Academic literature on the topic 'Interferon Regulator Faktor 1'

Sepsis is a severe systemic inflammatory response to infection. Endothelial activation and dysfunction play a critical role in the pathophysiology of sepsis and represent an important therapeutic target to reduce sepsis mortality. Interferon regulatory factor 1 (IRF-1) was recently identified as a downstream target of TNF-α-mediated signal transduction in endothelial cells. The aim of this study was to explore the importance of IRF-1 as a regulator of lipopolysaccharide (LPS)-induced endothelial proinflammatory activation. We found that renal IRF-1 was upregulated by LPS in vivo as well as in LPS-stimulated endothelial cells in vitro. Furthermore, we identified intracellular retinoic acid inducible gene-I (RIG-I) as a regulator of LPS-mediated IRF-1 induction. IRF-1 depletion specifically resulted in diminished induction of VCAM-1 in response to LPS, but not of E-selectin or ICAM-1, which was independent of NFκB signaling. When both IRF-1 and the RIG-I adapter protein mitochondrial antiviral signaling (MAVS) were absent, VCAM-1 induction was not additionally inhibited, suggesting that MAVS and IRF-1 reside in the same signaling pathway. Surprisingly, E-selectin and IL-6 induction were no longer inhibited by MAVS knockdown when IRF-1 was also absent, revealing a redundant endothelial activation pathway. In summary, we report an IRF-1-mediated proinflammatory signaling pathway that specifically regulates LPS-mediated VCAM-1 expression, independent of NFκB.

Cyclooxygenases (Cox) are rate-limiting enzymes that initiate the conversion of arachidonic acid to prostanoids. Cox-2 is the inducible isoform that is upregulated by proinflammatory agents, initiating many prostanoid-mediated pathological aspects of inflammation. In this study, we demonstrate that interferon (IFN)-γ alone or in synergy with lipopolysaccharide (LPS) or interleukin 1α induces Cox-2 expression in mouse peritoneal macrophages, which is paralleled by changes in Cox-2 protein levels and prostaglandin E2 (PGE2) release. Induction of Cox-2 was abrogated in macrophages that lack IFN regulatory factor (IRF)-1, consistent with an attenuated hepatic mRNA response in IRF-1−/− mice injected with LPS. Conversely, the absence of IRF-2 in macrophages resulted in a significant increase in both basal and inducible Cox-2 gene and protein expression as well as IFN-γ–stimulated PGE2 release, identifying IRF-2 as negative regulator of this promoter. Two IFN stimulation response elements were identified in the mouse Cox-2 promoter that were highly conserved in the human Cox-2 gene. Both bind endogenous IRF-1 and IRF-2 and regulate transcription in an IRF-1/2–dependent manner. Our data demonstrate conclusively the importance of IFN-γ as a direct activator and coactivator of the Cox-2 gene, and the central role of IRF-1/2 family members in this process.

The underlying mechanisms of thymocyte maturation remain largely unknown. Here, we report that serine/arginine-rich splicing factor 1 (SRSF1) intrinsically regulates the late stage of thymocyte development. Conditional deletion of SRSF1 resulted in severe defects in maintenance of late thymocyte survival and a blockade of the transition of TCRβhiCD24+CD69+ immature to TCRβhiCD24−CD69− mature thymocytes, corresponding to a notable reduction of recent thymic emigrants and diminished periphery T cell pool. Mechanistically, SRSF1 regulates the gene networks involved in thymocyte differentiation, proliferation, apoptosis, and type I interferon signaling pathway to safeguard T cell intrathymic maturation. In particular, SRSF1 directly binds and regulates Irf7 and Il27ra expression via alternative splicing in response to type I interferon signaling. Moreover, forced expression of interferon regulatory factor 7 rectifies the defects in SRSF1-deficient thymocyte maturation via restoring expression of type I interferon–related genes. Thus, our work provides new insight on SRSF1-mediated posttranscriptional regulatory mechanism of thymocyte development.

The RNase Regnase-1 is a master RNA regulator in macrophages and T cells that degrades cellular and viral RNA upon NF-κB signaling. The roles of its family members, however, remain largely unknown. Here, we analyzed Regnase-3–deficient mice, which develop hypertrophic lymph nodes. We used various mice with immune cell–specific deletions of Regnase-3 to demonstrate that Regnase-3 acts specifically within myeloid cells. Regnase-3 deficiency systemically increased IFN signaling, which increased the proportion of immature B and innate immune cells, and suppressed follicle and germinal center formation. Expression analysis revealed that Regnase-3 and Regnase-1 share protein degradation pathways. Unlike Regnase-1, Regnase-3 expression is high specifically in macrophages and is transcriptionally controlled by IFN signaling. Although direct targets in macrophages remain unknown, Regnase-3 can bind, degrade, and regulate mRNAs, such as Zc3h12a (Regnase-1), in vitro. These data indicate that Regnase-3, like Regnase-1, is an RNase essential for immune homeostasis but has diverged as key regulator in the IFN pathway in macrophages.

Aim of this thesis was to study the contribution of the hosts immune system during tumor regression. A wild-type rejection model was studied in which tumor regression is mediated through an adaptive, T cell host response (Research article 1). Additionally, the relationship between VACV infection and cancer rejection was assessed by applying organism-specific microarray platforms to infected and non-infected xenografts. It could be shown that tumor rejection in this nude mouse model was orchestrated solely by the hosts innate immune system without help of the adaptive immunity. In a third study the inflammatory baseline status of 75 human cancer cell lines was tested in vitro which was correlated with the susceptibility to VACV and Adenovirus 5 (Ad5) replication of the respective cell line (Manuscript for Research article 3). Although xenografts by themselves lack the ability to signal danger and do not provide sufficient proinflammatory signals to induce acute inflammation, the presence of viral replication in the oncolytic xenograft model provides the "tissue-specific trigger" that activates the immune response and in concordance with the hypothesis, the ICR is activated when chronic inflammation is switched into an acute one. Thus, in conditions in which a switch from a chronic to an acute inflammatory process can be induced by other factors like the immune-stimulation induced by the presence of a virus in the target tissue, adaptive immune responses may not be necessary and immune-mediated rejection can occur without the assistance of T or B cells. However, in the regression study using neu expressing MMC in absence of a stimulus such as a virus and infected cancer cells thereafter, adaptive immunity is needed to provoke the switch into an acute inflammation and initiate tissue rejection. Taken together, this work is supportive of the hypothesis that the mechanisms prompting TSD differ among immune pathologies but the effect phase converges and central molecules can be detected over and over every time TSD occurs. It could be shown that in presence of a trigger such as infection with VACV and functional danger signaling pathways of the infected tumor cells, innate immunity is sufficient to orchestrate rejection of manifested tumors
Ziel dieser Arbeit war, die Beteiligung des Wirts-eigenen Immunsystems bei der Tumoregression zu analysieren. Mittels eines Wildtyp-Regressionsmodells, wurde der Anteil des adaptiven Immunsystems studiert (Research-Artikel 1). Mit Hilfe von Organismus-spezifischen Mikroarrays und Genexpressionsanalysen konnte in einem Nacktmausmodell gezeigt werden, dass erfolgreiche, durch onkolytische VACV-vermittelte Tumortherapie auch ohne Beteiligung des adaptiven Immunsystems möglich ist (Research Artikel 2). In einer dritten Studie wurden 75 humane Tumorzelllinien auf ihren intrinsischen Entzündungsstatus hin getestet und bezüglich eines Zusammenhanges von diesem mit der Replikationsfähigkeit von VACV und Adenovirus 5 (Ad5) analysiert (Manuskript für den Research-Artikel 3). Obwohl Xenografts allein kein ausreichendes „Gefahrsignal“ geben und durch das Fehlen einer pro-inflammatorischen Stimulierung keine akute Entzündung verursachen können, ist die Infektion mit onkolytischem VACV ausreichend, um den Gewebe-spezifischen „Trigger“ darzustellen. In diesem Fall wird die Immunantwort aktiviert und nach der Hypothese des „Immunologic Constant of Rejection“ (ICR) geschieht dies, wenn eine chronische in eine akute Inflammation verändert wird. In dem beschriebenen onkolytischen Regressionsmodell ist die Präsenz des Virus ausreichend, um das Immunsystem zu aktivieren, d.h. die chronische Entzündung im Tumor in eine akute umzuwandeln. Dabei ist die adaptive Immunität mit T- und B-Zell-Aktivierung nicht notwendig für die Rückbildung des Tumors. In Abwesenheit eines solchen Stimulus, wie in der ersten Studie mit neu-exprimierenden MMCs, wird die Spezifität der adaptiven Immunantwort benötigt, um die akute Inflammation anzustoßen und die Tumorregression voranzutreiben. Zusammengefasst unterstützt diese Arbeit die Hypothese, dass die Mechanismen, die zu „tissue specific destruction“ (TSD) führen, in verschiedenen immunologischen Erkrankungen zwar divergieren, der Effektor-Mechanismus aber stets der Gleiche ist. Es zeigte sich, dass in Anwesenheit eines „triggers“, wie z.B. der VACV-Infektion und intakten „danger signaling pathways“ der Tumorzellen, die angeborene Immunität allein ausreicht, um die Tumorrückbildung zu vermitteln

Infektionen durch C. albicans auf den Schleimhäuten sind eine häufige Erkrankung bei Patienten mit einer Schwächung der T-Zellimmunität. Blutstrominfektionen mit der Hefe C. albicans (Candidämie) stellen, vor allem bei Patienten auf Intensivstationen, eine nach wie vor bedrohliche Komplikation mit hoher Letalität dar. Das pH-regulierte Antigen 1 (Pra1) ist ein Protein, das von C. albicans produziert wird, auf der Oberfläche des Pilzes gebunden vorkommt und auch vom Pilz in den Überstand sezerniert wird. Im humanen System bindet das Protein an T-Zellen an das Oberflächenprotein CD46. Es ist des Weiteren bekannt, dass das Pra1 an bestimmte Immunzellen der Maus (Monozyten und Phagozyten) binden kann. Eine Bindung an T-Zellen der Maus ist bisher nicht beschrieben. Eine genaue Charakterisierung der Interaktion von Pra1 mit Immunzellen der Maus ist interessant, da die Maus als biologischer Modellorganismus zur Erforschung der Infektion mit C. albicans dient. In dieser Arbeit konnte gezeigt werden, dass rekombinantes Pra1 (rPra1) auch an Maus-CD4+ T-Zellen binden kann. Es wurden Einflussfaktoren auf die gefundene Bindung von Pra1 an CD4+ T- Zellen gesucht. Als ein Einflussfaktor wurde Zink identifiziert. Pra1 kann an freies Zink binden und durch Zugabe von ZnCl2 während der Inkubation von Pra1 mit T-Zellen kann das Signal von gebundenem Pra1 an CD4+ T-Zellen erhöht werden. Aspf2, ein Protein aus Aspergillus fumigatus mit großer Homologie zu Pra1, kann nicht an diese Zellen binden. Im in-vivo-Experiment mit Tieren, die mit C. albicans infiziert wurden, konnte kein wildtypisches sezerniertes Pra1 gebunden an T-Zellen nachgewiesen werden. Zellkulturüberstände von C. albicans zeigten nach Inkubation in vitro mit T-Zellen ein Signal für gebundenes Pra1 an CD4+ T-Zellen. Die Bindungskinetik von Pra1 an T-Zellen zeigte eine über die Zeit der Inkubation konstante Zunahme des Signals von zellgebundenem rPra1 an CD4+ T-Zellen. In der off-Kinetik fand sich eine Abnahme des Signals über die Zeit bis an die Grenze der Nachweisbarkeit. Der Bindungspartner von Pra1 auf T-Zellen konnte nicht identifiziert werden. Die strukturell und funktionell verwandten Oberflächenproteine Crry, CD59a und CD55 wurden auf Bindungsfähigkeit an T-Zellen in entsprechenden Knockout- Mäusen getestet, konnten jedoch als Rezeptor für Pra1 ausgeschlossen werden. Durch die Bindung von sezerniertem Pra1 an neutrophile Granulozyten wird die Fähigkeit dieser Zellen zur Phagozytose eingeschränkt. Die Bindung von Pra1 an CD4+ T-Zellen führt zur Kostimulation der T-Zellen, also zur verstärkten Zellaktivierung und Proliferation. Durch die Zugabe von 10 μM Zinkchlorid wird die kostimulatorische Aktivität von Pra1 verstärkt. Während der Zellaktivierung von Effektor-Memory-CD4+ T-Zellen reduziert rPra1 die Sekretion von IFN-γ. Diese Reduktion von IFN-γ-produzierenden Zellen entsteht nicht durch einen Einfluss von Pra1 während der Zellaktivierung von naiven CD4+ T-Zellen zu Th1-Zellen und auch nicht durch die Auslösung von Apoptose in IFN-γ-produzierenden Th1-Zellen. Die Bindung von Pra1 an CD4+- T-Zellen, die über den T-Zell-Rezeptor aktiviert werden, reduziert in vitro die Sekretion des Zytokins. Zusätzlich werden weitere Zytokine in ihrer sezernierten Menge reduziert wie IL-2 und TNF-α
Infections caused by C. albicans on mucosal surfaces are a common disease in patients suffering from suppression of the T cell immune defense. Blood stream infections by the yeast C. albicans (candedemia) represent still a severe complication in patients in intensive care units with high rates of lethality. The pH-regulated antigen 1 (Pra1) is a protein produced by C. albicans which is present on the surface of the fungi and is secreted into the supernatant of fungal cultures as well. Pra1 can bind to human T cells via the surface protein CD46. It is known, that this protein can also bind to certain immune cells of mice (monocytes and phagocytes). Binding to T cells of mice is not yet known. A characterization of the interaction of Pra1 with immune cells of mice would be valuable, because mice act as a biological model system for the investigation of infections with C. albicans. In this paper, it could be shown that recombinant Pra1 (rPra1) can bind to mouse CD4+ T cells as well. After the finding that rPra1 can bind to CD4+ T cells, different parameters determining this binding have been studied. Zinc was found to be one influencing factor on the binding. Pra1 can bind free zinc ions and by the addition of ZnCl2 while incubating T cells with Pra1 the signal of bound Pra1 to CD4+ T cells could be increased. Aspf2, a protein from Aspergillus fumigatus with high homology to Pra1, was not able to bind to these cells. In in-vivo-experiments with animals infected with C. albicans, no wild-typic secreted Pra1 was found bound to T cells. Supernatant from C. albicans cultures produced, after incubation in vitro, a signal for cell-bound Pra1 on CD4+ T cells. Kinetics of the binding of rPra1 to T cells showed a constant increase of signal over the time of incubation. The off-kinetics revealed a decrease of cell-bound rPra1 over time to the edge of detectability. The receptor of Pra1 on T cells has not been identified yet. The structurally and functionally comparable surface proteins Crry, CD59a and CD55 were tested in knockout mice for each of these proteins and could be excluded as possible receptors. After binding of secreted Pra1 to neutrophilic granulocytes these cells experience a decreased capacity to phagocytose pathogens. The binding of Pra1 to CD4+ T cells leads to a costimulation of T cells, which results in increased cell activation and proliferation. This costimulatory capacity of Pra1 can be augmented by adding 10 μM zinc chloride. During activation of naïve CD4+ T cells Pra1 reduces the secretion of IFN-γ. The reduction of IFN-γ-producing cells is not due to an influence of Pra1 during cell activation of naive CD4+ T cells to Th1 cells and is also not due to induction of apoptosis in IFN-γ-producing Th1 cells. The binding of Pra1 to ex-vivo isolated CD4+ T cells reduces the in vitro secretion of IFN-γ after stimulating these cells via their T cell receptor. Additionally, the secretion of IL-2 and TNF-α was reduced

Magister Scientiae (Medical Bioscience) - MSc(MBS)
Breast cancer (BC) is one of the most common malignancies in women. Death results from treatment failure and metastatic disease. Thousands of lives might be saved if it was possible to detect and eliminate occult metastatic cells before they become clinically evident. Therefore, there is a critical need to identify new markers to improve treatment options for these patients. Genomic instability is the earliest indication of breast cancer and the use of genomic methodologies is a progress towards early detection and treatment, through the identification of biomarkers that can be translated into novel therapy targets. The interferon regulatory factor-1(IRF-1) gene, localized on chromosome 5q31.1, is believed to act as a tumor suppressor gene in breast cancer. The IRF-1 was found to be inactivated by single nucleotide polymorphism (SNP) in breast cancer suggesting that the loss of its function might be critical to the development of the disease. The phosphatidylinositol 3-kinase (PIK3) signaling pathway mediates key cellular functions and alterations of genes in this pathway, including PIK3CA, serine-threonine protein kinases (AKT1and AKT2), phosphatase and tensin homolog (PTEN), fibroblast growth factor receptor 2 (FGFR2) and ERBB2, whose expression have been demonstrated to be altered in breast cancer patients. In addition, these genes are linked to treatment resistance. vi In this study, we have investigated allelic loss of IRF-1 gene in primary tumors obtained from patients undergoing mastectomy at Groote Schuur hospital (Cape Town, South Africa). These samples were then further analyzed for the DNA copy number changes of specific genes involved in the PIK3/AKT signaling pathway. Statistical analysis has been performed in order to correlate genomic findings with clinical-histopathological and follow up information from the patients and to establish whether these genes can predict prognosis. Our data analysis has indicated that 46 cases (45.5%) out of 101 cases were informative for the IRF-1 dinucleotide marker used for LOH analysis (Figure 3.1). LOH was detected in 23 of these informative cases (23/46; 50%). No statistical significance was found between LOH at the IRF-1 locus and age (≤50 years or >50 years) (P value = 1.0000) and earlier stage (Stages I and II) (P value= 0.4982) based on Fisher’s exact test. Patients presented a high level of DNA copy number changes in genes involved in the PIK3/AKT pathway. The most frequent changes were observed in the PIK3CA and PTEN genes. PIK3CA presented high copy number in 36.8% of the cases. PTEN was observed with low copy number in 47.5% of the cases. This dissertation shows the effectiveness of genomic methodologies as means for the detection of early breast cancer progression in South African women. The PIK 3/AKT genes can validate the usefulness of breast cancer therapies.