IFN-gamma Gene Research Articles

Synthesis of recombinant gamma interferons resistant to proteolysis in the yeast Pichia pastoris

Genes encoding truncated versions of bovine and chicken gamma interferon genes lacking predicted protease cleavage sites at the C-terminus were constructed and expressed in the yeast Pichia Pastoris. The recombinant proteins possessed increased stability in comparison with the corresponding wild-type gamma interferons while retaining biological activity. The recombinant strains provide a useful tool for the purification of bovine and chicken gamma interferons for their use in veterinary applications.

The Interferon Gamma Gene Polymorphism In Acquired Aplastic Anemia and Its Association With HLA

IntroductionIdiopathic acquired aplastic anemia (AAA) is a rare and life-threatening disease, characterized by pancytopenia. Its immune-mediated physiopathology is not yet fully understood. However, important associations have been reported in AAA, which include the association with certain HLAs, the presence of a PNH clone in 40-50% of cases, occurrence of a hepatitis associated variant in about 5-10% of cases, the occurrence of interferon gamma (IFN-g) and tumor necrosis factor alpha (TNF-a) producing T cells in the peripheral blood and bone marrow cells. Not much is known about cytokine gene polymorphisms in AAA and its relationship with HLA alleles. The IFN-γ +874 A/T gene polymorphism, and specially the +874TT genotype, have been associated with elevated levels of IFN-γ production. The individuals can present 3 different phenotype (TT, TA or AA). Some groups have demonstrated that the TT (the IFN-gamma “hyper-producer type”) is possibly overrepresented in AA patients and correlates with susceptibility to the disease. In order to better understand the relationship between these immune parameters, we investigated the associations between IFN-g gene polymorphism in AAA patients and its relationship with the presence of HLADR15 (identified as of greater importance in our local patients). Materials and methodsIn this study we analyze the variations of the gene polymorphism at position +874 interferon in 30 consecutive patients with confirmed diagnosis of AAA,in 2012 and 2013, at the Federal University of the State of Bahia Hospital/Brazil. Diagnosis of AAA was confirmed by bone marrow biopsy. Patients also had their HLA typing and were tested for the IFN-gamma gene polymorphism at +874 T/A position using the ARMS methodology described by Pravica. As controls, 116 healthy individuals from the same population (Bahia/Brazil) were tested for these polymorphism. The analysis of the association between the gene polymorphism and the chances of developing AAA, and the polymorphism and the HLA-DR15, were performed using qui-square/exact fisher test. The results were expressed as OR. ResultsWe have found genotypic frequency of the A allele in 65%(n=39/60) in the aplastic patients (control 64.2%, n = 232, OR 1.05, p=0.87). The T allele was found in 35%(n =21/60)of the cases (control 34.6%, OR 0.95, p=0.87). Thus, there was no difference between the genotypic frequency of the alleles among patients with aplasia and the control group.In addition, there was NO difference between the frequency of the phenotype (TT, AT or AA) between patients with AAA and control group, respectively 16,5%, 36,5%, 46% (AAA, n =30) versus 18.1% , 35.3%, 46.6% (n = 116 control) (p=n.s.). The association of the polymorphism was tested and compared to the HLA-DR15. There was NO association between the IFN-gamma gene polymorphisms and the presence of HLADR15 (p =,90). ConclusionIn our brazilian cohort, the +874 T/A IFN-gamma gene polymorphism was Not associated with the onset of AAA. Also, there was No association between IFN-gamma gene polymorphism and the antigen HLA-DR15. TT phenotype, possibly the predisposing one for disease, was expressed only in 15% of patients with AAA. These findings corroborate the presence of a much more complex pathogenesis in AAA ,and that, regional differences in genetic and immune mechanisms may co-exist. Disclosures:No relevant conflicts of interest to declare.

Single nucleotide polymorphisms in the interferon gamma gene are associated with distinct types of retinochoroidal scar lesions presumably caused by Toxoplasma gondii infection

The association of single nucleotide polymorphisms (SNPs) in the interferon (IFN)-γ gene ( IFNG ) with different types of retinal scar lesions presumably caused by toxoplasmosis were investigated in a cross-sectional population-based genetic study. Ten SNPs were investigated and after Bonferroni correction, only the associations between SNPs rs2069718 and rs3181035 with retinal/retinochoroidal scar lesions type A (most severe scar lesions) and C (least severe scar lesions), respectively, remained significant. The associations of two different IFNG SNPs with two different types of retinal lesions attributable to toxoplasmosis support the hypothesis that different inflammatory mechanisms underlie the development of these lesions. The in vitro analysis of IFN-γ secretion by peripheral blood mononuclear cells stimulated with Toxoplasma gondii antigens was also investigated. The association between SNP rs2069718 and type A scar lesions revealed that differential IFN-γ levels are correlated with distinct genotypes. However, no correlation was observed with IFN-γ secretion levels and the SNP rs3181035 , which was significantly associated with type C scar lesions. Our findings strongly suggest that immunogenetic studies of individuals with congenital or postnatally acquired infection are needed to better understand the role of IFN-γ and its polymorphisms in the pathogenesis of ocular toxoplasmosis.

The interferon‐gamma (IFN‐γ) +874T allele reduces the risk of hepatitis B infection in an Asian population

Increasing evidence suggests that polymorphism of the interferon-gamma (IFN-γ) gene in the first intron at position +874 may be associated with chronic hepatitis B virus (HBV) infection and/or HBV clearance. However, the results of relevant studies have been inconsistent. To derive a more precise estimation of the association, we performed a meta-analysis. In total, 10 independent studies including 1661 chronic HBV-infected patients and 1142 controls were included in this meta-analysis. In studies following Hardy-Weinberg equilibrium (HWE), a significantly decreased risk of chronic HBV infection was associated with the IFN-γ+874TT genotype in the overall population (TT vs AA: odds ratio (OR)=0.714, 95% confidence interval (CI)=0.526-0.969, P=0.031) when compared with a spontaneously recovered population. Subgroup analysis by ethnicity revealed a similar association in Asian individuals (TT vs AA: OR=0.706, 95% CI=0.518-0.962, P=0.028). Moreover, when compared with a healthy control group, the 874T allele was associated with a significant lower risk of chronic HBV infection in the overall populations (TA vs AA: OR=0.439, 95% CI=0.193-0.997, P=0.049; TT+TA vs AA: OR=0.475, 95% CI=0.271-0.832, P=0.009) and in Asian individuals (TA vs AA: OR=0.862, 95% CI=0.744-0.999, P=0.048). In conclusion, the IFN-γ+874TT genotype and 874T allele reduce the risk of chronic HBV infection in Asian individuals.

120 : Loss of IFN-gamma 3′untranslated region AU-rich element affects B220+ cell populations in novel murine lupus model

Interferon-gamma (IFN-g) is a key player in immunoregulation, inflammation and autoimmunity. We created a mouse with a 162 nt deletion of an AU-rich element (ARE) region in the 3’UTR of the IFN-g gene. The ARE-deleted (ARE-Del) mice have chronic circulating serum IFN-g levels and develop a lupus-like disease characterized by nuclear antigen-specific autoantibodies and glomerulonephritis. B cells and plasmacytoid dendritic cells (pDCs) play a prominent role is systemic lupus erythematosus (SLE) and we find alterations in both B220+ cell types in these mice. The percentage of CD11c+, pDCA1+, B220+, Siglec H+ pDCs increases in bone marrow and spleens from ARE-Del mice. Furthermore, addition of IFN-g to flt-3 driven in vitro bone marrow cultures results in a dramatic increase in the pDC population with increases in IRF8, but not E2-2, mRNA expression that coincides with this expansion. In contrast, the total percentage of B220+, CD19+ B cells is reduced in spleens from ARE-Del mice. There is little impact on the CD23+ follicular B cell subset; however, CD21+ marginal zone (MZ) B cells are greatly reduced or absent. Crossing the ARE-Del mice with TLR7 or IFN-alpha receptor null mice restore MZ B cells to levels seen in WT mice indicating that interplay between type I and type II interferons plays a role in splenic B cell development. Current studies are underway to understand the molecular basis for how these interferons influence B cell maturation.

Polymorphisms of interferon gamma gene and risk of hepatocellular carcinoma in korean patients with chronic hepatitis B viral infection.

Increasing evidence supports the contribution of the pro-/anti-inflammatory cytokine balance and genetic factors to hepatocellular carcinoma (HCC). Here, we investigated whether genetic interferon gamma polymorphisms were associated with HCC in Korean patients with chronic hepatitis B. We genotyped a single nucleotide polymorphism (SNP, rs2430561, +874A/T) and a microsatellite (rs3138557, (CA)n repeat), located in the first intron of the interferon gamma gene, by direct sequencing and the gene scan method. A population-based case-control study of HCC was conducted and included 170 patients with chronic hepatitis and HCC, and 171 with chronic hepatitis B patients without hepatocellular carcinoma in a Korean population. Genotype and allele distributions of the interferon gamma gene SNP were associated with HCC. The frequencies of the AA genotype and the A allele were significantly increased in hepatocellular carcinoma subjects (p<0.05). Combined analysis using the genotype of rs2430561 and the number of microsatellites revealed that the frequencies of AT-CA12 and TT-CA12 increased significantly in hepatocellular carcinoma subjects (p<0.0001). Our results suggest that the interferon gamma gene may be a susceptibility gene and a risk factor for HCC in the Korean population.

Experimental Transmission ofSarcocystis muris(Apicomplexa: Sarcocystidae) Sporocysts from a Naturally Infected Cat (Felis catus) to Immunocompetent and Immunocompromised Mice

Cats serve as definitive hosts for zoonotic Toxoplasma gondii , a protozoan that threatens human reproductive health, but they also excrete sporocysts of related protozoan that pose no known human health risk. Here we provide the first definitive evidence for natural infection with the enzootic parasite Sarcocystis muris, one such enzootic parasite. Sporulated Sarcocystis sp. sporocysts were found in rectal contents of an adult feral cat ( Felis catus ) in Giza, Egypt. After these sporocysts were orally inoculated into 2 Swiss Webster mice, sarcocysts were found to have developed in skeletal muscles 114 days later. As observed through transmission electron microscopy, the cyst wall corresponded to Type 1, and the parasitophorous vacuolar membrane had tiny outpocketing of blebs (<200 nm thick) that were not invaginated into the interior of the cyst; these structures were identical to the sarcocyst wall described for a Costa Rican isolate of S. muris that has served as an experimental model for nearly 4 decades. Two parasite-free cats fed sarcocyst-infected muscles developed patent infections; fully sporulated sporocysts (10-11 × 7.0 μm) were found in the lamina propria of small intestines of cats killed 6 and 7 days postinoculation (PI). Interferon gamma gene knockout (KO) mice were orally inoculated with sporocysts from experimentally infected cats, and their tissues were examined histologically; sarcocysts were found in 5 KO mice killed 87, 115, 196, 196, 196 days PI, but no stages were seen in 5 KO mice 10, 14, 14, 18, and 39 days PI. Bradyzoites were released from intramuscular sarcocysts of a KO mouse killed 115 days PI and orally inoculated into 5 KO mice. No stage of Sarcocystis was found in any organ (including intestinal lamina propria) of KO mice killed 4, 8, 81, 190, and 190 days PI, confirming that the definitive host is required to complete the life cycle even in the case of immunodeficient mice. This is the first confirmation of S. muris infection in a naturally infected cat anywhere.

A new atypical genotype mouse virulent strain of Toxoplasma gondii isolated from the heart of a wild caught puma (Felis concolor) from Durango, Mexico

Nothing is known of the genetic diversity of Toxoplasma gondii circulating in wildlife in Mexico. In the present study, a mouse virulent T. gondii strain was isolated from the heart of a wild puma (Felis concolor). The puma was found roaming in outskirt of Durango City, Mexico and tranquilized for moving to a zoo. The puma died during translocation and a necropsy examination was performed. The puma had an antibody titer for T. gondii of 200 by the modified agglutination test. Its heart and brain tissue were bioassayed into 2 outbred Swiss Webster (SW) and 1 gamma interferon gene knockout (KO) mouse. The KO mouse and the 2 SW mice that became infected after inoculation with homogenate of puma heart died of acute toxoplasmosis 12, 19 and 20 days p.i. respectively and tachyzoites were found in lungs of all 3 mice. None of the 4 SW and 1 KO mouse inoculated with digest of the puma brain became infected with T. gondii. Tachyzoites from the lungs of mice were propagated in cell cultures. Tachyzoites from cell culture were inoculated into 5 SW; the mice died or had to be killed 14 days p.i. and a cat fed tissues of these mice shed T. gondii oocysts. Results of mortality and infectivity of tachyzoites and oocysts in SW mice indicated that the puma T. gondii strain (designated TgPumaMe1) was virulent for outbred mice. DNA isolated from culture-derived tachyzoites was characterized using 11 PCR-RFLP markers (SAG1, 5′- and 3′-SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico) revealed a new genotype (ToxoDB PCR-RFLP #222). Isolation of atypical genotype T. gondii from wild puma indicates that mouse virulent strains are circulating in wildlife in Mexico.

Association of Cytokine Gene Polymorphisms (IL6, IL10, TNF-&amp;#945;, TGF-&amp;#946; and IFN-&amp;#947;) and Graves’ Disease in Turkish Population

Cytokines play a crucial role in the pathogenesis of autoimmune thyroid disease, and recent studies have demonstrated an association between cytokine gene polymorphisms and Graves' Disease (GD) in different ethnic groups. The aim of the present study was to investigate the relationship of interleukin-6 (IL-6), IL-10, tumor necrosis factor-alpha (TNF-α), transforming growth factor-beta (TGF-β), and interferon-gamma (INF-γ) gene polymorphisms in the development of GD in Turkish population. A total of 224 subjects were included in the study comprising of 100 patients with GD (70 female, 30 male; mean age, 43.9 ±13.8 years) and 124 healthy subjects (81 female, 43 male); mean age, 37.8 ±10.2 years) without antithyroid autoantibodies or family history of autoimmune disorders. Genotyping was conducted by using PCR and sequence-specific primers. Statistical analysis showed a significant association between high TNF-α -308GA and IL-6 -174CC gene polymorphisms in patients with GD compared to control subjects (p=0.016, p=0.044, respectively). However, no significant differences were observed between GD and control subjects for IL-10, TGF-β, and INF-γ gene polymorphisms. TNF-α-308GA and IL-6 -174CC gene polymorphisms are involved in susceptibility to GD in Turkish population. The polymorphism hypothesis in pro-inflammatory cytokines might be involved in predisposition to GD.

Determination of the Cytokine Gene Polymorphism and Genetic Susceptibility in Tuberculosis Patients

Tuberculosis (TB) is a complicated disease in which biological, socioeconomical and environmental factors play role. Since only 10% of the individuals infected with Mycobacterium tuberculosis develop active disease, it has been suggested that host genetic factors may influence the risk for the development of TB. In this study, we aimed to investigate the presence and role of single nucleotide polymorphisms in the gene regions responsible for cytokine production, since these factors are considered to be associated with susceptibility or resistance to disease development. Single nucleotide polymorphisms were investigated by Amplification Refractory Mutational System (ARMS) Polymerase Chain Reaction (PCR) and PCR-Restriction Fragment Length Polymorphism (RFLP) methods. The presence of single nucleotide polymorphisms were analyzed in tumor necrosis factor alpha (TNF-α) gene promoter -308 G>A (rs1800629) region, interferon gamma (IFN-γ) gene +874 T>A (rs61923114) region, interleukin (IL)-12B p40 gene 1188 A>C (rs3212227) region, IL-10 gene promoter -1082 G>A (rs1800896) region and IL-4 gene promoter -590 C>T (rs2243250) region. A total of 84 patients (71 male, 13 female; mean age: 32.57 ± 15.94 years) whose clinical samples yielded M.tuberculosis complex growth, and 110 healthy blood donors (93 male, 17 female; mean age: 29.40 ± 11.56 years) as control group were included in this study. Of the patients, 76 (90.5%) were diagnosed as pulmonary and 8 (9.5%) as extrapulmonary TB. While 79 (94.1%) patients were newly diagnosed as TB, 5 (5.9%) patients had a TB history (relapsed TB). It was detected that acid-fast bacilli (AFB) were positive in 58 (69%) patients. According to the single nucleotide polymorphism results, gene frequencies could not be compared for TNF-a gene promoter -308 G>A region since healthy controls were in Hardy-Weinberg equilibrium while the patients were not. There were no statistically significant differences in allele and genotype distribution between the patients and healthy controls in IFN-γ gene +874 T>A region, IL-12B p40 gene 1188 A>C region, IL-10 gene promoter -1082 G>A region and IL-4 gene promoter -590 C>T region (p> 0.05). There were also no statistically significant differences between AFB positive (n= 58) and negative (n= 26) patients, and AFB positive (n= 56) and negative (n= 20) pulmonary TB patients (p> 0.05). In conclusion, no statistically significant differences were found associated with the susceptibility or resistance to TB with single nucleotide polymorphisms in the gene regions responsible for cytokine production in the study population. Only some of the single nucleotide polymorphisms of the gene regions responsible for cytokine release were investigated in our study. Therefore further detailed studies to investigate the polymorphisms in the genes that control the cytokine release and receptors specific for these cytokines, should be conducted. Although this study was performed in a relatively small sized population, these findings might provide a significant contribution to the epidemiologic data about the molecular immunology of TB in Turkey.

The peptide Ala-Glu-Asp-Gly and interferon gamma: Their role in immune response during aging

The decrease in interferon gamma expression by lymphocytes during aging is one of the main mechanisms leading to an immunodeficiency state in the elderly. Cell-penetrating geroprotective peptide Ala-Glu-Asp-Gly has the ability to activate proliferation of lymphocytes in the thymus during its aging. The nucleotide sequence that complementary contacts with Ala-Glu-Asp-Gly has been found in the promoter region of interferon gamma gene. Thus, the immunoprotective effect of the peptide can be explained by activation of interferon gamma production in T cells.

Molecular and Biological Characterization of First Isolates ofHammondia hammondifrom Cats from Ethiopia

Toxoplasma gondii oocysts are morphologically and antigenically similar to oocysts of another feline coccidian, Hammondia hammondi. The distinction between H. hammondi and T. gondii is important from an epidemiological perspective because all isolates of T. gondii are potentially pathogenic for humans and animals, whereas H. hammondi is not known to cause clinical disease in any naturally infected intermediate or definitive hosts. In the present report, H. hammondi (designated HhCatEt1 and HhCatEt2) oocysts were found microscopically in the feces of 2 of 36 feral domestic cats (Felis catus) from Addis Ababa, Ethiopia. Oocysts were orally infective to Swiss Webster and gamma interferon gene knockout mice; the inoculated mice developed tissue cysts in their muscles. Laboratory-raised cats fed mouse tissues of infected mice shed H. hammondi oocysts with a prepatent period of 5 days. The DNA extracted from sporulated oocysts reacted with H. hammondi-specific primers, and sequences were deposited in GenBank (accession nos. JX477424, and KC223619). This is the first report of isolation of H. hammondi from cats from the African continent.

Prevalence and Polymorphism in Interferon-γ Gene (CA) Repeats with Different Stages of Endometriosis

<b> </b>Background: Endometriosis is a female physical disorder that happens when cells from the lining of the womb (uterus) grow in other areas of the body i.e., outside of uterine cavity and leads to infertility. It represents a major personal and public health concern. The aim of the study was to investigate the prevalence and interferon-gamma gene CA-repeats polymorphism in patients with different stages of endometriosis.Methodology/Principal Findings: It is a case control prevalence study carried out in gynecology research centre. The mutational analysis of Interferon-γ gene CA repeats were tested for association in 356 affected women with different stages of disease and 372 women with no evidence of disease. The prevalence of endometriosis was accounted in 44.95% of infertile women. All the women were of South Indian origin and ascertained from the same infertility clinic. The broad IFN-gamma genotype and allele frequencies in all patients with different stages of endometriosis varied significantly from that in the control women (χ2 = 8.2690, 4 df, P =0.0822). The disparity in distribution was due to an increase in the a13 (114 bp) allele in the patients with endometriosis (χ2 =13.2394, p= 0.00027, OR=1.6656, 95.0% CI=1.263-2.1953). Conclusions/Significance: Significant difference was experiential in universal allele frequencies between the control women and specifically, women with endometriosis staging were tilted in the direction of minimal (39.04%) and mild disease (33.98%). Therefore, we conclude that interferon-gamma CA-repeat polymorphism may influence the likelihood of a woman developing different stages of endometriosis irrespective of its incidence.

Sarcocystis neurona infection in gamma interferon gene knockout (KO) mice: Comparative infectivity of sporocysts in two strains of KO mice, effect of trypsin digestion on merozoite viability, and infectivity of bradyzoites to KO mice and cell culture

The protozoan Sarcocystis neurona is the primary cause of Equine Protozoal Myeloencephalitis (EPM). EPM or EPM-like illness has been reported in horses, sea otters, and several other mammals. The gamma interferon gene knockout (KO) mouse is often used as a model to study biology and discovery of new therapies against S. neurona because it is difficult to induce clinical EPM in other hosts, including horses. In the present study, infectivity of three life cycle stages (merozoites, bradyzoites, sporozoites) to KO mice and cell culture was studied. Two strains of KO mice (C57-black, and BALB/c-derived, referred here as black or white) were inoculated orally graded doses of S. neurona sporocysts; 12 sporocysts were infective to both strains of mice and all infected mice died or became ill within 70 days post-inoculation. Although there was no difference in infectivity of sporocysts to the two strains of KO mice, the disease was more severe in black mice. S. neurona bradyzoites were not infectious to KO mice and cell culture. S. neurona merozoites survived 120min incubation in 0.25% trypsin, indicating that trypsin digestion can be used to recover S. neurona from tissues of acutely infected animals.

B cells participate on the early stages of immune response against intracellular bacterial pathogen Francisella tularensis

Event Abstract Back to Event B cells participate on the early stages of immune response against intracellular bacterial pathogen Francisella tularensis Zuzana Krocova1*, Klara Kubelkova1, Lenka Plzakova1, Lenka Zarybnicka1, Zuzana Sinkorova1 and Ales Macela1 1 Faculty of Military Health Sciences, University of Defense, Inst Molec Pathol, Czechia Francisella tularensis, as an intracellular bacterial pathogen, adhere, interact, and enter the range of phagocytic and non-phagocytic cells. Recently, we have demonstrated, using in vitro model, that both, vaccine strain F. tularensis LVS and virulent F. tularensis strain FSC200 are able to adhere and even to enter the human (Ramos) and mouse (A20) B cells where survive in non-replicative state. The entrance of F. tularensis into B cells required active participation of bacterium and engagement of B cell receptor. The in vivo analysis of the cellular response during early stages of F. tularensis FSC200 i.p. infection on murine model demonstrated that the cells responding early are the B1b cells and macrophages expressing CD11c marker. In parallel the monocyte-derived CD123+ F4/80+ dendritic cells started to express CD11b and disappeared from the site of infection. During this stage of infection F. tularensis induces increased expression of MHC class II molecules on B1a and B1b cells in peritoneum and activates genes for IL-12, TNF alpha, TGF beta, IFN-gamma, and iNOS at splenic B cells. Twenty-four hours after infection murine splenic B cells produce the specific anti-F. tularensis antibodies. Moreover, the co-cultivation of A20 B cells with the F. tularensis virulent strain FSC200 rendered them to be potent antigen presenting cells presenting F. tularensis antigens. Thus, considering the protective efficacy of circulating antibodies, production of cytokines, and potent antigen-presenting function of B cells, B cell-mediated, as well as T cell-mediated immunity plays an equivalent role in control of F. tularensis infection in mice. Acknowledgements Acknowledgement: The authors are supported by the grant from Grant Agency of Czech Republic No. P302/11/1631 References Crystal L. Jones, B. A. Napier, T. R. Sampson,a, A. C. Llewellyn,, M. R. Schroeder, and D. S. Weissb,c: Subversion of Host Recognition and Defense Systems by Francisella spp. Microbiol Mol Biol Rev. 2012 June; 76(2): 383–404. Krocova Z, Härtlova A, Souckova D, Zivna L, Kroca M, Rudolf E, Macela A, Stulik J. Interaction of B cells with intracellular pathogen Francisella tularensis. Microb Pathog. 2008 Aug;45(2):79-85. Kubelkova K, Krocova Z, Balonova L, Pejchal J, Stulik J, Macela A.: Specific antibodies protect gamma-irradiated mice against Francisella tularensis infection. Microb Pathog. 2012 Nov-Dec;53(5-6):259-68. doi: 10.1016/j.micpath.2012.07.006. Epub 2012 Jul 25. Yang Y, Ghosn EE, Cole LE, Obukhanych TV, Sadate-Ngatchou P, Vogel SN, Herzenberg LA, Herzenberg LA.: Antigen-specific antibody responses in B-1a and their relationship to natural immunity. Proc Natl Acad Sci U S A. 2012 Apr 3;109(14):5382-7. Keywords: B cells, Francisella tularensis, innate immunity, Antigen Presentation, Cytokines Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Host-pathogen interactions Citation: Krocova Z, Kubelkova K, Plzakova L, Zarybnicka L, Sinkorova Z and Macela A (2013). B cells participate on the early stages of immune response against intracellular bacterial pathogen Francisella tularensis. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00657 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 12 Jun 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. Zuzana Krocova, Faculty of Military Health Sciences, University of Defense, Inst Molec Pathol, Hradec Kralove, 500 01, Czechia, zuzana.krocova@unob.cz Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Zuzana Krocova Klara Kubelkova Lenka Plzakova Lenka Zarybnicka Zuzana Sinkorova Ales Macela Google Zuzana Krocova Klara Kubelkova Lenka Plzakova Lenka Zarybnicka Zuzana Sinkorova Ales Macela Google Scholar Zuzana Krocova Klara Kubelkova Lenka Plzakova Lenka Zarybnicka Zuzana Sinkorova Ales Macela PubMed Zuzana Krocova Klara Kubelkova Lenka Plzakova Lenka Zarybnicka Zuzana Sinkorova Ales Macela Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

2144 – IL-2, IL-6, IFN-gamma and TGF-beta genetic polymorphism with respect to susceptibility to schizophrenia

Schizophrenia is a chronic severe psychiatric disorder with multiple environmental and genetic determinants. Several reports indicate the possible role of immune system dysregulation in the pathogenesis of schizophrenia. An accumulating body of evidence indicates an association of schizophrenia and its psychopathology with altered cytokine production. The variation of the level of cytokines might be partly due to their functional gene polymorphism. The study was carried out to investigate the association of schizophrenia with the following cytokine polymorphisms: IL-2 (-330T/G), IL-6 (-174G/C), IFN-gamma (+874 T/A), TGF-beta (+869 T/C and +915 G/C). We included 130 patients diagnosed with schizophrenia and 184 controls in our study. The patients were evaluated for lifetime psychotic symptomatology using the Operational Criteria for Psychotic Illness (OPCRIT) checklist. The patients having history of traumatic brain injury, neurologic disorders, substance abuse or immune related diseases were excluded from the study by detailed medical examination. The polymorphisms were in HWE both in the cases’ and controls’ groups. In single marker analysis, we did not find an association for the tested polymorphisms. Our data do not support the role of IL-2, IL-6, IFN-gamma and TGF-beta gene polymorphisms in the predisposition to schizophrenia in the Polish population.

Kinetics of IFN-Gamma and TNF-Alpha Gene Expression and Their Relationship with Disease Progression after Infection withMycobacterium Tuberculosisin Guinea Pigs

PurposeGuinea pig is one of the most suitable animal models for Mycobacterium tuberculosis (M. tb) infection since it shows similarities to pulmonary infection in humans. Although guinea pig shows hematogenous spread of M. tb infection into the whole body, immunological studies have mainly focused on granulomatous tissues in lungs and spleens. In order to investigate the time-course of disease pathogenesis and immunological profiles in each infected organ, we performed the following approaches with guinea pigs experimentally infected with M. tb over a 22-week post-infection period.Materials and MethodsWe examined body weight changes, M. tb growth curve, cytokine gene expression (IFN-γ and TNF-α), and histopathology in liver, spleen, lungs and lymph nodes of infected guinea pigs.ResultsThe body weights of infected guinea pigs did not increase as much as uninfected ones and the number of M. tb bacilli in their organs increased except bronchotracheal lymph node during the experimental period. The gene expression of IFN-γ and TNF-α was induced between 3 and 6 weeks of infection; however, kinetic profiles of cytokine gene expression showed heterogeneity among organs over the study period. Histophathologically granulomatous lesions were developed in all four organs of infected guinea pigs.ConclusionAlthough IFN-γ and TNF-α gene expression profiles showed heterogeneity, the granuloma formation was clearly observed in every organ regardless of whether the number of bacilli increased or decreased. However, this protective immunity was accompanied with severe tissue damage in all four organs, which may lead to the death of guinea pigs.

Efficient Sleeping Beauty DNA Transposition From DNA Minicircles

DNA transposon-based vectors have emerged as new potential delivery tools in therapeutic gene transfer. Such vectors are now showing promise in hematopoietic stem cells and primary human T cells, and clinical trials with transposon-engineered cells are on the way. However, the use of plasmid DNA as a carrier of the vector raises safety concerns due to the undesirable administration of bacterial sequences. To optimize vectors based on the Sleeping Beauty (SB) DNA transposon for clinical use, we examine here SB transposition from DNA minicircles (MCs) devoid of the bacterial plasmid backbone. Potent DNA transposition, directed by the hyperactive SB100X transposase, is demonstrated from MC donors, and the stable transfection rate is significantly enhanced by expressing the SB100X transposase from MCs. The stable transfection rate is inversely related to the size of circular donor, suggesting that a MC-based SB transposition system benefits primarily from an increased cellular uptake and/or enhanced expression which can be observed with DNA MCs. DNA transposon and transposase MCs are easily produced, are favorable in size, do not carry irrelevant DNA, and are robust substrates for DNA transposition. In accordance, DNA MCs should become a standard source of DNA transposons not only in therapeutic settings but also in the daily use of the SB system.

Expression of Human Interferon-Gamma Gene in Human Tissue Culture Cells

ABSTRACTInterferon-gamma is a cytokine secreted by natural killer cells, CD4 TH1 cells and CD8 cytotoxic lymphocytes in response to mitogenic and antigenic stimuli. It is endowed with multiple biological activities and because of that is one of the first targets of recombinant DNA technology. Human interferon-gamma (hIFNγ) is expressed so far in Escherichia coli and various eukaryotic hosts. The disadvantages of the bacterial hIFNγ and unsatisfactory results from the eukaryotic expression were the reason to employ transient expression of the hIFNγ gene in CAP-T and HEKF cells, widely used for industrial production of human proteins. The obtained results demonstrated that (unlike with other human genes): i) cell viability decreased very fast and reached the critical level of 50% after five days only; ii) cell density did not exceed 0.7x105 cells/mL and sharply decreased after four days of cultivation; iii) the yield of hIFNy secreted in the supernatant did not exceed 5.8 mg/L.

新たな結核・ハンセン病ワクチン開発に向けて

The effectiveness of a vaccine against tuberculosis and leprosy is mainly judged by its capability to induce memory CD8 cytotoxic T cells (CTL). It has been reported that 'help' from CD4+ T cells is required to induce memory CTL. However, how CD4+ T cells instruct or support memory CTL during priming phase has not been resolved in detail. Therefore, we examined the helper function of CD4+ T cells in CTL differentiation. Peptide-25 is the major T cell epitope of Ag85B of Mycobacterium tuberculosis. We found that this peptide induced the expression of T-bet and TATA box binding protein-associated factor that can induce the chromatin remodeling of ifn-gamma gene, and as a result induced Th1 differentiation even in the absence of IFN-gamma and IL-12. Next, we established an in vitro CTL differentiation system using Peptide-25, Peptide-25 specific CD4+ T cells, OVA specific CD8+ T cells and splenic DC. By using this system, we found that CD4+ T cells activated DC even in the absence of IFN-gamma and CD40 ligand association, and the activated DC induced the functional differentiation of CTL. To identify the regulatory factors for DC activation, we analyzed the gene expression profile of helper CD4 T cells and identified 27 genes. Taken together, these results suggest that the inducing factors for Th1 differentiation are not indispensable to induce the functional differentiation of CTL.