn Marsh, Emma Harding, David A. Jacques, Matthew J. Gartner, Wen Shi Lee, Julie McAuley, Paola Vaz, Frank Sainsbury, Michelle D. Tate, Jane Sinclair, Allison Imrie, Stephen Rawlinson, Andrew Harman, Jillian M. Carr, Ebony A. Monson, Merilyn Hibma, Timothy J.Mahony, Thomas Tu, Robert J. Center, Lok Bahadur Shrestha, Robyn Hall, Morgyn Warner, Vernon Ward, Danielle E. Anderson, Nicholas S. Eyre, Natalie E. Netzler, Alison J. Peel, Peter Revill, Michael Beard, Alistair R. Legione, Alexandra J. Spencer, Adi Idris, Jade Forwood, Subir Sarker, Damian F. J. Purcell, Nathan Bartlett, Joshua M. Deerain, Bruce J. Brew, Sassan Asgari, Helen Farrell, Alexander Khromykh, Daniel Enosi Tuipulotu, David Anderson, Sevim Mese, Yaman Tayyar, Kathryn Edenborough, Jasim Muhammad Uddin, Abrar Hussain, Connor J. I. Daymond, Jacinta Agius, Karyn N. Johnson, Paniz Shirmast, Mahdi Abedinzadeshahri, Robin MacDiarmid, Caroline L. Ashley, Jay Laws, Lucy L. Furfaro, Thomas D. Burton, Stephen M. R. Johnson, Zahra Telikani, Mary Petrone, Justin A. Roby, Carolyn Samer, Andreas Suhrbier, April Van Der Kamp, Anthony Cunningham, Celeste Donato, Jackie Mahar, Wesley D. Black, Subhash Vasudevan, Roman Lenchine, Kirsten Spann, Daniel J. Rawle, Penny Rudd, Jessica Neil, Richard Kingston, Timothy P. Newsome, Ki Wook Kim, Johnson Mak, Kym Lowry, Nathan Bryant, Joanne Meers, Jason A. Roberts, Nigel McMillan, Larisa I. Labzin, Andrii Slonchak, Leon E. Hugo, Bennett Henzeler, Natalee D. Newton, Cassandra T. David, Patrick C. Reading, Camille Esneau, Tatiana Briody, Najla Nasr, Donna McNeale, Brian McSharry, Omid Fakhri, Bethany A. Horsburgh, Grant Logan, Paul Howley, Paul Youn

High Quality Long-Term CD4+ and CD8+ Effector Memory Populations Stimulated by DNA-LACK/MVA-LACK Regimen in Leishmania major BALB/c Model of Infection

Heterologous vaccination based on priming with a plasmid DNA vector and boosting with an attenuated vaccinia virus MVA recombinant, with both vectors expressing the Leishmania infantum LACK antigen (DNA-LACK and MVA-LACK), has shown efficacy conferring protection in murine and canine models against cutaneus and visceral leishmaniasis, but the immune parameters of protection remain ill defined. Here we performed by flow cytometry an in depth analysis of the T cell populations induced in BALB/c mice during the vaccination protocol DNA-LACK/MVA-LACK, as well as after challenge with L. major parasites. In the adaptive response, there is a polyfunctional CD4+ and CD8+ T cell activation against LACK antigen. At the memory phase the heterologous vaccination induces high quality LACK-specific long-term CD4+ and CD8+ effector memory cells. After parasite challenge, there is a moderate boosting of LACK-specific CD4+ and CD8+ T cells. Anti-vector responses were largely CD8+-mediated. The immune parameters induced against LACK and triggered by the combined vaccination DNA/MVA protocol, like polyfunctionality of CD4+ and CD8+ T cells with an effector phenotype, could be relevant in protection against leishmaniasis

Statement in Support of: "Virology under the Microscope-a Call for Rational Discourse".

Letter to the Editor OnlinePublPeter Speck ... Michael Beard ... et al

A Temporal Role Of Type I Interferon Signaling in CD8+ T Cell Maturation during Acute West Nile Virus Infection

A genetic absence of the common IFN- α/β signaling receptor (IFNAR) in mice is associated with enhanced viral replication and altered adaptive immune responses. However, analysis of IFNAR-/- mice is limited for studying the functions of type I IFN at discrete stages of viral infection. To define the temporal functions of type I IFN signaling in the context of infection by West Nile virus (WNV), we treated mice with MAR1-5A3, a neutralizing, non cell-depleting anti-IFNAR antibody. Inhibition of type I IFN signaling at or before day 2 after infection was associated with markedly enhanced viral burden, whereas treatment at day 4 had substantially less effect on WNV dissemination. While antibody treatment prior to infection resulted in massive expansion of virus-specific CD8+ T cells, blockade of type I IFN signaling starting at day 4 induced dysfunctional CD8+ T cells with depressed cytokine responses and expression of phenotypic markers suggesting exhaustion. Thus, only the later maturation phase of anti-WNV CD8+ T cell development requires type I IFN signaling. WNV infection experiments in BATF3-/- mice, which lack CD8-α dendritic cells and have impaired priming due to inefficient antigen cross-presentation, revealed a similar effect of blocking IFN signaling on CD8+ T cell maturation. Collectively, our results suggest that cell non-autonomous type I IFN signaling shapes maturation of antiviral CD8+ T cell response at a stage distinct from the initial priming event

Virulence in Murine Model Shows the Existence of Two Distinct Populations of Brazilian Vaccinia virus Strains

Brazilian Vaccinia virus had been isolated from sentinel mice, rodents and recently from humans, cows and calves during outbreaks on dairy farms in several rural areas in Brazil, leading to high economic and social impact. Some phylogenetic studies have demonstrated the existence of two different populations of Brazilian Vaccinia virus strains circulating in nature, but little is known about their biological characteristics. Therefore, our goal was to study the virulence pattern of seven Brazilian Vaccinia virus strains. Infected BALB/c mice were monitored for morbidity, mortality and viral replication in organs as trachea, lungs, heart, kidneys, liver, brain and spleen. Based on the virulence potential, the Brazilian Vaccinia virus strains were grouped into two groups. One group contained GP1V, VBH, SAV and BAV which caused disease and death in infected mice and the second one included ARAV, GP2V and PSTV which did not cause any clinical signals or death in infected BALB/c mice. The subdivision of Brazilian Vaccinia virus strains into two groups is in agreement with previous genetic studies. Those data reinforce the existence of different populations circulating in Brazil regarding the genetic and virulence characteristics