8^th European Immunology Conference June 29-July 01, 2017 Madrid, Spain

Tabriz University of Medical Sciences, Tabriz, Iran

Introduction: Breast cancer is the second most common cancer in the world, as it accounts for 25% of the total cancers in women. The pursuit for a viral cause for human breast cancer has generated substantial disagreement. The aims of our study were to explore the presence of high – risk HPV type (16- 18) and HMLTV -DNA in breast tissue in a group of Iranian women with and without breast cancer. Material & method: Paraffin-embedded specimens from 65 cancerous breast cancer cases and 65 cases with benign breast lesions were examined for the presence of HPV and HMLTV -DNA by the nested polymerase chain reaction. Results: In our study all cancerous and non- cancerous breast samples were negative for the HMLTV –DNA, but HPV-6 was detected in 17 (26.2%) cases with breast cancer. This data is similar to reports that found no evidence of high-risk HPVs in breast cancer. Conclusion: The data presented in our study indicate a strong need for more epidemiological studies correlating different HPV types and HMLTV in Iranian women with breast cancer. Keywords: Human MMTV-Env like sequences, Breast cancer, Human papillomavirus.

Adriana Rodríguez Cruz burned in Mexico, 1989. Awarded bachelor's degree in Biology with honors by Universidad Nacional Autónoma de México (UNAM), 2012. Education abroad in 2010 at University of California Berkeley, U.S.A. Research internships in 2013 at University of Arizona, U.S.A., and in 2014 at Charité Universitäts Medizin Berlin, Germany. Currently studying PhD in Biomedical Sciences at UNAM with the project "Evaluation of the signaling pathway of macrophages CD3+ TCRαβ+/TCRαβ- and their function as pro-inflammatory macrophages"; to identify the mechanisms of activation and cellular function of these macrophages, as well as their implication in the physiopathology of pulmonary tuberculosis.

Background: It was reported that 5-15% of macrophages isolated from peripheral blood mononuclear cells (PBMC) express the TCRαβ receptor. Using monocyte derived macrophages (MDM), from healthy donors, and infected in vitro with BCG it was observed that the percentage of TCR+ macrophages increased in response to the infection (1). Preliminary results from our group, in mice, showed that after an intravenous BCG-infection there is an increase in the recruitment of two subpopulations of macrophages: CD11b+CD3+TCRαβ- and CD11b+CD3+TCRαβ+, at moment the characterization of these subpopulations has not been assessed. Methodology: PBMC were obtained from healthy donors, CD14+ cells were obtained through inmunomagnetic positive selection. After 7 days in culture MDM were obtained and characterized by flow cytometry. The MDM CD3+ TCRαβ+ and TCRαβ- subpopulations were evaluated for the coexpression of: CD80, CD86, CD11B, CD68, CD14, CD3, TCRαβ, TCRϒδ, HLA-I, HLA-II, CD1a, CD1b, CD1c, CD1d, CCR4, CCR7, CXCR1, CD16, and tmTNF. Findings: Our data shows that both of these subpopulations express efficiently the HLA-I, HLA-II. Moreover, the subpopulation CD3+TCRαβ+ showed an increase in the expression of CD1a, CD1b, CD1c and CD1d molecules, however in CD3+TCRαβ- this expression was absent. The expression of pro-inflammatory molecules CD16 and tmTNF had a stronger augment in the CD3+TCRαβ+ subpopulation. Chemokine receptors were measured and CCR4, CCR7 and CXCR1 were expressed 3 times more in CD3+TCRαβ+ compared to CD3+TCRαβ-. Conclusion: The CD3+TCRαβ+ MDM are efficient cells to present peptide antigens, however the expression of CD1 family molecules suggests that CD3+TCRαβ+ could play an important role in the presentation of lipid antigens. Probably, overexpression of pro-inflammatory molecules and chemokines receptors on CD3+TCRαβ+ is used by the MDM to favor a pro-inflammatory function. The phenotypic characterization of CD3+TCRαβ+ provides evidence that this subpopulation could be crucial in pathologies where lipids and inflammatory environment trigger a signal, such as tuberculosis.

Anne Schink earned her M.Sc. degree in Toxicology in 2012 from the Technical University, Kaiserslautern, Germany. Her thesis title was “Biomarkers to characterize and monitor skeletal muscle toxicity in the rat”. Afterwards, she held a position as consultant at the a-tune software AG, Darmstadt, Germany. In August 2015, she enrolled her Ph.D. studies and is actively included in the research work at the Max Planck Institute for Chemistry, Mainz, Germany and the Institute of Translational Immunology, University Medicine, Mainz, Germany. Her research focus is the identification of anti-inflammatory herbal extracts and their active compounds, which can alleviate different chronic diseases.

Acute and especially chronic inflammation underlies the development and progression of most severe diseases, such as inflammatory bowel disease (IBD), allergic asthma/chronic obstructive pulmonary disease (COPD), and organ fibrosis that leads to dysfunction and death. Regardless of the origin, inflammatory processes on the molecular level can be described as a cyclic events, involving innate immune activation, e.g. via Toll-like receptors (e.g. the TLR4-complex) and reactive oxygen species (ROS), and adaptive immunity directed to target tissue. Activation of TLR4 triggers secretion of pro-inflammatory cytokines and chemokines, which further activate these immune responses. To date, no effective orally active TLR4 antagonists are available for experimental or clinical application. The aim of our study was to identify anti-inflammatory herbal drugs and their active compounds that are stable in the gastrointestinal tract and preferably possess antagonistic TLR4 activity, or interfere with downstream inflammatory signaling pathways. To this aim, we screened more than 100 plant ethanolic extracts in TLR4-transfected reporter cells challenged with lipopolysaccharide (LPS) to identify their anti-inflammatory activity. Twenty-eight promising extracts were additionally confirmed in LPS-stimulated THP-1 monocytes for their dose-dependent anti-inflammatory activity. To detect TLR4 antagonistic activity or interference with the downstream inflammatory cascade, these extracts were also tested in TLR2- and TLR4-transfected HEK cell lines, which permits pathway differentiation, since both receptors share similar downstream signaling upon stimulation. Promising anti-inflammatory herbal extracts were fractionated by high-performance liquid chromatography-diode array detector (HPLC-DAD) technology and specific compounds in the active fractions were identified. Pure compounds were tested for TLR4 antagonistic activity in the cell culture systems described above. Select identified active compounds will be discussed.

Seong Kug EO’s lab has focused on unveiling how hosts response to pathogen infection. They have used various infectious models to prove host responses upon pathogenic infection. In recent, EO’s lab has found the detailed pathway that IFN-I signal pathway orchestrated environments to provide effective protection against mucosal viral infection (PLoS Pathog., 2016). Moreover, EO’s lab is expert on viral acute encephalitis caused by flaviviral infection. They have got many reports to unveil how immune system works on viral encephalitis caused by Japanese encephalitis virus (J. Neuroinflammation, 2014 and 2016).

Japanese encephalitis (JE), a neuroinflammation caused by zoonotic JE virus, is the major cause of viral encephalitis worldwide, and poses an increasing threat to global health and welfare. To date, however, there has been no report describing the regulation of JE progression using immunomodulatory tools for developing therapeutic strategies. We tested whether blocking the 4-1BB signaling pathway would regulate JE progression using murine JE model. Blocking the 4-1BB signaling pathway significantly increased resistance to JE and reduced viral burden in extraneural tissues and the CNS, rather than causing a detrimental effect. In addition, treatment with 4-1BB agonistic antibody exacerbated JE. Furthermore, JE amelioration and reduction of viral burden by blocking the 4-1BB signaling pathway was associated with an increased frequency of IFN-II-producing NK and CD4+ Th1 cells as well as increased infiltration of mature Ly-6Chi monocytes in the inflamed CNS. More interestingly, DCs and macrophages derived from 4-1BB KO mice showed potent and rapid IFN-I innate immune responses upon JEV infection, which was coupled to strong induction of PRRs (RIG-I, MDA5), transcription factors (IRF7), and antiviral ISG genes (ISG49, ISG54, ISG56). Further, the ablation of 4-1BB signaling enhanced IFN-I innate responses in neuron cells, which likely regulated viral spread in the CNS. Finally, we confirmed that blocking the 4-1BB signaling pathway in myeloid cells derived from hematopoietic stem cells (HSCs) played a dominant role in ameliorating JE. In support of this finding, HSC-derived leukocytes played a dominant role in generating the IFN-I innate responses in the host. Blocking the 4-1BB signaling pathway ameliorates JE via divergent enhancement of IFN-II-producing NK and CD4+ Th1 cells and mature Ly-6Chi monocyte infiltration, as well as an IFN-I innate response of myeloid-derived cells. Therefore, regulation of the 4-1BB signaling pathway with antibodies or inhibitors could be a valuable therapeutic strategy for the treatment of JE.

Sandra Sánchez has her expertise in molecular biology and bioinformatics, and is a passionate of biochemistry and science communication. Her novel approximation to the function of a drug on a bilayer membrane can help to better understanding of our murine models and how this can impact our knowledge on the systemic lupus erythematosus disease. She has built her abilities after years of experience in research, project administration and protocols development in the pharmaceutical industry, teaching of Biochemistry and Genetics for physicians, translation of pharmaceutical documents and participation in science fairs for kids. This work was possible due to her willingness to collaborate with external researchers, as Laura Domínguez from FQ-UNAM and Cornelio Yañez from CIC-IPN, who gave her the tools and knowledge to use bioinformatics and a Mexican supercomputer to realize this project.

Statement of the Problem: Systemic lupus erythematous (SLE) is an autoimmune, chronic and multifactorial disease. For SLE study, some animal models had been developed, and our lab has one. Our model was obtained by administering a different lipid bilayer structure called non-bilayer phospholipid arrangements. These non-bilayer phospholipid arrangements can be stabilized with drugs such as chlorpromazine or procainamide and specifically in female mice, causing a disease very similar to human SLE [1-3]. To help understand how these lipid structures in the development of the disease involved, we used bioinformatics tools to understand how they form, and now we’ll try to extrapolate this to explain some of the symptoms faced by patients with SLE. Methodology & Theoretical Orientation: A molecular simulation was established using all the conditions used in our murine model with chlorpromazine and was created with GROMACS software [4]. Analyses of results were made with GROMACS utilities, proprietary Python programs and VMD tools [5]. Findings: With our simulation strategy, we were able to observe differences between diverse lipidic environments with one or various molecules of chlorpromazine. Also, we have started new simulations to improve our strategy and get more useful information for our molecular knowledge of the models and the etiology of the human disease. Conclusion & Significance: These findings help us to understand how the models are triggered and give us clues about how to improve them for a further research of how SLE is initiated in humans. That probably will contribute to the improvement of the models and support our novel theory about the etiology of the SLE disease in humans.

Eduardo Martins de Sousa holds a bachelor's degree in Biomedicine, a Masters degree in Tropical Medicine area of concentration in Immunology by the Institute of Tropical Pathology and Public Health of the Federal Goiás University. PhD in Tropical Medicine area of concentration in Immunology by the Institute of Tropical Pathology and Public Health of the Goiás Federal University, being part of a sandwich doctorate held at the Institute of Molecular and Cellular Biology of the University of Porto - Portugal. Currently, Professor of the Postgraduate Program in Parasitary Biology (Masters degree) of the University Center of Maranhão (UNICEUMA). He is an Associate Professor of Post-Graduate Program in Biodiversity and Biotechnology of the Bionorte Network (PPG-BIONORTE) (Doctoral Level). He has experience in Immunology, with emphasis in Applied Immunology, working mainly on the following topics: Mycobacterium tuberculosis, Mycobacterium massiliense, ELISA, experimental infection, vaccine, flow cytometry, real-time PCR and mice.

Tuberculosis (TB) is a major cause of the global infectious disease-related morbidity and mortality. In 2014, 9.6 million new TB cases were estimated throughout the world, among which 1.5 million deaths occurred due to the infection. In the same year, the incidence of TB in Brazil was 44 cases per 100,000 inhabitants, placing Brazil 16th among the 22 countries comprising 80% of TB cases in the worldwide. The development of new tools for rapid and accurate diagnosis of active pulmonary tuberculosis (TB) is considered by many countries to be a strategy for controlling the disease. The aim of this study was to apply the enzyme-linked immunosorbent assay (ELISA) for measuring anti-CMX IgM antibodies in individuals with active pulmonary TB and healthy controls in an area endemic for TB. Analysis of the clinical signs and symptoms showed that most individuals with active pulmonary TB more frequently presented with a cough (94,7%), weight loss (83,4%), and, to a lesser extent, hemoptysis (37%). In addition, we evaluated the tests that were used to diagnose TB. The chest X-ray (100%) and sputum smear microscopy (80%) were the most frequently used tests for diagnosis; in contrast, only 33% of patients underwent computerized tomography, and microbiological culture was performed for 12%. Patients with active pulmonary TB had higher titers of anti-CMX IgM antibodies (optical density (OD = 0.502 ± 0.281; p = 0.0001) than healthy controls (OD = 0.200 ± 0.125). The cut-off for IgM-ELISA was determined using ROC curve analyzes (AUC=0.868) with a sensitivity of 80.1% and a specificity of 78.2%. These results suggest that the recombinant protein CMX can be used as a serological marker for screening individuals suspected of having active pulmonary TB.

Federal University of Bahia, Brazil

Rationale: IL-10 is an important regulatory cytokine with a protective role in allergies. This study aim to verify if IL-10 gene polymorphisms interferes in IL-10 cytokine production according to mite stimuli and atopy/asthma status. Methods: 1119 subjects from Salvador, Brazil, were genotyped using 2.5HumanOmni Beadchip from Illumina. IL-10 production by whole blood culture with the following mite stimuli: Blomia tropicalis and Dermatophagoides pteronyssinus was measured by ELISA. Asthma status was defined by ISAAC questionary. Atopy was determined through skin prick test to regional aeroallergens and specific IgE levels to B. tropicalis, D. pteronyssinus, Blatella germanica and Periplaneta americana. Statistical analyses were done using PLINK 1.9 and SPSS 22.1. Results: B. tropicalis was the mite with the higher frequency of sensitized individuals (34,26%) and the bigger frequency of IL-10 responders (93,1% against 21,3% to D. pteronyssinus). There was no association between asthma severity and B. tropicalis IL-10 induction. SPT ≥ 3mm for B. tropicalis had a positive correlation with IL-10 production in response to this stimulus (r = 0.126, p = 0.031). None genetic variant was associated with IL-10 production by B. tropicalis, furthermore one variant rs3024496 (C allele) was associated with greater skin reactivity (OR 1.33; 95% CI, 1.02-1.73). Conclusion: The absence of association between IL-10 polymorphism and this cytokine production can be explained by the lack of the promoter region of the gene on these analyses. Our findings did not support the IL-10 regulatory role once we did not find differences between IL-10 cytokine production according to atopy/asthma status, we otherwise found a positive correlation between IL-10 production and SPT positivity and an high frequency of IL-10 producers in B. tropicalis stimulus response. Now we question what IL-10 function in allergy caused by B. tropicalis, once this mite stimulus IL-10 production and there is association with more allergic biomarkers.

Alana Alcântara Galvão is graduated in pharmacy, Master in Interactive processes of the organs and systems by the Federal University of Bahia (UFBA) and a PhD student of Immunology at UFBA. Actually working in Laboratory of Allergy and Acarology (LAA), her research covers areas immuno-epidemiology, immunomodulation and immunogenetics. She collaborate with projects: Research on the prevalence of respiratory allergies and their risk factors in children from rural areas in the city of São Francisco do Conde, Bahia (2010); An asthma cohort in children and adolescents of the city of Salvador - Bahia, SCAALA (Social Change in Asthma and Allergy in Latin America) (2012); Immunogenetic study on possible associations between Vitamin D, atopy and asthma in children and adolescents of Salvador, Bahia (2014). The objective this study is identify the association of vitamin D with asthma and atopy, highlighting genetic variants in vitamin D pathway.

Introdution: Vitamin D presents a highlighted immune system activity. Their deficiency or insufficiency has been associated with low asthma control. Also, genetics determinants on the vitamin D pathway have been associated with asthma. In this study, we investigated associations between vitamin D serum levels with atopy and asthma, as well as polymorphism genes of the vitamin D pathway. Methods: 25 (OH) vitamin D was quantified from 970 of 11-17 years old Brazilians by ELISA. Asthma diagnosis was obtained by ISAAC, Phase III questionaries. Specific IgE detection to allergens was performed by ImmunoCAP; DNA was extracted and genotyped using 2.5 HumanOmni Biochip from Illumina. Statistical analyses were performed using PLINK 1.9 and SPSS 22.1 programs. Results: The prevalence of vitamin D insufficiency was 64% in 76 asthmatic and 62.5% in 446 atopic individuals, however there was no significant association between vitamin D and this outcomes. Negative correlation was found between vitamin D and specific IgE levels to Dermatophagoides pteronyssinus. on atopic subjects (r = -0.11, p =0.04). Genetic variants in CYP2R1 gene, rs7935792 (C allele) (Beta 1.66; 95% CI 0.20-3.11) and rs7129781 (C allele) (Beta 1.55; 95% CI 0.07-2.96), were associated with vitamin D serum levels. In addition, the same variants had suggestive protection on asthma, but it was not significant (OR 0.74; 95% CI 0.39;1.39; OR 0.73; 95% CI 0.38;1.37, respectively). VDR variants rs7965397 (G allele) was positively associated with atopy (OR 1.43; 95% CI, 1.07-1.92); rs4328262 (G allele) (OR 1.44; 95% CI 1.09-1.90) and asthma rs2408876 (C allele) (OR 2.31; 95% CI; 1.18-4.53); rs2238317 (T allele) (OR 2.19; 95% CI 1.02-4.72). Conclusions: Vitamin D can modulate the immune system and reduced allergic biomarkers. Genetic variants in CYP2R1 regulated vitamin D levels, and must prevent asthma symptoms. Genetic variants in VDR were associated with asthma and atopy susceptibility, maybe by modifying VDR expression.

Eduardo Martins de Sousa holds a bachelor's degree in Biomedicine, a Masters degree in Tropical Medicine area of concentration in Immunology by the Institute of Tropical Pathology and Public Health of the Federal Goiás University. PhD in Tropical Medicine area of concentration in Immunology by the Institute of Tropical Pathology and Public Health of the Goiás Federal University, being part of a sandwich doctorate held at the Institute of Molecular and Cellular Biology of the University of Porto - Portugal. Currently, Professor of the Postgraduate Program in Parasitary Biology (Masters degree) of the University Center of Maranhão (UNICEUMA). He is an Associate Professor of Post-Graduate Program in Biodiversity and Biotechnology of the Bionorte Network (PPG-BIONORTE) (Doctoral Level). He has experience in Immunology, with emphasis in Applied Immunology, working mainly on the following topics: Mycobacterium tuberculosis, Mycobacterium massiliense, ELISA, experimental infection, vaccine, flow cytometry, real-time PCR and mice.

Infectious diseases continue to be one of the biggest health problems in the world, affecting millions of people annually. M. abscessus and other species of rapidly growing mycobacteria (RGM) are naturally resistant to antimicrobial compounds and disinfectants because they have an impermeable cell wall composed by peptideoglycan and mycolic acids. These RGM are responsible for various hospital outbreaks worldwide, causing lung infections in patients with cystic fibrosis, chronic lung disease (bronchiectasis, nodules and cavitations), post-surgical infections, and skin and soft tissue infections in immunocompromised patients. The resistance of M. abscessus (Mabs) to the medications used in current therapy challenges the search for new treatment strategies. Previous studies on the search for new natural compounds with antimicrobial action highlighted the potential of Bixa orellana (urucum). The seeds of this plant are already used in folk medicine for treating heart disease, gastrointestinal problems, respiratory infections. In this study, we evaluated potential anti-inflammatory activities of hydroalcoholic (BoEH) and ethyl acetate (BoEA) extracts of B. orellana leaves, using a murine model of peritonitis induced by heat killed Mabs. C57BL/6 mice were orally treated with different concentrations of BoEH or BoEA. After one hour, peritonitis was induced by inoculation of 1x108 CFU of heat killed Mabs. BoEH and BoEA inhibited the migration of total leukocytes (Figure 1A - B), migration of polymorphonuclear cells (Figure 1C - D) and mononuclear cells (Figure 1E - F) into the peritoneum in the periods analyzed 4 and 24 hours after the induction of peritonitis. Our results suggest anti-inflammatory actions of the extracts tested, indicating this plant as natural source of compounds with potential for pharmacological and biotechnological applications.

Dr Yap Chui Sun joined the Department of Clinical Translational Research, Singapore General Hospital (SGH) in 2013, and has been working on reprogramming mesenchymal stem cells. Her first postdoctoral position was at Duke-NUS Medical School, where she was studying the contribution of micro RNAs on thyroid hormone function. She obtained her Master’s degree in Molecular Immunology at the National University of Singapore and Ph.D. degree in Molecular Oncology at Brown University (U.S.A.).

Mesenchymal stromal cell (MSC) therapy has been shown to be effective in phase I/II clinical trials in the treatment of graft versus host disease (GVHD) after allogeneic hematopoietic cell transplantations. However, MSC trials still face major challenges, such as complex and time-consuming manipulation, requiring a good manufacturing practice facility, difficult and expensive to produce etc. In a screen of MSC-derived factors with serial factorial designs, we first time identified two MSC-derived factors, CXCL5 and CCL24 inhibitor (antibody), which exhibited synergistic immunomodulation effect in mixed lymphocyte reaction. This two-factor (2F) cocktail also showed excellent in vivo immunosuppressive effect in ameliorating GVHD symptoms and improving survival. A xenograft GVHD animal model was created by injecting 400×106cells/kg of cryopreserved human PBMCs into NSG mice respectively. Four consecutive treatments were given on day-10, day-14, day-17 and day-21 post-transplantation. The 2F cocktail exhibited excellent immunosuppressive effect as it could improve mice 36-day survival from 19.0% with severe symptoms to 61.9% with mild symptoms (p<0.01). It was significantly better than BM-MSCs (8.3%, p<0.001) and Cyclosporine A (26.1%, p<0.05). Synergistic effect was again observed between those two factors (CXCL5, 18.2%; anti-CCL24, 9.1%; p<0.05). The 2F cocktail treatment reduced cytotoxic T lymphocytes (CTLs), T helper 1 (Th1) cells, Th17 cells, NK cells in the circulation and macrophages in the spleen, but did not affect human hematopoietic stem cells (HSCs) reconstitution in the bone marrow. Concurrently, it reduced pro-inflammatory cytokine IFN-γ, IL-1β, IL-6, IL-12, TNF-α, IL-17A, IL-8, MIP-1β and MCP-1 in the circulation. In conclusion, the efficacy of a novel identified 2F cocktail was validated in an in vivo xenograft GVHD model. It demonstrated a robust immunosuppressive effect and kept the development of GVHD under control. The 2F cocktail could be a potential chemically defined substitute for MSCs in GVHD therapy.

Emilia Andrade Belitardo is physiotherapist, specialist in respiratory physiotherapy by the Federal University of São Paulo and Master in Immunology by the Federal University of Bahia (UFBA). She is currently a PhD student of the Immunology at UFBA, working in the Laboratory of Allergy and Acarology (LAA). Her research interests include immuno-epidemiology, clinical immunology, immunomodulation and immunogenetics. She collaborates with two major research projects: (i) an asthma cohort in children and adolescents of the city of Salvador - Bahia, SCAALA (Social Change in Asthma and Allergy in Latin America); and (ii) a case-control study developed in partnership with the Center of Excellence for Asthma (UFBA). The aim of this study is to investigate biomarkers for understanding the different asthma phenotypes. E-mail: emilia_mandrade@hotmail.com

Rationale: Asthma is a chronic and heterogeneous disease presenting various phenotypes. The aim of this study was to assess airway inflammation among subjects with asthma by counting cells and measuring cytokines in the sputum to search for associations with clinical features of the disease. Methods: We studied 66 subjects, divided into three asthma subgroups [16 with severe asthma resistant to treatment (SAR), 22 with severe asthma controlled with treatment (SAC), 19 with mild to moderate asthma (MMA)] and a group with no asthma (NA) including 9 subjects. Total cellularity of the sputum samples was counted using a hemocytometer and differential cytology was observed in cytospin. Measurements of cytokines were performed by Luminex (Upstate / Millipore system “Flex kit”). Statistical analysis was performed using nonparametric tests. Results: Sputum of SAR had higher percentage of neutrophils as compared with MMA (p = 0.05) and higher percentage of eosinophils compared with NA (p = 0.02). TNF was increased in SAR compared to NA, MMA and SAC (p=0.001). Subjects with asthma treated with high doses of inhaled corticosteroids presented higher levels of TNF (p = 0.02), IL-6 (p = 0.01) and IL-5 (p = 0.03). Increased TNF production was associated with reduced lung function before and after bronchodilator as measured by FEV1% [p = 0.00 for both), FEV1/FVC% (p = 0.03 and p=0.02 respectively) and FEF25-75% (p = 0.00 for both). Conclusion: Increased inflammatory cytokines (TNF, IL-6 and IL-5) and number of inflammatory cells (neutrophils and eosinophils), were associated with SAR, and high levels of TNF were associated with worse lung function. Our findings support the concept these subjects may have an end phenotype of asthma related to airway inflammation that goes beyond the Th2 type response, and that it is indeed resistant to high doses inhaled corticosteroids, requiring a different approach to treatment.

As a post Doc my focus was to study therapeutic strategies towards successful xenotransplantation. I was involved in two main projects (supported by the government) related to the development of the first pre-clinical nonhuman primate study of solid organ xenotransplantation in Korea. This was done using genetically engineered pigs expressing multiple human complement and coagulation regulatory proteins in order to overcome the immunological and physiological barriers against successful xenotransplantation.

Introduction: In kidney, heart and islet transplantation the Rhesus monkey (Macaca mulatta, RM) has been shown to be an excellent preclinical model that can provide the basis for new immunosuppressive protocols for clinical studies. However, there remain relatively few liver transplant (LT) models in nonhuman primates. In this study, we analyzed the immune cell populations of PBMC and secondary lymphoid organs along with livers of normal rhesus monkeys and compared them to those of rejecting liver transplanted recipient’s following withdrawal of immunosuppression. Methods & Results: We undertook six allogeneic ABO compatible orthotopic LT in monkeys using six normal donor monkey livers. We collected tissues including lymph-node, spleen, and blood from which we isolated immune cells for FACS analysis along with the liver from the recipient. We found that CD4 or CD8 naïve T cells were normally seen at low levels (13.89±8.67 or 1.50±1.44 respectively) and memory T cells were seen at high levels (76.12±11.40 or 98.0±1.60) in the liver rather than lymphoid organs or PBMC. However, regulatory cells such as CD4+FoxP-3+ T cells and CD8+CD28- cells remained in high numbers (0.77±0.54 and 34.99±6.40) in the liver but not in lymph node or PBMC. These result demonstrate that the liver has rather unique immunological properties compared to other organs. We also compared CD4/8 T sub-populations in normal or rejected livers and the various tissues showed that naïve cells were dramatically decreased in spleen, lymph node and PBMC of rejected transplanted monkeys but rather their memory cells were increased in all tissues and PBMC. Conclusion: We have shown that the normal liver has large numbers of C4Tregs or CD8+CD28- or MDSC which are known immune suppressive cells at much higher levels than other lymph node or peripheral blood. Memory T cell populations in rejected livers or lymphoid organs were expressed at significantly higher levels than those seen in normal tissues including as seen in the peripheral blood.

Gayane Manukyan is researcher in the National Academy of Sciences of the Republic of Armenia and a group leader of a Group of Molecular and Cellular Immunology Institute of Molecular Biology. Currently she is postdoctoral fellow in the Department of Immunology, Medical Faculty, Palacky University, Olomouc, Czech Republic. She has strong expertise in flow cytometry and studies focused on innate immunity and imunology of inflamatory and infectious diseases.

Besides a classical role in antimicrobial functions, emerging evidence indicates that neutrophils could have an effect on chronic and progressive diseases such as leukemias. Nowadays, there is limited information about the function of circulating neutrophils in the chronic lymphocytic leukemia (CLL). The aim of the present study was to study functional properties of circulating neutrophils in CLL. Eighteen CLL patients and 17 healthy controls were enrolled in the study. Priming of neutrophils with LPS as well as oxidative stress capacity were analyzed using flow cytometry. Spontaneous and induced with fMLP and PMA oxidative stress was measured using dihydrorhodamine 123. Despite fMLP and PMA significantly induced ROS production by neutrophils in both healthy (P<0.01) and CLL groups (P<0.01 and P<0.001, respectively), stimulation with both inducers has led to a maximum ROS-release in neutrophils from CLL patients compared to healthy ones (P<0.05 and P<0.01, respectively). Spontaneous production of ROS by CLL neutrophils was also increased (P<0.05) compared with healthy neutrophils. LPS up-regulated TLR2 in healthy cells (P<0.05), while TLR2 expression was down-regulated in CLL cells (P<0.05). LPS exposure of isolated neutrophils from healthy group induced production of IL-1β and TNF-α (P<0.05). In contrast, LPS-stimulated CLL neutrophils failed to induce releasing of both cytokines. LPS-induced production of IL-1β and TNF-α in CLL group was lower than those released by neutrophils from healthy group (P<0.05). Taken together, circulating neutrophils in CLL patients have altered functional properties of which may account for the heightened sensitivity to bacterial infection as well as influence the disease course. Future studies are needed to prove our observations. Grant support: MZ ČR VES16-32339A, IGA_LF_2017_009.

Gabriela Gabcova is a PhD student of Immunology at Palacky University Olomouc, Czech Republic. Her main research activity includes immunophenotyping of cells in peripheral blood and synovial fluids by flow cytometry. She has experience with development of multi-colours panels for flow cytometry. She is primarily focused on disorders of connective tissue (osteoarthritis, rheumatoid arthritis) and hematological malignancies (chronic lymphocytic leukemia, Hodgkin lymphoma).

Chronic lymphocytic leukemia (CLL) cell is characterized by a progressive accumulation of long-lived CD5+ B lymphocytes in bone marrow/lymph nodes, whose survival requires exogenous activation signals such as chemokines. Growing number of studies show that chemokines and their receptors, in addition to trafficking, have much broader influence on neoplastic cells, such as cell growth, differentiation and survival. To gain more insights into the chemokine receptor network in CLL, we characterized the expression pattern of 16 canonical and 4 atypical chemokine receptors in peripheral blood mononuclear cells (PBMC) of CLL patients (n=88) and healthy subjects (n=34) by using quantitative RT-PCR. The expression of CXCR3, CXCR4, CXCR5, CXCR7, and CCR7 was confirmed by 6-color flow cytometry. Among deregulated receptors, 5 receptors (CCR7, CCR10, CXCR3, CXCR4, CXCR5) were up-regulated and 9 receptors (CCR2-CCR6, CCR8, CCRL2, CXCR1, CXCR2) down-regulated in CLL; the expression of others did not differ between CLL and controls (P>0.05). We have also analyzed differences in expression pattern in CLL groups subdivided according cytogenetics (13q and 17q deletions). In patients with del(17q) having worse prognosis, we observed higher mRNA levels of CXCR6, CXCR7 and CCR10 comparing to del(13q) associated with good prognosis. In conclusion, differential expression patterns of chemokine receptors suggest the relevance of the network of these receptors in CLL pathogenesis. The potential of chemokine/chemokine receptor network as determinant for clinical outcome and novel therapies is worth exploring. Grant support: MZ ÄŒR VES16-32339A, IGA_LF_2017_009.

Guan-Ming Ke is a licensed veterinarian and teaches at the National Pingtung University of Science & Technology as an associate professor. He focuses on animal vaccine development and production, continuously applying new technologies to develop new types of vaccines and to better the manufacturing process. His work aims to turn research into applicable technologies that will aid in disease prevention and improve both animal and human welfare.

Newcastle disease virus (NDV), also known as avian paramyxovirus serotype 1, is a member of the family Paramyxoviridae and causes a highly contagious respiratory, neurological, or enteric disease in chickens. Currently, genotype VII strains are the predominant virulent strains. Although the different genotypes of NDV all belong to one serotype, it is still difficult to confer cross-protection under stressful environmental conditions. The aim of this work is to generate an attenuated NDV strain and develop a cell culture-derived genotype VII ND inactivate vaccine against current virulent NDV strains. After the 40th passage, we found that the MDT of NDV strains is higher than 120 hours, the intracerebral pathogenicity indexes (ICPI) had decreased to 0-0.08, and TCID50 has reached 108. The attenuated strain, KGM-01, is then inactivated and mixed with oil adjuvants for vaccine evaluation. Hemagglutination inhibition test showed that the inactivated vaccine elicited high antibody titer two weeks after immunization and no virus shedding was detected using real-time PCR when challenged with the Sato strain and field type VII strains. Thus, KGM-01 is a low virulent, type VII genotype strain with

Chi-Chi WEN holds a master’s degree in veterinary medicine from National Taiwan University. She is a research fellow at Graduate Institute of Animal Vaccine Technology of National Pingtung University of Science and Technology. Her research and interest include molecular biology and biotechnology, continuing in the research of veterinary medicine. She focuses on development of animal vaccines, designs new type of vaccines with biotechnology, with the hope to help animals avoid suffering.

The foot and mouth disease (FMD) is a highly contagious viral disease of Artiodactyla, causing severe economy lost worldwide. In this study, the structural protein VP1, VP2, VP3 and VP4 of FMD serotype O were produced with the concentration between 250ug/mL to 550ug/mL by using baculovirus expression vector system (BEVS), which improved the low productivity problem of eukaryotic expression system. The concentration of the anti-FMD antibodies in animal sera can be distinguished by using purified VP1 proteins, which were coated on the ELISA plates, showing that the recombinant VP1 protein has good antigenicity. The virus like particles (VLPs) was observed under electric microscope by using the mixture of the four proteins and pH value adjustment. Presently, those four structural proteins were tested alone or in mixture in pig experiment. In conclusion, this subunit vaccine has the potential to provide protection against FMD.

I am Master Student of the University of Costa Rica; I am developing a research project on different Immunology aspects regarding the infectious disease called Brucellosis. My work's main objective is to study the role of polymorphonuclear neutrophils as “Trojan Horse” vehicles during brucellosis using a bone marrow murine model and the chronicity and persistence of the disease. During my work, I have demonstrated effort, dedication and a very positive attitude toward research and academic work. I have acquired expertise in elisas, flow cytometry, bacterial infections, cell culture, cell differentiation, cell infection through cells, fluorescence microscopy. I am co-author of a publication (I & I 84: 1712) regarding the role of neutrophils during brucellosis.

Brucella abortus is a facultative intracellular pathogen that causes chronic infections. Neutrophils (PMNs) are the first cells that encounter Brucella after invasion, however, Brucella resist their killing action and induce premature cell death of these leukocytes. It has been described that B. abortus persist in bone marrow at chronic stages of infection. Nevertheless, the role of PMNs in bone marrow persistence has not been studied. Here we show that B. abortus organisms are able to persist in murine bone marrow even at the “declining stages of chronic infection”. B abortus were observed inside a PMN/monocyte cell type at very low rates. Additionally, we demonstrate that murine bone marrow PMNs phagocyte antibody-opsonized B abortus and die quickly after infection. These dying infected PMNs show increased adhesion and are readily taken up by RAW 264.7 macrophages. When ex vivo macrophage infections were performed, B. abortus were more infective and replicated at higher rates when macrophages were infected through PMNs than when infected with Brucella only. In general, proinflammatory and anti-inflammatory cytokines showed increased values in macrophages infected through PMNs but only after 24 hours of infection, when Brucella has already reached their replication niche inside the cell. Our results, support the notion that infected bone marrow PMN might behave as vectors for Brucella persistence in bone marrow in a non-logistic way.

Mansoura University, Egypt

Statement of the problem: Rheumatoid arthritis (RA) is a multi-organ inflammatory disorder. A reduction in life expectancy in RA patients is primarily due to myocardial disease which is clinically silent. There is increasing interest in autoimmune diseases, especially their relationship with cardiovascular disease. RA in particular has been considered an independent risk factor for coronary artery disease in recent years. Various studies have aimed to clarify important aspects of risk stratification and treatment options in patients with rheumatoid arthritis. TDE offers the promise of an objective measure to quantify regional and global LV function. So, it is important for using a non-invasive technique and can be serially followed over time for cardiac risk progression and detecting patients at the greatest risk of cardiac morbidity and mortality Methodology& theoretical orientation: case control design that has been carried out on rheumatoid arthritis (RA) patients and control. All patients evaluated clinically and echocardiographically (M mode, trans-mitral and tissue Doppler). and all echo-parameters were correlated with various clinical data. Findings: Sensitivity of tissue Echo compared to conventional echo in diagnosis of diastolic dysfunction in RA patients is weak. Tissue Doppler finding were not related to DAS28CRP. Tei index showed significant positive correlation with disease duration. Conclusion & significance: Diagnostic accuracy of Tie index by tissue echocardiography is weak diagnostic. Myocardial dysfunction in RA is a matter of time and not related to disease activity.

Vaccine & Infectious Disease Organization-International Vaccine Centre, University of Saskatchewan, Saskatchewan, Canada

Statement of the Problem: The microorganism Nessieria gonorrhoeae is an obligate human pathogen and its adherence to host cells is essential for its pathogenesis. We devised a flow cytometry-based method to quantify the adherence of piliated N. gonorrhoeae strain F62 to human cervical ME-180 cells. Methodology: Piliated N. gonorrhoeae F62 were collected after 10 to 12 hours of growth then stained with cell-permeable fluorescent dye 5′-carboxyfluoroscein succidyl ester (CFSE). The bacteria were incubated with 0.5 μl of 5 mM CFSE in 2.5 ml of PBS and incubated at 37 °C for 15 min. ME-180 cells were incubated for 2 hours with fluorescent, piliated N. gonorrhoeae (multiplicity of the infection 1:100) then the ME-180 cells were washed with phosphate buffer saline to remove loosely adherent bacteria. Flow cytometry was used to quantify the percentage of ME-180 associated with CFSE+ fluorescent bacteria and a minimum of 30,000 events were recorded. Finding: Results indicated that 19.2% +/- 0.99 (n=4) ME-180 cells were associated with the fluorescent, piliated bacteria. To assess whether antibodies specific for N. gonorrhoeae blocked their adherence to ME-180 cells, rabbit hyper-immune anti serum was raised against heat-killed piliated N. gonorrhoeae F62. Adherence Efficiency, the percentage of cell-associated CFSE+ bacteria divided by the total input CFSE+ bacteria ranged between 37-47% (n=5). Heat-inactivated hyperimmune serum, at 1:10 to 1:80 dilutions, significantly inhibited gonococcal adherence by 6 and 3 fold, respectively. Heat-inactivated negative rabbit serum was significantly (3 to 5 folds) less effective at preventing bacterial adherence suggesting that antibody specificity and not a non-specific serum component was involved. Flow cytometric analysis was amenable to the quick, easy and high-throughput quantification of N. gonorroheae association with eukaryotic cells. These approaches may be adapted for use in in vitro and in vivo adherence studies related to gonococcal pathogenesis.

Kira Ziegler obtained her Diploma in Biology form the Johannes Gutenberg-University, Mainz, Germany in 2015. The topic of her Thesis was “Expression and recovery of two recombinant proteins in Escherichia coli for cancer vaccination”. Following this, she started her PhD thesis at the Max Planck Institute for Chemistry, Mainz, Germany, in the same year. Her research focus is the allergenic effect of nitration on wheat derived alpha amylase trypsin inhibitors, bridging with this topic atmospheric science and fundamental medical research.

Over the past decades environmental pollution and allergy incidence have been increasing on a global scale, implicating that they are interconnected. Air pollutants e.g. nitrogen dioxide and ozone are capable of chemically modifying airborne allergens, particularly under humid summer smog conditions. As demonstrated for birch pollen Betv1, these nitrated allergens are known to have enhanced allergic potential. We demonstrated that wheat derived alpha-amylase-trypsin inhibitors (ATIs), which previously were identified as major allergens of baker’s asthma, are also strong activators of the intestinal innate immune system when ingested with wheat products. Moreover, via their innate immune stimulatory activity these ATIs also promote experimental allergies. As air pollution and fertilizers can lead to nitration of these ATIs in the living grains, the aim of this project was to elucidate the effect of nitrated ATIs on innate and adaptive immunity. Therefore, a HeLa TLR4 dual reporter cell line was stimulated with untreated ATIs vs nitrated ATIs. Furthermore, human monocyte-derived dendritic cells (DC) were exposed to ATIs or nitrated ATIs and changes in specific DC maturation markers and cytokine patterns were analyzed by flow cytometry or multiplex ELISA. Additionally, T cell proliferation after co-cultivation with different ATI-treated autologous DC was determined. In all these different in vitro systems we could demonstrate a stronger stimulatory capacity of nitrated ATIs in comparison to native ATIs, indicating that nitration of an antigen/allergen not only affects its allergenicity but also its immunogenicity.

Landa Carla: I am a PhD student. Now, I´m working with a murine lupus model induced by lipidic antigens. We are interested in the immune response, especially in the participation of NKT cells and plasmacytoid dendritic cells (pDC) in this autoimmune disease. We had found an increase in the percentage of NKT cells y pDC in secondary lymphoid organs at 10, 20 and 30 days after being injected in mice with non-bilayer phospholipid arrangements and we already know that these antibodies are produced via germinal centers in this murine model that resembles human lupus.

Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease where animal models are used to study its pathogenesis. We have developed a mouse model of autoimmune disease resembling human lupus by the injection of liposomes with non-bilayer phospholipid arrangements (NPA). Also we described the presence of IgG antibodies against NPA in the serum of mice with lupus and patients with SLE. In this work, we determined by citofluorometry the presence of plasmacytoid dendritic cells (pDC) the main producer of type I interferons, and of NKT cells in the secondary lymphoid organs of mice 30 and 60 days after the injection of liposomes with or without NPA induced with 8 mM promazine. In both groups of mice injected with liposomes bearing NPA a significant increase of pDC cells (5-fold) was found which correlates with the high concentration of type I interferon previously detected in mice with lupus and in human SLE. A significant increase of NKT (3-fold) was detected at 30 days, specifically in the NKT subpopulation CD4+ which is known to cooperate with B cells in response to lipid antigens; this increase suggests its probable involvement in adaptive immune responses which lead to the production of anti-NPA IgG antibodies. The increase of pDC and NKT cells found by cytometry in secondary lymphoid organs in this work, suggests their involvement in the formation of anti-NPA IgG antibodies and the development of the disease resembling human lupus.

Jong-Yeup Kim has his expertise in evaluation and passion in improving the Otorhinolaryngology (Snoring, septic disease, sinusitis, tonsillitis, nasal molding, and allergic rhinitis). His open and contextual evaluation model based on responsive constructivists creates new pathways for improving nose disease. He has built this model after years of experience in research, evaluation, teaching and administration both in hospital and education institutions.

Predicting which patients are at a higher risk for recurrent chronic rhinosinusitis with nasal polyps (CRSwNP) is one of the most challenging problems in clinical rhinology. A direct association between nasal polyp and eosinophil/neutrophil counts was reported. This study aimed to identify difference of eosinophils and neutrophils for formation polyp by DNA methylation in CRS. We have previously shown that KRT 19, NR2F2, ADAMTS1, and ZNF222 levels are changed in nasal polyps (NPs) of patients with chronic rhinosinusitis (CRS) in patients. A study was performed from 30 patients with CRS with bilateral NP examining the prognostic role of eosinophil and neutrophil levels. 30 patients with CRS were classified by the rate of eosinophils and neutrophils in tissue. The methylated genes detected by DNA methylation microarray were validated by methylation-specific polymerase chain reaction (PCR), bisulfite sequencing, and real-time PCR. DNA methylation microarray identified 43,674 CpG islands in 518 genes. Specific genes were found to have a hypermethylated signal, and some genes were significantly hypomethylated in the promoter region in eosinophils compared with neutrophils. Real-time PCR showed that the expression levels of gene were changed in eosinophils compared with neutrophils. We clearly demonstrated that all two subgroups of CRSwNP had characteristic differences in DNA methylation, which allows for pathophysiologically meaningful differentiations with likely therapeutic consequences. Further studies are needed to confirm the significance of these epigenetic factors in the mechanisms underlying NP formation.

Simona Valova studied Molecular Biology and Genetics at Faculty of Science, Masaryk University, Brno, the Czech Republic. She is currently in her third year of Ph.D. studies in Physiology and Pathological Physiology at Faculty of Medicine, Masaryk University. She works in the team of Prof. Lydie Izakovicova Holla, that focuses on variability in candidate genes for multifactorial diseases, including periodontitis, recurrent aphthous stomatitis, diabetes mellitus or gastroesophageal reflux disease.

Statement of the Problem: Recurrent aphthous stomatitis (RAS) is a multifactorial disease with an unclear etiopathogenesis, resulting from the interplay between genetic and environmental factors.1 As the dysregulation of the immune system can play a role in the RAS development2, single nucleotide polymorphisms (SNPs) in the genes for immune and inflammatory molecules were studied.3,4 The NOD-like receptor (NLRP3) gene, encoding the component of the inflammasome, has been proposed as one of the candidate genes for RAS.5 The aim of our study was to investigate three SNPs (rs4612666, rs10754558, rs3806265) in NLRP3 gene in patients with RAS and healthy controls in the Caucasian population Methodology: A total of 200 Czech subjects were enrolled in this case-control study. 143 healthy controls, 57 patients with RAS were genotyped by method based on polymerase chain reaction using 5′ nuclease TaqMan® assays. Clinical parameters such as complete blood count, levels of immunoglobulins including allergen-specific immunoglobulin E or presence of antibodies against cytomegalovirus, Epstein-Barr virus were determined in RAS patients. Findings: Although no significant differences in the NLRP3 (rs10754558, rs3806265) allele and genotype frequencies between patients with RAS and controls were observed, statistically significant differences in NLRP3 rs4612666 genotype frequencies between subjects with RAS and controls were found. Carriers of NLRP3 rs4612666 TT genotype had a higher risk of developing RAS in comparison to subjects with CT + CC genotypes (OR = 16.71, 95% CI = 1.96-142.14, P = 0.0024). No association between NLRP3 haplotypes and RAS was detected. Conclusion & Significance: In contrast to the previous study5, associations between NLRP3 (rs10754558, rs3806265) polymorphisms and RAS were not confirmed. However, we suggest that NLRP3 rs4612666 polymorphism can strongly influence the risk of developing RAS in the Czech population. Acknowledgements: This study was supported by the grants AZV 15-29336A, GACR GB14-37368G and project MUNI/A/0948/2016.

Seung-Heon Hong works as a Professor at the Department of Oriental Pharmacy, College of Pharmacy, Wonkwang University, Iksan, Korea. Since 2005, Hong has been an Editor of 'Oriental Pharmacy and Experimental Medicine' and an Editorial Board of 'Evidence-based Complementary and Alternative Medicine'. His research interest is to investigate pharmacological effect of herbal medicine on cancer, allergic inflammation, and obesity.

Lipins are phosphatidic acid phosphatases involved in synthesis of phospholipids and triglycerides., Although they regulate cellular levels of important signaling lipids. Lipin-1 contributes positively to macrophage stimulation through TLR4, and other TLRs, by affecting MAPKs and AP-1 activation and, as a consequence, the generation of pro-inflammatory factors. Lipin-2 reduces pro-inflammatory signaling induced by saturated fatty acids in macrophages. Here we examined whether LPIN2 mRNA inhibition affects TLR mediated inflammatory signaling in HT29, a colon cancer cell line. The LPIN2 siRNA pre-treatment reduced the up-regulated defensins stimulated by TLR ligands, LPS and flagellin. And the increased level of IL-8 mRNA by LPS and R848 were more increased by LPIN2 mRNA inhibition. And LPS and R848 induced JNK and ERK phosphorylation whose expressions were more elevated by Lipin2 inhibition. On the other hand, the lipid transcription factors like PPARγ and PGC1α did not changed by LPIN2 siRNA pre-treatment. Taken together, LPIN2 inhibition aggravates TLR ligands induced inflammatory signaling through ERK and JNK phosphorylation.

Immune regulation has important roles in various immune diseases. The authors have studied the regulatory mechanisms and function of TIM-3 in various cells and in an in vivo tumor model. The authors revealed the involvement of MEK and c-jun in TIM-3 expression by CD4+ T cells. Additionally, they reported that the efficacy of tumor vaccine can be up-regulated by TIM-3 pathway blockade and the IL-2 production is decreased in CD4+ T cells expressing TIM-3 through NFAT dephosphorylation and AP-1 transcription.

T cell immunoglobulin- and mucin-domain-containing molecule-3 (TIM-3) is well known as one of the immune check point molecules. TIM-3 expression is increased on exhausted T cells and senescent T cells in numerous immune diseases including cancers. However, the regulatory mechanisms of TIM-3 expression in cancers have not been well studied. Using Jurkat T cells, we examined TIM-3 regulatory mechanisms in condition similar to tumor microenvironment. TIM-3 mRNA and protein levels were increased by co-culture of Jurkat T cells with tumor cell lines and by incubation of them in tumor cell conditioned media. Given that cyclic adenosine monophosphate (cAMP) can be transferred from tumor cells to T cells, we examined the effect of cAMP signaling on TIM-3 expression. It was promoted by intracellular elevation of cAMP concentration and activation of cAMP downstream pathways. Further, inhibition of cAMP downstream pathway attenuated TIM-3 expression in Jurkat T cells cultured in tumor-CM as well as in Jurkat T cells stimulated with a cAMP elevating agent. Conclusively, this study suggests that TIM-3 expression in Jurkat T cells may be induced by tumor CM through activation of cAMP pathway.

Young Sang Kim, a professor in Biochemistry department in Chungnam National University, finished his PhD at University of Illinois at Chicago in 1990 and continued postdoctoral research at Yale University for 2 years. His research interests focus to develop a strategy for selective activation of tumor associated antigen(TAA)-specific cytotoxic T lymphocytes. He evaluates anti-tumor effect of tumor cell vaccines engineered to express cytokines on tumor cell surface as a membrane-bound form instead secretory form. In this way he expects the membrane-bound form of cytokine on tumor cells may function as a co-stimulatory molecule to TAA-specific cytotoxic T lymphocytes. He published more than 70 scientific papers last 20 years.

Interleukin-17A is a member of the IL-17 family, and is known as CTLA8 in the mouse. It is produced by T lymphocytes and NK cells and has proinflammatory roles, inducing cytokine and chemokine production. However, its role in tumor biology remains controversial. We investigated the effects of locally produced IL-17A by transferring the gene encoding it into mouse tumor cells including B16 melanoma, and MethA fibrosarcoma, either in a secretory or a membrane-bound form. Expression of the membrane-bound form on CT26 colon cancer cells dramatically enhanced their proliferation in vitro. The enhanced growth was shown to be due to an increased rate of cell cycle progression: after synchronizing cells by adding and withdrawing colcemid, the rate of cell cycle progression in the cells expressing the membrane-bound form of IL-17A was much faster than that of the control cells. Both secretory and membrane-bound IL-17A induced the expression of Sca-1 on the cancer cells, which is commonly associated with aggressive phenotype of cancer cells. When tumor clones were grafted into syngeneic BALB/c mice, the tumor clones expressing the membrane-bound form IL-17A grew rapidly; those expressing the secretory form also grew faster than the wild type CT26 cells, but slower than the clones expressing the membrane-bound form. These results indicate that IL-17A promotes tumorigenicity, in part, by enhancing cell cycle progression. This finding should be considered in treating tumors and immune-related diseases.

Sung-Joo Park is a professor of Wonkwang University in South Korea, and majored in Korean traditional medicines and immunology. Prof.Park has many focuses on inflammatory diseases such as pancreatitis, sepsis, obesity, and asthma. Prof.Park has many experimental models and molecule detection techniques to examine the pathophysiology and possible drug of the diseases. Recently, Prof.Park mostly studied about acute pancreatitis and sepsis and reported many papers about them.

Guggulsterone (GS) is a phytosterol that has been used to treat inflammatory diseases such as colitis, obesity, and thrombosis. Although many previous studies have examined the anti-inflammatory activities of GS, the specific effect and mechanism of GS on lipopolysaccharide (LPS)-induced inflammation and endotoxemia have not been examined. Therefore, we investigated the anti-inflammatory action of GS on LPS-induced inflammatory responses on murine peritoneal macrophages and in septic mice. In the mouse endotoxemia model, GS prolonged survival and inhibited inflammatory mediators such as interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) and also inhibited the organ injury. In murine peritoneal macrophages, GS significantly inhibited production of nitric oxide (NOS) and COX-derived prostaglandin PGE2, as well as IL-1β and IL-6 and TNF-α. Furthermore, the anti-inflammatory activities of GS were mainly through heme oxygenase-1 (HO-1) induction, which was mediated by GSH depletion and ROS production from endoplasmic Reticulum (ER). The ROS mediated by GS caused the phosphorylation of GSK3β (ser9/21) and p38 and leads to translocation of nuclear factor (erythroid-derived 2)-like 2 (NRF2) which ultimately could induce HO-1. These results suggest that GS exerts anti-inflammatory responses through ROS -HO-1 axis.

My name is Amare Aregay. I am currently a PhD student at department of Gasteroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover Germany under supervision of prof.Dr Heiner Wedemeyer. Before commencing my Phd project I did my master degree at Wageningen University research center and did my year-long masters thesis at Erasmus Medical center, department of Viroscience, The Netherlands. My current Phd work focuses on cellular immune response (specifically T and NK cell response) towards chronic HCV infection in the context of IFN-free DAA therapy and liver transplantation.

Hepatitis C virus (HCV) persists and sets-up chronicity in majority of infected patients. Fortunately, new IFN-free direct-acting antiviral (DAA) therapies resulted in rapid and sustained clearance of HCV from infected patients. However, the impact of HCV clearance on HCV-specific CD8+ T cell responses remain yet to be understood. Owing to the rapid cessation of HCV replication and ensuing abrupt clearance of viral antigens mediated by IFN-free DAAs, we aimed at investigating the possible repercussions thereof on exhausted HCV-specific CD8+ T cells during chronic hepatitis C. We could show, by employing multimer-based magnetic bead enrichment technique that unlike activation markers that increased, ex-vivo surface expressions of co-regulatory markers remain unaffected following HCV clearance. Upon 10 day peptide stimulation in-vitro, the overall frequency of dextramer positive CD8+ T cells increased from baseline to 24 weeks after treatment in patients without advanced liver disease despite the fact that majority (55%) of patients did not show increase in proliferation. Meanwhile, HCV-specific CD8+ T cells proliferative capacity was not restored in patients with advanced liver disease. In addition, cytokines secretion and degranulation of HCV-specific CD8+ T cells remain unaffected following HCV clearance. .Importantly however, blockade of PDL1 pathway as well as PDL1/TIM3 double blockade resulted in enhanced proliferation and cytokine secretion by HCV-specific CD8+ T cells after IFN-free DAA therapy. Interestingly, HCV-specific CD8+ T cells that did not show increase in proliferation upon peptide stimulation alone could preferentially increase their proliferation and cytokine secretion upon blockade of PDL1 pathway. Taken together, our data implies that despite rapid HCV clearance, IFN-free DAA therapy does not fully re-constitute the altered phenotype and function of HCV-specific CD8+ T cells in chronic HCV. However, combining PDL1 or PDL1/TIM3 blocking therapy with IFN-free DAA therapy might possibly confer a functional and protective virus-specific CD8+ T cell response against re-infection.

Doctor Gerard Pasternak graduated from the Medical University Wroclaw in 2008. Since 2015 he has been the assistant 3rd Department and Clinic of Paediatrics, Immunology and Rheumatology of Developmental Age, Wroclaw Mediacal University and the Department of Immunology and Pediatrics, Provincial Hospital J. Gromkowski, Wroclaw (since 2010). He serves as a didactic assistant professor in the Department. Participates in conducting activities in the field of primary immunodeficiency for students of III-VI of the year, and takes part in the preparation and conducting of workshops for patients with PID and their families. Professional interests and research are concentrated on the diagnosis and treatment of primary and secondary immunodeficiencies, with particular emphasis on deficit of the IgG subclasses.

Statement of the Problem: Deficits in disorders of humoral immunity associated with a deficit of antibodies are the most common primary immunodeficiency. In the case of patients with primary and secondary immunodeficiencies, the total IgG and IgG subclasses measurements are used in patients diagnosis. Results of measurements show the discrepancy between the sum of IgG subclasses (IgGsum) and total IgG The purpose of this study is to analyze the influence of gender, age and IgG abnormality on the value of this discrepancy. Methodology & Theoretical Orientation: The group of patients was 648 children (aged 6 months to 18 years) referred to the Department of Clinical Immunology and Pediatrics for the purpose of diagnosis of immune disorders. For all children, measurements of the total IgG and the IgG subclasses (IgG1, IgG2, IgG3, IgG4) were conducted.The group of patients was divided into subgroups according togender, age (under 6 years of age, in the age range between 6.5 and 12 years and in the age range between 12.5 and 18 years) and IgG abnormality (below the normal range, normal and above the normal range).Findings: Statistical analysis (Kruskal–Wallis test)indicated that the all three parameters (age, gender and IgG abnormality) have a statistically significant effect on discrepancy between the IgGsum and total IgG.The higher average value of discrepancy between the IgGsum and total IgG was recognized in female group.The percentage of patients with IgG greater than IgGsum decreases as the age increases. The average value of discrepancy between the IgGsum and total IgG is highest for the group of age between 12.5 and 18 years. In the group of patients with normal IgG, the average value of discrepancy between the IgGsum and total IgG is lower than for children with IgG abnormality.

Anna Petrackova is a PhD student of Immunology at Palacky University Olomouc, Czech Republic. Her research is focused on gene expression studies of inflammatory mediators in autoimmune disorders and hematological malignancies & Dr. Eva Kriegova is an immunologist at the Faculty of Medicine and Dentistry, Palacky University Olomouc, Czech Republic. She is leader of Molecular Immunology group and her research interest is focused on molecular mechanisms of immune processes ongoing in inflammatory lung disorders and autoimmune diseases, hematological malignancies and orthopaedic disorders.

Statement of the Problem: Systemic lupus erythematosus (SLE) is a remarkably heterogeneous autoimmune disease. The greatest challenges remain in management of organ damage and lupus nephritis (LN), one of the most feared phenotypes in SLE. Despite tremendous effort, our knowledge on serum protein pattern in this severe SLE manifestations is still limited. The purpose of this study is to investigate the serum protein pattern of organ damage and LN in SLE using a highly sensitive multiplex Proximity Extension ImmunoAssay (PEA) on 92 inflammation-related proteins. Methodology & Theoretical Orientation: We enrolled 75 Czech patients with SLE, subgroups were formed according to the organ damage (assessed by the SLICC/ACR damage index, SDI) (SDI≥1, n=42; SDI=0, n=33) and biopsy-proven presence of LN (no LN, n=48; LN, n=27). The serum levels of 92 inflammation-related proteins were assessed by PEA (Proseek Multiplex, Olink Bioscience, Sweden). Statistical tests (Student t-test, Benjamini-Hochberg correction) were performed using GenEx (Sweden), P-value≤0.05 was considered as significant. Findings: SLE patients with organ damage had elevated serum levels of IL-8, CCL2, IL-6, CCL11, FGF21, MMP10, IL-18, CCL3, FGF5, and FGF23 comparing to those without organ damage. Of these, enhanced levels of CCL11, MMP10, and CCL2 were informative for identification of patients with organ damage. Importantly, IL-8 and MMP10 also correlated with disease activity (r≤0.355, P≤0.002). Comparing patients with/without LN, elevated levels of CSF1, sCX3CL and GDNF, sCD40 (Pcorr<0.05) were detected and showed usefulness in prediction of this severe SLE manifestation. Although all upregulated proteins correlated with disease activity, the best correlation was observed for sCX3CL1 and GDNF (r≥0.403, P≤0.0003). Conclusion & Significance: This highly sensitive PEA analysis identified serum pattern of organ damage and LN, with many novel candidate proteins detected. Their exact role and suitability as biomarkers in SLE deserves further investigation. Gant support: MZ CR VES 15-28659A, IGA_LF_2017_009.