High Levels Of Interferon-alpha Research Articles

OMIP-051 - 28-color flow cytometry panel to characterize B cells and myeloid cells.

OMIP-051 - 28-color flow cytometry panel to characterize B cells and myeloid cells.

Plasmacytoid Dendritic Cells (pDCs) From HIV Controllers Produce Interferon-α and Differentiate Into Functional Killer pDCs Under HIV Activation

Human immunodeficiency virus (HIV) controllers spontaneously control viremia and CD4 T-cell depletion in contrast to viremic patients. After HIV exposure, plasmacytoid dendritic cells (pDCs) produce high levels of interferon alpha (IFN-α) and express the apoptotic ligand TRAIL (tumor necrosis factor-related apoptosis inducing ligand). Simian models have shown that prolonged high levels of IFN-α production could be responsible for AIDS progression. We studied pDC activation in response to human immunodeficiency virus (HIV) using flow cytometry and 3D microscopy. We show here that pDCs from controller patients produced higher levels of IFN-α in response to HIV than pDCs from viremic patients but similar levels to pDCs from healthy donors. Because binding of HIV to CD4 is essential for pDC activation, the low CD4 expression by pDCs from viremic patients may explain the weak IFN-α response to HIV. Three-dimensional microscopy revealed that pDCs from controllers and healthy donors expressed intracellular TRAIL that is relocalized to the membrane after HIV exposure. In contrast, pDCs from viremic patients expressed membrane TRAIL without any stimulation. We demonstrate that, in response to HIV, pDCs from controller patients produce IFN-α, express membrane TRAIL, and induce apoptosis of T-cell lines.

Interferon Regulatory Factor 5 in the Pathogenesis of Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by multiple genetic risk factors, high levels of interferon alpha (IFN-α), and the production of autoantibodies against components of the cell nucleus. Interferon regulatory factor 5 (IRF5) is a transcription factor which induces the transcription of IFN-α and other cytokines, and genetic variants of IRF5 have been strongly linked to SLE pathogenesis. IRF5 functions downstream of Toll-like receptors and other microbial pattern-recognition receptors, and immune complexes made up of SLE-associated autoantibodies seem to function as a chronic endogenous stimulus to this pathway. In this paper, we discuss the physiologic role of IRF5 in immune defense and the ways in which IRF5 variants may contribute to the pathogenesis of human SLE. Recent data regarding the role of IRF5 in both serologic autoimmunity and the overproduction of IFN-α in human SLE are summarized. These data support a model in which SLE-risk variants of IRF5 participate in a “feed-forward” mechanism, predisposing to SLE-associated autoantibody formation, and subsequently facilitating IFN-α production downstream of Toll-like receptors stimulated by immune complexes composed of these autoantibodies.

Interferon‐α mediates suppression of C‐reactive protein: Explanation for muted C‐reactive protein response in lupus flares?

C-reactive protein (CRP) is synthesized by hepatocytes in response to interleukin-6 (IL-6) during inflammation. Despite raised IL-6 levels and extensive systemic inflammation, serum CRP levels remain low during most viral infections and disease flares of systemic lupus erythematosus (SLE). Because both viral infections and SLE are characterized by high levels of interferon-alpha (IFNalpha), the aim of this study was to determine whether this cytokine can inhibit the induction of CRP. The interference of all 12 IFNalpha subtypes with CRP promoter activity induced by IL-6 and IL-1beta was studied in a CRP promoter- and luciferase reporter-transfected human hepatoma cell line, Hep-G2. CRP secretion by primary human hepatocytes was analyzed by enzyme-linked immunosorbent assay. CRP promoter activity was inhibited by all single IFNalpha subtypes, as well as by 2 different mixtures of biologically relevant IFNalpha subtypes. The most prominent effect was seen using a leukocyte-produced mixture of IFNalpha (56% inhibition at 1,000 IU/ml). The inhibitory effect of IFNalpha was confirmed in primary human hepatocytes. CRP promoter inhibition was dose dependent and mediated via the type I IFN receptor. Transferrin production and Hep-G2 proliferation/viability were not affected by IFNalpha. The current study demonstrates that IFNalpha is an inhibitor of CRP promoter activity and CRP secretion. This finding concords with previous observations of up-regulated IFNalpha and a muted CRP response during SLE disease flares. Given the fundamental role of both IFNalpha and CRP in the immune response, our results are of importance for understanding the pathogenesis of SLE and may also contribute to understanding the differences in the CRP response between viral and bacterial infections.

Impact of Secondary Structure of Toll-Like Receptor 9 Agonists on Interferon Alpha Induction

Oligodeoxynucleotides containing a CpG motif and double- or multistranded structure-forming sequences act as agonists of Toll-like receptor 9 (TLR9) and induce high levels of interferon alpha (IFN-alpha) in addition to other Th1-type cytokines. In the present study, we evaluated three highly effective IFN-alpha-inducing agonists of TLR9 to determine the type of duplex structures formed and the agonist's ability to induce immune responses, including IFN-alpha induction, in human cell-based assays and in vivo in mice and nonhuman primates. Thermal melting studies showed that two of the agonists evaluated had a single melting transition with similar hyperchromicity in both heating and cooling cycles, suggesting the formation of intermolecular duplexes. A third agonist showed a biphasic melting transition in the heating cycle and a monophasic melting transition with lower hyperchromicity during the cooling cycle, suggesting the formation of both intra- and intermolecular duplexes. All three agonists induced the production of Th1-type cytokines and chemokines, including high levels of IFN-alpha, in human peripheral blood mononuclear cell and plasmacytoid dendritic cell cultures. Subcutaneous administration of the two intermolecular duplex-forming agonists, but not the intramolecular duplex-forming agonist, induced cytokine secretion in mice. In nonhuman primates, the two agonists that formed intermolecular duplexes induced IFN-alpha and IP-10 secretion. On the contrary, the agonist that formed an intramolecular duplex induced only low levels of cytokines in nonhuman primates, suggesting that this type of structure formation is less immunostimulatory in vivo than the other structure. Taken together, the present results suggest that oligonucleotide-based agonists of TLR9 that form intermolecular duplexes induce potent immune responses in vivo.

Glomerular accumulation of plasmacytoid dendritic cells in active lupus nephritis: Role of interleukin‐18

Defective circulating dendritic cells (DCs) have been described in systemic lupus erythematosus (SLE) and correlated with high levels of interferon-alpha (IFNalpha). DCs are differentiated as being either myeloid or plasmacytoid, according to chemokine expression and the tendency to migrate toward inflamed tissue. We investigated the potential role of interleukin-18 (IL-18) in driving the glomerular migration of DCs in lupus nephritis (LN) and in affecting the ability of DCs to induce an imbalance in the Th1:Th2 ratio. DC subsets were characterized by flow cytometry and defined as either myeloid or plasmacytoid according to the expression of CD11c/blood dendritic cell antigen 1 (BDCA-1) and CD123/BDCA-2, respectively. The serum Th1:Th2 profile was studied by enzyme-linked immunosorbent assay. IL-18 receptor (IL-18R) and other chemokine receptors were analyzed by flow cytometry. Glomerular levels of IL-18/IL-18R and the presence of plasmacytoid DCs and myeloid DCs were investigated by immunohistochemical analysis. The number of peripheral plasmacytoid DCs was decreased in patients with SLE compared with control subjects, and this defect in the number of DCs was correlated with LN. Patients with LN showed a prevalent Th1 response, with high production of IL-18, IL-12 and IFNgamma. Only plasmacytoid DCs expressed IL-18R. Patients with severe LN showed a high accumulation of IL-18 within glomeruli in association with the presence of plasmacytoid DCs, whereas myeloid DCs were almost absent. A deficient number of peripheral plasmacytoid DCs correlated with high levels of Th1 cytokines and was associated with LN. Both serum and glomerular IL-18 were increased in LN. It is suggested that the high level of expression of IL-18R by peripheral plasmacytoid DCs allows the DCs to relocate within glomeruli under IL-18 stimulation and triggers the resident T cells, thus promoting renal damage.

Development of a continuous IL-7-dependent murine pre-B cell line PB-1 suitable for the biological characterisation and assay of human IL-7

Human interleukin-7 (IL-7) is a cytokine that appears to be critical for early T- and B-cell development and although IL-7 is currently under investigation as a therapeutic agent in a variety of hematolymphopoietic disorders, there have been few instances of the detection or investigation of this cytokine using a biological assay. This has been due, in the main, to the lack of a widely available, stable, easy to maintain and use, IL-7 responsive cell line. We have developed a pre-B-cell line, PB-1, from murine bone marrow, that is dependent on IL-7 for growth and has been maintained continually for up to 1 year without loss of responsiveness. The cells survive freezing and reviving, having been stored for periods of up to 4 years. The IL-7 bioassay is reproducible and sensitive, able to reliably detect 50 pg/ml IL-7. The assay is completely unresponsive to any other stimulatory cytokines tested and is not affected by a wide variety of inhibitory cytokines, with the exception of high levels of interferon alpha. The assay can be made completely specific for human IL-7 by including specific neutralizing antibodies for IL-7 and has been shown to be suitable for the estimation of IL-7 in both plasma and serum samples.

Proinflammatory cytokines and elastase-alpha-1-antitrypsin in Argentine hemorrhagic fever.

Argentine hemorrhagic fever (AHF) is a disease caused by Junin virus. In the acute phase, patients present hematologic and neurologic involvement with high levels of interferon-alpha and tumor necrosis factor-alpha (TNF-alpha. Nineteen patients with a confirmed diagnosis of AHF were studied: six severe, four moderate and nine mild cases. Serum levels of interleukin-6 (IL-6), IL-6 soluble receptor (IL-6sR), IL-8, IL-10, and elastase-alpha1-antitrypsin complex (E-alpha 1AT) were assayed by ELISAs. Levels of IL-6, IL-8, and IL-10 were high in nine, 12, and 13 patients, respectively, while levels of IL-6sR were high in two patients and low in one patient. Seven patients had increased levels of E-alpha1AT. Significant correlations were found between levels of both IL-8 and IL-10 with those of TNF-alpha as well as between IL-8 and E-alpha 1AT. These data demonstrate activation of pro-inflammatory and anti-inflammatory cytokine pathways, and statistical analysis showed differences among the clinical forms of illness. This study shows that IL-8 plays an essential role in neutrophil activation in AHF patients as demonstrated in other infectious diseases.

Aicardi-Goutières syndrome: an update and results of interferon-alpha studies.

Twenty-seven patients with familial encephalopathy with calcification of the basal ganglia and chronic cerebrospinal fluid (CSF) lymphocytosis (Aicardi-Goutières syndrome) are reviewed. In 19 children, the onset was within the first 4 months of life. Most patients had normal head circumference at birth, but 21 developed microcephaly between 3 and 12 months. Neuroimaging showed severe and progressive brain atrophy in all patients. The extent and intensity of the calcification was variable even in the same sibship. CSF lymphocytosis persisted beyond 12 months of age in 7 children. High levels of interferon-alpha were found in serum and CSF in 14 patients. The higher CSF levels suggest intrathecal synthesis. Tubuloreticular inclusions related to the presence of interferon were found in 4 additional children. The 19 patients still alive (6 older than 10 years) are profoundly disabled. However, the syndrome may present with individual variations in severity, rapidity of evolution, and imaging features. Neuropathological examination in 2 patients failed to detect significant inflammatory lesions and showed only foci of necrosis and wide-spread demyelination. This study supports an autosomal recessive inheritance for this syndrome. The high level of interferon-alpha is not explained but may play a role in the pathogenesis of the disorder.

Central interferon-alpha inhibits natural killer cytotoxicity through sympathetic innervation.

The brain has been known to produce high levels of interferon-alpha (IFN-alpha) during viral infections. We investigated the central and peripheral mechanisms of the brain IFN-alpha-induced suppression of natural killer (NK) cytotoxicity in the rat. The activity of NK cells in the spleen and the peripheral blood decreased 30-120 min after intracerebroventricular (icv) injection of recombinant human IFN-alpha of > 1,000 U but not after its intraperitoneal injection. This effect was antagonized by pretreatment with icv naltrexone (NLTX). Splenic denervation was observed to completely abolish the IFN-alpha-induced suppression of NK activity, whereas bilateral adrenalectomy did not. Furthermore, this immunosuppression was blocked by an icv injection of an antagonist of corticotropin-releasing factor (CRF), alpha-helical CRF-(9-41). The icv injection of CRF resulted in reduced NK activity, which was not affected by NLTX. The results suggest that brain IFN-alpha activates the CRF system through central opioid receptors and thereby suppresses the NK cytotoxicity predominantly through splenic sympathetic innervation.

High Levels of Interferon Alpha in the Sera of Children with Dengue Virus Infection

We measured the levels of interferon alpha (IFN alpha) in the sera of Thai children hospitalized with dengue hemorrhagic fever (DHF) or dengue fever (DF) to examine the role of IFN alpha in dengue virus infections of humans. The percentage of patients who had detectable levels of IFN alpha (> or = 3 U/ml) was higher in patients with DHF (80%, P < 0.001) and in patients with DF (60%, P < 0.001) than in healthy Thai children (7%). The levels of IFN alpha were higher in patients with DHF and in patients with DF on the first few days after the onset of fever than in healthy Thai children. The average levels of IFN alpha in patients with DHF were high two days before defervescence, decreasing gradually until the day of defervescence. There was a subset of patients with DHF who had increasing levels of IFN alpha after defervescence. However, the levels of IFN alpha in patients with DF were not high after fever subsided. The levels of IFN alpha were not different among children with DHF grades 1, 2 and 3. Among patients with DHF, T lymphocytes were activated to a higher degree in high IFN alpha producers than in low IFN alpha producers. These results indicate that similarly high levels of IFN alpha are produced in vivo during the acute stages of DHF and DF, and that high levels of IFN alpha remain after fever subsides in some patients with DHF, but not in patients with DF.

Lymphocyte-derived adrenocorticotropin is insufficient to stimulate adrenal steroidogenesis in hypophysectomized rats.

Cells of the immune system can produce and respond to peptide hormones associated with the endocrine system. However, the physiological significance of these endocrine-immune interactions is not known. It has been postulated that cells of the immune system, when stimulated with viruses that induce interferon-alpha, produce sufficient levels of ACTH to stimulate adrenal steroidogenesis and, thus, function as an auxiliary source of ACTH that may have a role in the response to stress. However, we have confirmed that levels of ACTH-related peptides produced by immunocompetent cells are far lower than those produced by the pituitary, raising questions about the ability of lymphocyte-derived ACTH to stimulate adrenal function. Furthermore, we have rigorously examined this issue using intact and hypophysectomized rats treated with Newcastle disease virus. Although high levels of interferon-alpha were produced by both intact and hypophysectomized rats, and the plasma corticosterone concentration increased dramatically in intact animals, corticosterone remained undetectable in hypophysectomized rats. The lack of a corticosterone response in these animals was not due to adrenal insensitivity to ACTH, as shown by a normal rise in corticosterone following Cosyntropin injection 8 h after hypophysectomy. The findings demonstrate that levels of ACTH produced by nonpituitary sources in response to viral infection are not sufficient to stimulate adrenal steroidogenesis.