- Email: [email protected]
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Abstract / Cytokine 63 (2013) 243–314 However, IL-2 is not required to maintain these cells in the skin and skin-draining lymph nodes. Conversely, IL-7 is essential for maintaining mTregs in skin in the steady state. These results elucidate the fundamental biology of mTregs and show that IL-7 plays an important role in their survival in skin. Support: Erwin Schroedinger Fellowship from the Austrian Science Fund to I.K.G. National Institutes of Health (NIH) Grant 1K08AR062064-01, Burroughs Wellcome Career Award for Medical Scientists, and Scleroderma Research Foundation to M.D.R. NIH Grants P01 AI35297, R01 AI73656, and U19 AI56388 to A.K.A. http://dx.doi.org/10.1016/j.cyto.2013.06.180
178 Characterization of PD-1+Tim-3+ ‘Exhausted’ CD8 T cells in mouse models of systemic lupus erythematosus Tom McCaughtry, Isharat Yusuf, Sandra Gallagher, Ronald Herbst, Laura Carter, Yue Wang, Respiratory, Inflammation & Autoimmunity, MedImmune LLC., Gaithersburg, MD, United States Systemic lupus erythematosus is an autoimmune disorder characterized by the presence of autoantibodies and inflammation that leads to tissue damage. Whether CD8 T cells play a role in the pathogenesis of SLE is not clear. The accumulation of ‘exhausted’ CD8 T cells with impaired effector function and distinguished by coexpression of PD-1, Tim-3, and Lag-3 has been described under a number of settings including chronic viral infection, sepsis, and cancer. In the current study, we investigated the phenotype of CD8 T cells in multiple mouse models of lupus. We report an age-dependent accumulation of ‘exhausted’ CD8 T cells in multiple mouse models of lupus. The phenotype of these cells is defined by expression of PD-1, along with a substantial population of cells co-expressing PD-1 with Tim-3 and Lag-3. A similar accumulation of ‘exhausted’ CD8 T cells was found in SLE1, NZBWF1, Sanroque, and MRL/ lpr strains and was significantly greater than that found in age-matched C57Bl/6 mice. Furthermore PD-1+ CD8 T cells were distributed throughout the body in both secondary lymphoid organs as well as a variety of tissues, such as liver and kidney. These cells exhibited altered cytokine production upon ex vivo stimulation with more cells capable of producing Granzyme B, IFNc, and IL-10, and fewer cells producing TNFa or IL-2. Moreover, these cells exhibited impaired survival after anti-CD3/CD28 costimulation. The frequency of PD-1+ CD8 T cells showed a strong positive correlation with the frequency of GC B cells as well as PD-1+ CD4 T cells. These results suggest that ‘exhausted’ CD8 T cells may contribute to the onset or exacerbation of disease. Alternatively, impaired CD8 T cell function may be a secondary consequence of the disease that may result in increased susceptibility to pathogen infection. Future studies in human and animal models are warranted. Disclosure: all authors are employees of MedImmune.
285
that modulation of miRNA may provide an alternative mechanism for the use of IL10 therapeutically.
http://dx.doi.org/10.1016/j.cyto.2013.06.182
180 The TNF family member TL1A: A key player in type 2 immunity in both adaptive and innate lymphocytes Françoise Meylan a, Eric Hawley a, Luke Barron b, Jillian Barlow a,b,c,d, Cuyian Tan c, Pallavi Penumetcha a, Arianne Richard a, Xi Chen a,b,c,d, William E. Paul a,b,c,d, Thomas A. Wynn b, Igal Gery c, Andrew McKenzie d, Richard M. Siegel a, a Immunoregulation Section, Autoimmunity Branch, NIAMS, USA, b Immunopathogenesis Section, LPD, NIAID, USA, c Experimental Immunology Section, NEI, National Institutes of Health, Bethesda, MD, USA, d Laboratory of Molecular Biology Cambridge University, Cambridge, UK The TNF-family cytokine TL1A costimulates T cells through its receptor DR3. TL1A polymorphisms and increased TL1A expression has been linked to Rheumatoid Arthritis and Inflammatory Bowel Disease. TL1A-DR3 interactions are required for diverse mouse models of autoimmune disease. The effects of TL1A on T cell differentiation and its role in innate lymphocyte biology has not been explored. During T cell activation, we have found that TL1A costimulation specifically promotes production of the allergy-promoting cytokine IL-9 through a mechanism dependent on IL-2 and STAT5 signaling, and TL1A promotes pathology in mouse models of asthma and ocular inflammation. Transgenic mice chronically expressing TL1A spontaneously develop small intestinal pathology characterized by high levels of IL-13, muscular and goblet cell hyperplasia, and infiltration with immune cells. These histological changes are dependent on IL-13 but not T cells or commensal flora. We find that the major IL13 producing cells in TL1A transgenic mice lack T and B cell lineage markers and are phenotypically similar to ‘type 2’ innate lymphocytes (ILC2), which promote allergic and anti-parasitic responses in mice. ILC2 express surface DR3 and produce IL-13 in response to TL1A ex vivo. However, ILC2 can be expanded normally in DR3 deficient mice by IL-25 or IL-33, or Nippostrongylus brasiliensis, and DR3 deficient mice clear Nippostrongylus normally from the intestine. TL1A can thus induce IL-13 production by innate lymphocytes through mechanisms distinct from parasitic infection. Taken together, these data demonstrate that TL1A coordinately enhances type 2 immunity in both innate and adaptive lymphocytes and may be a good target for therapy in allergic diseases.
http://dx.doi.org/10.1016/j.cyto.2013.06.183
http://dx.doi.org/10.1016/j.cyto.2013.06.181 181 Ribavirin enhances and prolongs IFN signaling and ISG expression in virallyinfected human lung epithelial cells 179 MIRNA analysis reveals novel insights into IL10 function Claire E. McCoy a,b, Susan Quinn b, Michael P. Gantier a, Kirsten Fairfax c, Luke A. O’Neill b, Bryan R.G. Williams a, a Monash Institute of Medical Research, Monash University, Clayton, VIC, Australia, b School of Biochemistry and Immunology, Trinity College Dublin, Ireland, c Department of Immunology, Alfred Hospital, Monash University, VIC, Australia IL10 is a pluripotent cytokine that has multiple functions on cells of the immune system. In most cases, IL10 acts as an anti-inflammatory cytokine, dampening the inflammatory response by down-regulating a subset of pro-inflammatory genes induced by TLR signalling. In B cells, IL10 appears to have an additional role enhancing proliferation, differentiation and class-switch recombination required for appropriate responses to antigens. Although the overall mechanism of IL10 action is known, there still remain many gaps in our knowledge. Furthermore, many attempts have failed to utilize IL10 for the treatment of inflammatory conditions. In an effort to understand if miRNAs can shed light on IL10 function, we performed a miRNA array on B cells. We discovered that IL10 can repress key TLR-induced miRNA, such as miR155, miR-146a and miR-9 as well as up-regulating a range of unknown miRNA. Bioinformatic and experimental analysis revealed that genes such as SHIP1 and Aicda, two well-known miR-155 targets in macrophages and B cells respectively, can be regulated by IL10 through its modulation of miR-155. Analogously, target analysis of specific miRNAs identified in the array, predict that IL10 can regulate a range of previously uncharacterised genes required for its effect on immune cell function. These findings not only highlight a novel mechanism of action for IL10 but suggest
A.N. Morrow, K.C. Zoon, National Institute of Allergy and Infectious Diseases, Bethesda, MD, USA Combination treatments of IFN and ribavirin are successful in treating viral infections, however the mechanism of action for how these agents work together remains unclear. To examine this, we utilized vesicular stomatitis virus (VSV) infection on A549 human lung epithelial cells as an experimental model. After 15 h of pretreatment, we found that a combination of IFN-alpha2a and ribavirin results in complete protection from virus-induced CPE whereas these treatments alone failed to protect cells from viral challenge. The combination treatment led to a 3-log decrease in VSV titer at 24 and 48 h. We analyzed STAT activation and protein expression in cell lysates harvested at 8, 24, and 48 h post-infection and found that treating cells with ribavirin alone or in combination with IFN-alpha2a resulted in increased and prolonged activation of pSTAT1 and pSTAT2. Ribavirin alone or in combination with IFN-alpha2a resulted in increases in total STAT1, STAT2, and IRF9 protein expression compared to IFN-alpha2a alone. In addition, expression of MxA, IFIT3, PKR, and ISG15 showed enhanced expression in the presence of ribavirin alone or in combination with IFN-alpha2a when compared to IFN-alpha2a alone. The increases in STAT activation and ISG expression were not present in uninfected cells, suggesting that the enhanced activity induced by ribavirin required an active viral infection. This prolonged and enhanced activity of IFN signaling and induction of gene expression may play a role in how IFN and ribavirin work together to treat viral infections. http://dx.doi.org/10.1016/j.cyto.2013.06.184
178 Characterization of PD-1+Tim-3+ ‘Exhausted’ CD8 T cells in mouse models of systemic lupus erythematosus Tom McCaughtry, Isharat Yusuf, Sandra Gallagher, Ronald Herbst, Laura Carter, Yue Wang, Respiratory, Inflammation & Autoimmunity, MedImmune LLC., Gaithersburg, MD, United States Systemic lupus erythematosus is an autoimmune disorder characterized by the presence of autoantibodies and inflammation that leads to tissue damage. Whether CD8 T cells play a role in the pathogenesis of SLE is not clear. The accumulation of ‘exhausted’ CD8 T cells with impaired effector function and distinguished by coexpression of PD-1, Tim-3, and Lag-3 has been described under a number of settings including chronic viral infection, sepsis, and cancer. In the current study, we investigated the phenotype of CD8 T cells in multiple mouse models of lupus. We report an age-dependent accumulation of ‘exhausted’ CD8 T cells in multiple mouse models of lupus. The phenotype of these cells is defined by expression of PD-1, along with a substantial population of cells co-expressing PD-1 with Tim-3 and Lag-3. A similar accumulation of ‘exhausted’ CD8 T cells was found in SLE1, NZBWF1, Sanroque, and MRL/ lpr strains and was significantly greater than that found in age-matched C57Bl/6 mice. Furthermore PD-1+ CD8 T cells were distributed throughout the body in both secondary lymphoid organs as well as a variety of tissues, such as liver and kidney. These cells exhibited altered cytokine production upon ex vivo stimulation with more cells capable of producing Granzyme B, IFNc, and IL-10, and fewer cells producing TNFa or IL-2. Moreover, these cells exhibited impaired survival after anti-CD3/CD28 costimulation. The frequency of PD-1+ CD8 T cells showed a strong positive correlation with the frequency of GC B cells as well as PD-1+ CD4 T cells. These results suggest that ‘exhausted’ CD8 T cells may contribute to the onset or exacerbation of disease. Alternatively, impaired CD8 T cell function may be a secondary consequence of the disease that may result in increased susceptibility to pathogen infection. Future studies in human and animal models are warranted. Disclosure: all authors are employees of MedImmune.
285
that modulation of miRNA may provide an alternative mechanism for the use of IL10 therapeutically.
http://dx.doi.org/10.1016/j.cyto.2013.06.182
180 The TNF family member TL1A: A key player in type 2 immunity in both adaptive and innate lymphocytes Françoise Meylan a, Eric Hawley a, Luke Barron b, Jillian Barlow a,b,c,d, Cuyian Tan c, Pallavi Penumetcha a, Arianne Richard a, Xi Chen a,b,c,d, William E. Paul a,b,c,d, Thomas A. Wynn b, Igal Gery c, Andrew McKenzie d, Richard M. Siegel a, a Immunoregulation Section, Autoimmunity Branch, NIAMS, USA, b Immunopathogenesis Section, LPD, NIAID, USA, c Experimental Immunology Section, NEI, National Institutes of Health, Bethesda, MD, USA, d Laboratory of Molecular Biology Cambridge University, Cambridge, UK The TNF-family cytokine TL1A costimulates T cells through its receptor DR3. TL1A polymorphisms and increased TL1A expression has been linked to Rheumatoid Arthritis and Inflammatory Bowel Disease. TL1A-DR3 interactions are required for diverse mouse models of autoimmune disease. The effects of TL1A on T cell differentiation and its role in innate lymphocyte biology has not been explored. During T cell activation, we have found that TL1A costimulation specifically promotes production of the allergy-promoting cytokine IL-9 through a mechanism dependent on IL-2 and STAT5 signaling, and TL1A promotes pathology in mouse models of asthma and ocular inflammation. Transgenic mice chronically expressing TL1A spontaneously develop small intestinal pathology characterized by high levels of IL-13, muscular and goblet cell hyperplasia, and infiltration with immune cells. These histological changes are dependent on IL-13 but not T cells or commensal flora. We find that the major IL13 producing cells in TL1A transgenic mice lack T and B cell lineage markers and are phenotypically similar to ‘type 2’ innate lymphocytes (ILC2), which promote allergic and anti-parasitic responses in mice. ILC2 express surface DR3 and produce IL-13 in response to TL1A ex vivo. However, ILC2 can be expanded normally in DR3 deficient mice by IL-25 or IL-33, or Nippostrongylus brasiliensis, and DR3 deficient mice clear Nippostrongylus normally from the intestine. TL1A can thus induce IL-13 production by innate lymphocytes through mechanisms distinct from parasitic infection. Taken together, these data demonstrate that TL1A coordinately enhances type 2 immunity in both innate and adaptive lymphocytes and may be a good target for therapy in allergic diseases.
http://dx.doi.org/10.1016/j.cyto.2013.06.183
http://dx.doi.org/10.1016/j.cyto.2013.06.181 181 Ribavirin enhances and prolongs IFN signaling and ISG expression in virallyinfected human lung epithelial cells 179 MIRNA analysis reveals novel insights into IL10 function Claire E. McCoy a,b, Susan Quinn b, Michael P. Gantier a, Kirsten Fairfax c, Luke A. O’Neill b, Bryan R.G. Williams a, a Monash Institute of Medical Research, Monash University, Clayton, VIC, Australia, b School of Biochemistry and Immunology, Trinity College Dublin, Ireland, c Department of Immunology, Alfred Hospital, Monash University, VIC, Australia IL10 is a pluripotent cytokine that has multiple functions on cells of the immune system. In most cases, IL10 acts as an anti-inflammatory cytokine, dampening the inflammatory response by down-regulating a subset of pro-inflammatory genes induced by TLR signalling. In B cells, IL10 appears to have an additional role enhancing proliferation, differentiation and class-switch recombination required for appropriate responses to antigens. Although the overall mechanism of IL10 action is known, there still remain many gaps in our knowledge. Furthermore, many attempts have failed to utilize IL10 for the treatment of inflammatory conditions. In an effort to understand if miRNAs can shed light on IL10 function, we performed a miRNA array on B cells. We discovered that IL10 can repress key TLR-induced miRNA, such as miR155, miR-146a and miR-9 as well as up-regulating a range of unknown miRNA. Bioinformatic and experimental analysis revealed that genes such as SHIP1 and Aicda, two well-known miR-155 targets in macrophages and B cells respectively, can be regulated by IL10 through its modulation of miR-155. Analogously, target analysis of specific miRNAs identified in the array, predict that IL10 can regulate a range of previously uncharacterised genes required for its effect on immune cell function. These findings not only highlight a novel mechanism of action for IL10 but suggest
A.N. Morrow, K.C. Zoon, National Institute of Allergy and Infectious Diseases, Bethesda, MD, USA Combination treatments of IFN and ribavirin are successful in treating viral infections, however the mechanism of action for how these agents work together remains unclear. To examine this, we utilized vesicular stomatitis virus (VSV) infection on A549 human lung epithelial cells as an experimental model. After 15 h of pretreatment, we found that a combination of IFN-alpha2a and ribavirin results in complete protection from virus-induced CPE whereas these treatments alone failed to protect cells from viral challenge. The combination treatment led to a 3-log decrease in VSV titer at 24 and 48 h. We analyzed STAT activation and protein expression in cell lysates harvested at 8, 24, and 48 h post-infection and found that treating cells with ribavirin alone or in combination with IFN-alpha2a resulted in increased and prolonged activation of pSTAT1 and pSTAT2. Ribavirin alone or in combination with IFN-alpha2a resulted in increases in total STAT1, STAT2, and IRF9 protein expression compared to IFN-alpha2a alone. In addition, expression of MxA, IFIT3, PKR, and ISG15 showed enhanced expression in the presence of ribavirin alone or in combination with IFN-alpha2a when compared to IFN-alpha2a alone. The increases in STAT activation and ISG expression were not present in uninfected cells, suggesting that the enhanced activity induced by ribavirin required an active viral infection. This prolonged and enhanced activity of IFN signaling and induction of gene expression may play a role in how IFN and ribavirin work together to treat viral infections. http://dx.doi.org/10.1016/j.cyto.2013.06.184