Baylor Institute for Immunology Research (BIIR): Baylor Symposium and Workshop on Human Immunology and Biodefense Abstracts

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First Baylor Symposium and Workshop on Human Immunology and Biodefense, November 6-7, 2004: Videos and Abstracts

Videos from the Symposium are available for you to watch online.

	Module 1: Welcome Video in MOV format: 1:44 minutes, 4.66 MB Video in WMX format: 1:44 minutes, 5.30MB
	Module 2: Human Immunology and Biodefense Centers Video in MOV format: 14.09 minutes, 39.67 MB Video in WMX format: 14.09 minutes, 45.50 MB
	Module 3: Human T Memory Cells Video in MOV format: 50:02 minutes, 139.94 MB Video in WMX format: 50:02 minutes, 158.58 MB
	Module 4: Human DC Subsets Video in MOV format: 45:26 minutes, 126.54 MB Video in WMX format: 45:26 minutes, 146.30 MB
	Module 5: Deconstructing the Molecular Basis of T Cell Recognition and the Rapid Assessment of Human T Cell Responsiveness Video in MOV format: 51:08 minutes, 142.28 MB Video in WMX format: 51:08 minutes, 152.84 MB
	Module 6: Influenza Virus Video in MOV format: 39:40 minutes, 110.53 MB Video in WMX format: 39:40 minutes, 128.33 MB
	Module 7: Immune Response to Smallpox Video in MOV format: 48:09 minutes, 134.37 MB Video in WMX format: 48:09 minutes, 156.38 MB
	Module 8: Patterns of Host Response to Systemic Infection Video in MOV format: 44:22 minutes, 123.70 MB Video in WMX format: 44:22 minutes, 141.88 MB
	Module 9: CD1 Antigen Presenting Molecules Video in MOV format: 50:17 minutes, 140.21 MB Video in WMX format: 50:17 minutes, 164.05 MB
	Module 10: Models to Study Class A Pathogens Video in MOV format: 29:11 minutes, 81.30 MB Video in WMX format: 29:11 minutes, 94.08 MB
	Module 11: Morning Workshop Introduction: Measuring Human T Cell Immunity Video in MOV format: 37:58 minutes, 109.66 MB Video in WMX format: 37:58 minutes, 127.69 MB
	Module 12: Luminex In Vitro and In Vivo Video in MOV format: 14:15 minutes, 39.74 MB Video in WMX format: 14:15 minutes, 44.06 MB
	Module 13: Modulation of tumor-specific T cell response with dendritic cell vaccination Hideki Ueno, John E. Connolly, Luis Vence, Joseph Fay, Karolina Palucka, and Jacques Banchereau, Baylor Institute for Immunology Research. Abstract Video in MOV format: 18:50 minutes, 51.47 MB Video in WMX format: 18:50 minutes, 59.19 MB
	Module 14: Global analysis of the immune response to vaccinia for directed design of new vaccines Jonathan Duke-Cohan, Pedro Reche, Elizabeth Witten and Ellis Reinherz, Dana-Farber Cancer Institute, Department of Medical Oncology and Harvard Medical School. Abstract Video in MOV format: 15:04 minutes, 41.93 MB Video in WMX format: 15:04 minutes, 48.59 MB
	Module 15: T cell-dependent IFN- g production by NK cells in response to influenza A virus Xiao-Song He,1 Monia Draghi,1 Kutubuddin Mahmood,2 Tyson H. Holmes,1George W. Kemble,2 Cornelia L. Dekker,1 Ann M. Arvin,1 Peter Parham,1 and Harry B. Greenberg1, 1Stanford University School of Medicine, 2Medimmune Vaccines. Abstract Video in MOV format: 19:13 minutes, 53.43 MB Video in WMX format: 19:13 minutes, 62.17 MB
	Module 16: Gene Expression Patterns in Blood Leukocytes from Patients with Acute Infections Discriminate the Class of Pathogen Windy Allman, Wendy Chung, Octavio Ramilo, Knut M.Wittkowski, Asuncion Mejias, Bernard Piqueras, Damien Chaussabel, Lynda Bennett, Virginia Pascual, Jacques Banchereau, and A. Karolina Palucka, Baylor NIAID Cooperative Center for Translational Research on Human Immunology and Biodefense and Baylor Institute for Immunology, and Division of Pediatric Infectious Diseases, UT Southwestern Medical Center and Children's Medical Center of Dallas and General Clinical Research Center, The Rockefeller University (K.M.W.). Abstract Video in MOV format: 10:42 minutes, 29.87 MB Video in WMX format: 10:42 minutes, 33.82 MB
	Module 17: Influenza virus induces a programmed response in human blood dendritic cells, which control immune effectors attraction Bernard Piqueras, John Connolly, Heidi Freitas, Casey Glaser, Windy Allman, A. Karolina Palucka, and Jacques Banchereau, Baylor Institute for Immunology Research Abstract Video in MOV format: 13:11 minutes, 48.23 MB Video in WMX format: 13:11 minutes, 55.83 MB
	Module 18: Afternoon Workshop Introduction: Measuring Human T Cell Immunity Video in MOV format: 29:24 minutes, 93.48 MB Video in WMX format: 29:24 minutes, 107.75 MB
	Module 19: CMKLR1 Expression and Chemerin-directed Chemotaxis Distinguish Plasmacytoid from Myeloid Dendritic Cells in Human Blood Brian A. Zabel, Amanda M. Silverio, Eugene C. Butcher, Laboratory of Immunology and Vascular Biology, Department of Pathology, Stanford University School of Medicine, and Center for Molecular Biology and Medicine, Veterans Affairs Palo Alto Health Care System. Abstract Video in MOV format: 10:32 minutes, 29.25 MB Video in WMX format: 10:32 minutes, 33.20 MB
	Module 20: Proteases of Dendritic Cells and Dendritic Cell Subsets Nathanael McCurley, Ph.D. Candidate, Mellman Lab,Section of Immunobiology, Yale University Abstract Video in MOV format: 15:29 minutes, 54.63 MB Video in WMX format: 15:29 minutes, 60.98 MB
	Module 21: Regulation of Dendritic cell maturation by E-cadherin/ ß-catenin signaling Aimin Jiang, and Ira Mellman, Department of Cell Biology, Ludwig Institute for Cancer Research, Yale University School of Medicine Abstract Video in MOV format: 26:22 minutes, 74.07 MB Video in WMX format: 26:22 minutes, 86.49 MB
	Module 22: Respiratory Syncytial Virus (RSV) Affects Myeloid Dendritic Cell (mDC) Function Michelle Gill1,2, John Connolly1, Yaming Xue1, Vikram Bhuta1,2, Octavio Ramilo1,2, and Jacques Banchereau1,2. Baylor Institute for Immunology Research1 and Division of Pediatric Infectious Diseases, UT Southwestern Medical Center2 Abstract Video in MOV format: 14:04 minutes, 39.04 MB Video in WMX format: 14:04 minutes, 44.51 MB
	Module 23: Human breast cancer and melanoma differentially modulate dendritic cells in OncoHumouse Caroline Aspord, Jacques Banchereau, Mike Gallegos, Sasha Tindle, Florentina Marches and A. Karolina Palucka. Baylor Institute for Immunology Research. Abstract Video in MOV format: 9:53 minutes, 27.33 MB Video in WMX format: 9:53 minutes, 31.91 MB
	Module 24: Literature mining tools for knowledge discovery in microarray data Damien Chaussabel, Charlie Quinn and Jacques Banchereau, Baylor Institute for Immunology Research Abstract Video in MOV format: 18:09 minutes, 50.64 MB Video in WMX format: 18:09 minutes, 58.83 MB
	Langerhans Cells within CD34-DC efficiently prime melanoma specific CD8+ T cells E Klechevsky, Ueno H, Reiter R, Palucka A, Banchereau J. BIIR and Technion. Abstract
	Global Cytokine and Chemokine Analysis for High Throughput Immunomonitoring John Connolly, Baylor Institute for Immunology Research Abstract

CMKLR1 Expression and Chemerin-directed Chemotaxis Distinguish Plasmacytoid from Myeloid Dendritic Cells in Human Blood

Brian A. Zabel, Amanda M. Silverio, Eugene C. Butcher, Laboratory of Immunology and Vascular Biology, Department of Pathology, Stanford University School of Medicine, Stanford, CA 94305, and Center for Molecular Biology and Medicine, Veterans Affairs Palo Alto Health Care System, Palo Alto, CA 94304.

The organization and function of the immune system is established through precise positioning of white blood cells throughout the body. Leukocyte-expressed serpentine chemoattractant receptors play an integral role in an elegant, multi-component system of leukocyte traffic control. For some cells types, such as naive CD4+ T cells, the trafficking profile is well characterized: CCR7, L-selectin, and LFA-1 facilitate transit between blood and lymph nodes. For other cell types, such as plasmacytoid dendritic cells (pDC), their systemic positioning is not well understood. pDC are versatile cells of the immune response, secreting type I interferons and differentiating into potent immunogenic or tolerogenic antigen-presenting cells. pDC can express adhesion and chemokine receptors for lymphoid tissues, but are also recruited by unknown mechanisms during tissue inflammation. We use a novel monoclonal antibody specific for serpentine receptor CMKLR1 to evaluate its expression by circulating leukocytes in man. We show that CMKLR1 is expressed by circulating pDC in human blood, whereas myeloid DC (mDC) as well as lymphocytes, monocytes, neutrophils and eosinophils are negative. We identify a major serum agonist activity for CMKLR1 as chemerin, a proteolytically activatable attractant and the sole known ligand for CMKLR1, and show that chemerin is activated during blood coagulation and attracts pDC but not mDC in ex vivo chemotaxis assays. We conclude that CMKLR1 expression and chemerin-mediated chemotaxis distinguish circulating pDC from mDC, providing a potential mechanism for their differential contribution to or regulation of immune responses at sites of bleeding or inflammatory protease activity.

Gene Expression Patterns in Blood Leukocytes from Patients with Acute Infections Discriminate the Class of Pathogen.

Windy Allman, Wendy Chung, Octavio Ramilo, Knut M.Wittkowski, Asuncion Mejias, Bernard Piqueras, Damien Chaussabel, Lynda Bennett, Virginia Pascual, Jacques Banchereau, and A. Karolina Palucka, Baylor NIAID Cooperative Center for Translational Research on Human Immunology and Biodefense and Baylor Institute for Immunology, and Division of Pediatric Infectious Diseases, UT Southwestern Medical Center and Children's Medical Center of Dallas all in Dallas, Texas; and General Clinical Research Center, The Rockefeller University (K.M.W.), New York, NY.

Different classes of pathogens trigger specific pattern-recognition receptors differentially expressed on multiple cells of the immune system, many of which are present in the blood. We surmised that this would elicit distinct mRNA biosignatures in blood leukocytes of infected humans. To this end, we analyzed gene-expression profiles in blood samples from 50 pediatric patients with acute infection (Influenza virus, gram-negative bacterium E. coli and gram-positive bacterium S. aureus) using Affymetrix U133A GeneChips containing 14,500 unique genes or expression-signature tags. In a univariate analysis, Fisher's exact test or Mann-Whitney U test, genes were ranked on the basis of their ability to discriminate the class of pathogen. An independent set of samples from 26 patients with various gram-negative and gram-positive bacterial infection was analyzed using the K-nearest neighbors clustering (k-NN) for class identification. We identified distinct gene expression patterns that allow discrimination between three classes of pathogens we tested, i.e., Influenza virus, gram-negative and gram-positive bacteria, which are among the most common infections leading to hospitalization. Differential patterns of expression of 736 genes were identified between patients with influenza A and patients with bacterial infections (p<.01). As expected, patients with Influenza A infections had higher expression of genes known to be induced by IFN-alpha. In class discrimination, 39 of the top ranked genes, identified infection type, influenza or bacterial, with 94% accuracy. 583 genes were found to have differential expression between E.coli and S.aureus samples (p<.05). In class discrimination, 34 of the top ranked genes, identified infection type, gram-positive or gram-negative, with 91% accuracy. Real-time RT-PCR quantitation of the expression level of nine selected discriminative genes confirmed the microarray results. Thus, assessing the host response to the invading microbe through transcriptome analysis of blood leukocytes presents a novel approach to the diagnosis of acute infection.

Global analysis of the immune response to vaccinia for directed design of new vaccines

Jonathan Duke-Cohan, Pedro Reche, Elizabeth Witten and Ellis Reinherz, Dana-Farber Cancer Institute, Department of Medical Oncology and Harvard Medical School, Boston MA 02115

As a rational approach to designing new smallpox vaccines, we have initiated a long-term study to follow the immune response to vaccination and re-challenge. The aim is to to design and develop methodology to characterize the cell-mediated response and humoral responses at both molecular and functional levels. The developed resources include a suite of bioinformatics utilities to predict HLA Class I-binding epitopes within the orthopox virus proteome. We have synthesized 458 peptides derived from the vaccinia open reading frames, and shared with variola, that are predicted to bind to the HLA-A2 supertype and which we are now testing for recognition by CD8+ cells from immunized individuals. We have established a reliable protocol using intracellular IFN-g synthesis as a measure of peptide recognition to determine the changing target repertoire of vaccinia-reactive CD8+ T cells after virus exposure. We also have set up a reliable technique for directly quantitating binding of vaccinia-derived peptides to HLA Class I. Having identified peptide epitopes, we will then be able to use peptide-MHC tetramers to isolate reactive T cells and analyse the T cell receptor a/b chain usage by spectratyping. We have also initiated an extensive effort to use in vitro coupled transcription/translation to manufacture a complete representation of the vaccinia proteome as a microarray, allowing us to follow the serological response to vaccinia. These techniques will enable us to identify poxvirus targets likely to be highly immunogenic for both arms of the immune response, and which may then be used to design recombinant protein immunization protocols not dependent upon live or attenuated virus. Furthermore, the established techniques will function as a model program for developing rational recombinant-based vaccines against any infectious agent.

Global Cytokine and Chemokine Analysis for High Throughput Immunomonitoring

John Connolly, Baylor Institute for Immunology Research, Dallas, Texas

Coordinated changes in cytokine networks, characteristic of specific immune responses, may represent predictive indicators during the course of immunomonitoring. By integrating automated high throughput cytokine multiplex assays with complex data visualization and analysis techniques, state specific bio-signatures can be identified in patient serum and cell culture samples. The descriptive and predictive value of these signatures in the context of autoimmune and cancer immunotherapy will be discussed. Novel techniques based on this platform will also be described.

Human breast cancer and melanoma differentially modulate dendritic cells in OncoHumouse

Caroline Aspord, Jacques Banchereau, Mike Gallegos, Sasha Tindle, Florentina Marches and A. Karolina Palucka. Baylor Institute for Immunology Research, Dallas, TX, USA.

Human tumors are infiltrated by dendritic cells. Yet tumor growth indicates the lack of effective anti-tumor immunity. To understand how tumors modulate DC function, we have developed a novel model to study the in vivo interactions between human cancer and human dendritic cells in an immunodeficient mouse. NOD-SCID ?2m-/- mice engrafted with human CD34+ hematopoietic progenitors develop myeloid and plasmacytoid DCs. Human tumor cell lines, such as breast cancer and melanoma, can develop in these animals (OncoHumouse). We demonstrate that these tumors differentially attract human DC and monocytes / macrophages in vivo. Indeed, breast cancer attracts larger numbers of human DCs and promotes their maturation and migration to the draining lymph nodes. This differential modulation of human DCs could be explained by different chemokine environnement observed in vivo as breast tumor secreted mostly MCP1 and IL6 and melanoma produced IL8. DC were functional as assessed by their capacity to induce allogeneic CD4 proliferation ex vivo and their ability to prime T cells against the tumor in vivo. Furthermore, DC isolated from breast tumor and its draining lymph nodes, as opposed to DC from spleen or BM, induced the production of high level of IL4, IL13 and TNF? by naive CD4 T cells, suggesting that breast tumor modulate DC to induce an inflammatory Th2 response. Thus, OncoHumouse can be used to study the development and modulation of tumor immunity.

Influenza virus induces a programmed response in human blood dendritic cells, which control immune effectors attraction

Bernard Piqueras, John Connolly, Heidi Freitas, Casey Glaser, Windy Allman, A. Karolina Palucka, and Jacques Banchereau, Baylor Institute for Immunology Research

Host response to viral infection involves distinct effectors of the innate and adaptive immunity, whose mobilization needs to be coordinated to insure protection. Dendritic cells (DCs) are considered essential in the control of immune responses. Here we show that Influenza virus triggers in human plasmacytoid and myeloid DCs, a coordinated chemokine secretion program with three successive waves. The first one, occurring at early time points (2-4 hours), includes chemokines potentially attracting effector cells like neutrophils, cytotoxic T and NK cells (CXCL16, CXCL1, CXCL2, and CXCL3). The second one occurs within 8 to 12 hours and includes chemokines attracting effector memory T cells (CXCL8, CCL3, CCL4, CCL5, CXCL9, CXCL10, and CXCL11). The third wave, which occurs after 24-48 hours, probably when DCs have reached the lymphoid organs, includes CCL19, CCL22 and CXCL13, which attract naïve T and B lymphocytes. Thus, human blood DCs carry a common program of chemokine production, which allows a coordinated attraction of the different immune effectors upon viral infection.

Langerhans Cells within CD34-DC efficiently prime melanoma specific CD8+ T cells

E Klechevsky, Ueno H, Reiter R, Palucka A, Banchereau J. BIIR, Dallas,TX and Technion, Haifa, Israel.

Culturing CD34 HPCs with GM-CSF and TNFa yields CD34-DCs, which include two DC subsets CD14+ Int DC and CD1a+ LCs, with different phenotype and functions. While CD14 IntDCs, uniquely, induce the differentiation of naïve B cells into IgM secreting cells, no unique functions have been identified for LCs. We analyzed the ability of CD34-DC subsets, to prime autologous naïve CD8+ T cells. T cells were cultured for up to 3 weeks with sorted subsets of HLA-A201+CD34-DCs pulsed with MART-1(26-35) and GP100(209-217) peptides. T cell expansion is measured using i) tetramer binding and , ii) CTL activity against melanoma cells. CD1a+LCs appear to be more efficient than CD14+ intDCs at priming high precursor frequency MART-1 specific CD8+T cells. The efficient priming capacity of CD1a+LCs is further confirmed when measuring the expansion of the low frequency gp100-specific T cells. Tetramer decay experiments showed that CD8+T cells primed by CD1a+LCs are of higher avidity than those primed by CD14+intDCs. CTLs primed by CD1a+LCs were more efficient in killing HLA-A*0201 Me275 melanoma cells. Thus, distinct subsets of CD34-DCs show distinct capacity to prime melanoma-specific CD8+T cells.

Literature mining tools for knowledge discovery in microarray data

Damien Chaussabel, Charlie Quinn and Jacques Banchereau, Baylor Institute for Immunology Research, Dallas, TX, USA.

The rate-limiting step in high throughput experimentation is neither data acquisition nor analysis, but rather our ability to interpret data on a genome-wide scale. Explosion of data sampling capacity combined with ever increasing publication rates is rapidly transforming biology into a science of information management. We are developing mining strategies and tools to accelerate knowledge discovery in gene expression microarray databases. It involves the implementation of a suite of literature analysis applications: 1) A literature curation tool allows us to address the serious limitations caused by an unruly gene nomenclature. 2) We subsequently developed a dependable gene-centered literature database that can be queried using a stand alone text browser. Literature search results returned by this application are indexed by gene. 3) Gene lists generated experimentally and/or through queries against our literature database can be imported in a literature mining application for downstream analyses. These will include "literature profiling", an original text mining algorithm that groups genes based on patterns of term occurrences in abstracts. A second aspect of our activities is centered on BIIR's extensive microarray data collection. In order to leverage this invaluable resource we will develop a gene expression data repository specifically designed to integrate information accumulated across dozens of studies and hundreds of genechip experiments. Altogether, these unique resources give us the opportunity to design novel mining strategies for microarray data analysis, which focus on hypothesis generation and knowledge discovery.

Modulation of tumor-specific T cell response with dendritic cell vaccination

Hideki Ueno, John E. Connolly, Luis Vence, Joseph Fay, Karolina Palucka, and Jacques Banchereau, Baylor Institute for Immunology Research, Dallas, TX, USA, 75204

To assess T cell responses in melanoma patients vaccinated with killed tumor cells loaded dendritic cell (DC) vaccines, we have developed a high-throughput strategy designated as EPIMAX. The principle of EPIMAX is to measure simultaneously cell proliferation and secretion of multiple cytokines that distinguish Type 1, Type 2 cytokines and IL-10 using the cytokine multiplex technology. Briefly, PBMCs stained with CFSE are cultured with overlapping peptide libraries from selected tumor antigens. After 36-48 h, cytokines in culture supernatants are measured. After 4 additional days of culture, cell proliferation is analyzed by a flowcytometer after staining with anti-CD4 and CD8 mAbs. After determining peptide clusters capable of inducing immune response, another culture is started with individual peptides from that cluster. Thus, this methodology permits us to assess 1) specificity and breadth of anti-tumor immune responses, and 2) function of specific T cells as measured by cytokine secretion and proliferation. After the identification of the peptide and the type of T cell responses, the frequency of specific T cells is measured by analyzing intracellular cytokines in PBMCs stimulated with this peptide together with anti-CD28/CD49d mAbs. We have found in metastatic melanoma patients multiple NY-ESO1, MART-1, and TRP-1 specific CD4+ and CD8+ T cells which recognize novel epitopes. Furthermore, by comparing pre- and post-vaccine PBMCs, we observed; 1) priming of tumor antigen-specific CD4+ and CD8+ T cells, 2) release of melanoma-specific CD8+ T cells from anergic state, and 3) increase in frequency of melanoma-specific CD8+ T cells, as well as 4) elimination of tumor antigen-specific IL-10 producing CD4+ T cells. Thus, DCs loaded with killed tumor cells can induce and/or alter anti-tumor immune responses in metastatic melanoma patients.

This work was supported by NIH grant P01CA84512.

Proteases of Dendritic Cells and Dendritic Cell Subsets

Nathanael McCurley, Ph.D. Candidate, Mellman Lab,Section of Immunobiology, Yale University

In order to function efficiently for the presentation of antigen, dendritic cells (DCs) modulate their antigen processing capacities to preserve antigen for later presentation. Utilizing mouse bone marrow-derived DCs, it has previously been found that DCs utilize a variety of methods to perform this function of antigen preservation, including the attenuation of lysosomal pH and the regulation of lysosomal protease expression. Our recent initial characterization of protease expression in human DC subsets shows a clear variation in expression from one DC subset to another. Plasmacytoid DCs, blood DCs, and CD34+-derived DCs show a remarkably low level of expression of lysosomal proteases, somewhat comparable to what has been found with mouse bone marrow-derived DCs. Conversely, monocyte-derived DCs have a higher level of expression of these proteases that is comparable to the expression level found in macrophages. Given that subsets of DCs reside in a variety of locations within an organism and thus have different timescales for antigen processing and presentation, it is also becoming clear that individual subsets may differ in the regulation of antigen processing.

Regulation of Dendritic cell maturation by E-cadherin/ ß -catenin signaling

Aimin Jiang, and Ira Mellman, Department of Cell Biology, Ludwig Institute for Cancer Research, Yale University School of Medicine, New Haven, CT 06520

As the sentinels of the immune system, immature dendritic cells (DCs) are distributed in peripheral tissues where they continuously sample the environment. To initiate an efficient immune response, however, DCs must undergo a process of terminal differentiation termed "maturation" prior, during, or following their migration to lymph nodes. It has long been observed that both human and murine DC cultures often form clusters, and the disruption of which results in their maturation, suggesting a coordination of maturation and migration (adhesion). Here we show that the murine bone marrow-derived CD11c+ DCs, also expressed the E-cadherin/catenin adhesion complexes. Disruption of the DC clusters leads to maturation, which can be inhibited when treated with anti-E-cadherin antibody, indicating that the E-cadherin adhesion system is also involved in DC maturation. Translocation of ß-catenin from membrane to cytosol is detected after cluster disruption of DCs, suggesting a possible role of ß-catenin for E-cadherin signaling. Accumulation of cytosolic ß-catenin and subsequent translocation into the nucleus has previously shown to regulate gene expression by transactivation of the TCF/LEF transcription family and could be responsible for the DC maturation program. Indeed, treatment with a pharmacological drug, SB216763, a specific inhibitor of GSK-3 b -which phosphorylates ß-catenin leading to degradation of ß-catenin, results in murine DC maturation by accumulation of cytosolic ß-catenin not bound to E-cadherin. Furthermore, DCs retrovirally transfected with either wild type or non-phosphorylation mutant (stabilized) ß-catenin-GFP exhibited spontaneous maturation as compared to non-transfected DCs, confirming the role of ß-catenin in DC maturation. Together, these results suggest the involvement of E-cadherin/ ß-catenin signaling pathway in DC maturation. Phosphorylation of I k B and p38 MAPK was induced upon LPS stimulation but not by cluster disruption, suggesting that E-cadherin/ ß-catenin singals DC maturation through a distinct mechanism than the TLR-mediated maturation process. Thus, our studies have identified the E-cadherin/catenin adhesion system as a novel pathway other than the TLR-mediated signaling that the DCs utilize for their maturation.

Respiratory Syncytial Virus (RSV) Affects Myeloid Dendritic Cell (mDC) Function

Michelle Gill1,2, John Connolly1, Yaming Xue1, Vikram Bhuta1,2, Octavio Ramilo1,2, and Jacques Banchereau1,2. Baylor Institute for Immunology Research1 and Division of Pediatric Infectious Diseases, UT Southwestern Medical Center2, Dallas, Texas.

Respiratory syncytial virus (RSV) is the leading respiratory pathogen in infants and young children worldwide. RSV infection does not induce long-term protective immunity and therefore repeated RSV infections occur throughout life. In addition to the considerable morbidity caused by acute RSV infections, numerous studies have described a strong association between RSV bronchiolitis in infancy and subsequent abnormal pulmonary function. Exaggeration of the immune response to RSV infection is thought to play a significant role in the development of these long-term pulmonary abnormalities. Despite decades of research, however, our understanding of the immune responses induced by RSV remains limited. The focus of our study is to investigate the effect of RSV on the immune system in the context of human myeloid dendritic cells (mDCs). Using human mDCs purified from the blood of adult donors, we have found that mDCs cultured with RSV secrete significantly greater amounts of IL-6, IL-8, IL-12p40, TNF-alpha, and MIP 1- alpha as compared to mDCs cultured with no virus or with influenza A. Phenotypic analysis of RSV-exposed mDCs revealed that although CD80, CD83, CD86, CD40, and HLA-ABC are upregulated on mDCs cultured with RSV, surface HLA-DR is not upregulated by RSV exposure on mDCs. Furthermore, the ability of mDCs to induce CD4+ and CD8+ T cell proliferation in an alloreaction is significantly reduced by RSV. We therefore conclude that RSV modifies the phenotype and function of mDCs and thus alters the development of T cell immune responses.

T cell-dependent IFN- g production by NK cells in response to influenza A virus

Xiao-Song He,1 Monia Draghi,1 Kutubuddin Mahmood,2 Tyson H. Holmes,1 George W. Kemble,2 Cornelia L. Dekker,1 Ann M. Arvin,1 Peter Parham,1 and Harry B. Greenberg1, 1Stanford University School of Medicine, Stanford, CA 94305 , 2Medimmune Vaccines, Mountain View, CA 94303

The interaction between the adaptive immune response and innate immune response during influenza A (fluA) infection is poorly understood. We have developed a cytokine flow cytometry assay to simultaneously investigate the IFN- g response of T cells and NK cells to fluA. When PBMC from fluA immune adult donors were incubated with fluA, IFN- g was produced by fluA-specific memory T cells as well as by both CD56dim and CD56bright subsets of NK cells. Purified NK cells did not produce IFN- g in response to fluA, while depletion of T cells abolished the IFN- g production by NK cells. In 25 donors tested, the frequency of IFN- g producing NK cells correlated significantly with the frequency of fluA-specific T cells. A subset of the fluA-specific IFN- g -producing T cells also produced IL-2. The fluA-induced IFN- g production of NK cells could be suppressed by anti-IL-2 antibody, while recombinant IL-2 replaced the helper function of T cells for IFN- g production by NK cells. These results indicate that IL-2 produced by fluA-specific T cells is involved in the T cell-dependent IFN- g response of NK cells to fluA. Taken together, our findings suggest that at an early stage of recurrent fluA infection, NK-mediated innate immunity to the virus could be enhanced by preexisting fluA-specific memory T cells.