Raymond P. Donnelly

The expanded family of class II cytokines that share the IL-10 receptor-2 (IL-10R2) chain

Several novel interleukin (IL)‐10‐related cytokines have recently been discovered. These include IL‐22, IL‐26, and the interferon‐λ (IFN‐λ) proteins IFN‐λ1 (IL‐29), IFN‐λ2 (IL‐28A), and IFN‐λ3 (IL‐28B). The ligand‐binding chains for IL‐22, IL‐26, and IFN‐λ are distinct from that used by IL‐10; however, all of these cytokines use a common second chain, IL‐10 receptor‐2 (IL‐10R2; CRF2‐4), to assemble their active receptor complexes. Thus, IL‐10R2 is a shared component in at least four distinct class II cytokine‐receptor complexes. IL‐10 binds to IL‐10R1; IL‐22 binds to IL‐22R1; IL‐26 binds to IL‐20R1; and IFN‐λ binds to IFN‐λR1 (also known as IL‐28R). The binding of these ligands to their respective R1 chains induces a conformational change that enables IL‐10R2 to interact with the newly formed ligand‐receptor complexes. This in turn activates a signal‐transduction cascade that results in rapid activation of several transcription factors, particularly signal transducer and activator of transcription (STAT)3 and to a lesser degree, STAT1. Activation by IL‐10, IL‐22, IL‐26, or IFN‐λ can be blocked with neutralizing antibodies to the IL‐10R2 chain. Although IL‐10R2 is broadly expressed on a wide variety of tissues, only a subset of these tissues expresses the ligand‐binding R1 chains. The receptors for these cytokines are often present on cell lines derived from various tumors, including liver, colorectal, and pancreatic carcinomas. Consequently, the receptors for these cytokines may provide novel targets for inhibiting the growth of certain types of cancer.

Unique functions of the type II interleukin 4 receptor identified in mice lacking the interleukin 13 receptor α1 chain

The interleukin 4 receptor (IL-4R) is a central mediator of T helper type 2 (TH2)–mediated disease and associates with either the common γ-chain to form the type I IL-4R or with the IL-13R α1 chain (IL-13Rα1) to form the type II IL-4R. Here we used Il13ra1−/− mice to characterize the distinct functions of type I and type II IL-4 receptors in vivo. In contrast to Il4ra−/− mice, which have weak TH2 responses, Il13ra1−/− mice had exacerbated TH2 responses. Il13ra1−/− mice showed much less mortality after infection with Schistosoma mansoni and much more susceptibility to Nippostrongylus brasiliensis. IL-13Rα1 was essential for allergen-induced airway hyperreactivity and mucus hypersecretion but not for fibroblast or alternative macrophage activation. Thus, type I and II IL-4 receptors exert distinct effects on immune responses.

Interleukin-26: An IL-10-related cytokine produced by Th17 cells

IL-26 is classified as a member of the IL-10 cytokine family because it has limited sequence homology to IL-10 and the IL-10-related cytokines. The human IL-26 gene, IL26, is located on chromosome 12q15 between the genes for two other important class-2 cytokines, IFNG (IFN-γ) and IL22 (IL-22). IL-26 is often co-expressed with IL-22 by activated T cells, especially Th17 cells. It signals through a heterodimeric receptor complex composed of the IL-20R1 and IL-10R2 chains. IL-26 receptors are primarily expressed on non-hematopoietic cell types, particularly epithelial cells. Signaling through IL-26 receptor complexes results in the activation of STAT1 and STAT3 with subsequent induction of IL-26-responsive genes. The biological functions of IL-26 have only begun to be defined.

Interferon-lambda (IFN-λ) induces signal transduction and gene expression in human hepatocytes, but not in lymphocytes or monocytes

This study compared the ability of IFN‐α and IFN‐λ to induce signal transduction and gene expression in primary human hepatocytes, PBLs, and monocytes. IFN‐α drug products are widely used to treat chronic HCV infection; however, IFN‐α therapy often induces hematologic toxicities as a result of the broad expression of IFNARs on many cell types, including most leukocytes. rIFN‐λ1 is currently being tested as a potential alternative to IFN‐α for treating chronic HCV. Although IFN‐λ has been shown to be active on hepatoma cell lines, such as HepG2 and Huh‐7, its ability to induce responses in primary human hepatocytes or leukocytes has not been examined. We found that IFN‐λ induces activation of Jak/STAT signaling in mouse and human hepatocytes, and the ability of IFN‐λ to induce STAT activation correlates with induction of numerous ISGs. Although the magnitude of ISG expression induced by IFN‐λ in hepatocytes was generally lower than that induced by IFN‐α, the repertoire of regulated genes was quite similar. Our findings demonstrate that although IFN‐α and IFN‐λ signal through distinct receptors, they induce expression of a common set of ISGs in hepatocytes. However, unlike IFN‐α, IFN‐λ did not induce STAT activation or ISG expression by purified lymphocytes or monocytes. This important functional difference may provide a clinical advantage for IFN‐λ as a treatment for chronic HCV infection, as it is less likely to induce the leukopenias that are often associated with IFN‐α therapy.

A novel role for IL-22R1 as a driver of inflammation

The interleukin (IL)-22R1 chain of the heterodimeric IL-22 receptor is not expressed on normal leukocytes, but this receptor is expressed on T cells from anaplastic lymphoma kinase-positive (ALK(+)) anaplastic large cell lymphoma (ALCL) patients. To investigate the consequences of aberrant expression of this receptor on lymphocytes, we generated transgenic mice that express IL-22R1 on lymphocytes. The health of these animals progressively deteriorated at 8 to 12 weeks of age, as they displayed respiratory distress, rough coat and sluggish movement, and subsequent lethality due to multiorgan inflammation. The IL-22R1 transgenic animals developed neutrophilia that correlated with increased levels of circulating IL-17 and granulocyte colony-stimulating factor. In addition, these mice had increased serum IL-22 levels, suggesting that T cells expressing IL-22R1 generate IL-22 in a positive autoregulatory loop. As a result of the mouse model findings, we analyzed circulating cytokine levels in ALK(+)ALCL patients and detected elevated levels of IL-22, IL-17, and IL-8 in untreated patient samples. Importantly, IL-22 and IL-17 were undetectable in all patients who were in complete remission after chemotherapy. This study documents a previously unknown role of IL-22R1 in inflammation and identifies the involvement of IL-22R1/IL-22 in ALK(+)ALCL.

Interferon-lambda and therapy for chronic hepatitis C virus infection

Interferon (IFN)-α, a type-I IFN, is widely used to treat chronic hepatitis C virus infection, but the broad expression of IFN-α receptors often leads to adverse reactions in many organs. Here, we examine IFN-λ, a type-III IFN, as a therapeutic alternative to IFN-α. Like IFN-α, IFN-λ also induces antiviral activity in hepatocytes, but might induce fewer adverse reactions because its receptor is largely restricted to cells of epithelial origin. We also discuss the recent discovery of single nucleotide polymorphisms (SNPs) near the human IFN-λ3 gene, IL28B, that correlate strongly with the ability to achieve a sustained virological response to therapy with pegylated IFN-α plus ribavirin in patients with chronic hepatitis C.

An essential role for IFN-β in the induction of IFN-stimulated gene expression by LPS in macrophages.

TLR agonists such as LPS and poly(I:C) induce expression of type I IFNs, such as IFN‐α and ‐β, by macrophages. To examine the role of IFN‐β in the induction of ISGs by LPS, we compared the ability of LPS to induce ISGF3 activity and ISG expression in bone marrow–derived macrophages from WT and Ifnb1−/− mice. We found that LPS treatment activated ISGF3 and induced expression of ISGs such as Oas1, Mx1, Ddx58 (RIG‐I), and Ifih1 (MDA5) in WT macrophages, but not in macrophages derived from Ifnb1−/− mice or Ifnar1−/− mice. The inability of LPS to induce activation of ISGF3 and ISG expression in Ifnb1−/− macrophages correlated with the failure of LPS to induce activation of STAT1 and ‐2 in these cells. Consistent with these findings, LPS treatment also failed to induce ISG expression in bone marrow–derived macrophages from Stat2 KO mice. Although activation of ISGF3 and induction of ISG expression by LPS was abrogated in Ifnb1−/− and Ifnar1−/− macrophages, activation of NF‐κB and induction of NF‐κB‐responsive genes, such as Tnf (TNF‐α) and Il1b (IL‐1β), were not affected by deletion of either the IFN‐β or IFN‐αR1 genes. These findings demonstrate that induction of ISGF3 activity and ISG expression by LPS is critically dependent on intermediate production of IFN‐β and autocrine signaling through type I IFN receptors.

Distinct and overlapping genomic profiles and antiviral effects of Interferon-λ and -α on HCV-infected and noninfected hepatoma cells.

Summary.u2002 Recently, several SNPs in the region of the IL28B (IFN‐λ) gene have been associated with spontaneous clearance of hepatitis C virus (HCV) and enhanced cure rates for IFN‐alfa‐based therapies, suggesting a potential correlation between IFN‐λ and the ability to clear HCV. To understand the mechanism of IFN‐λ’s as compared to IFN‐α’s antiviral activity, we performed a comprehensive analysis of their anti‐HCV effects, whole genome transcriptome profiling with validation, and signalling of IFN‐α and IFN‐λ using J6/JFH‐1 and Huh7.5 cells in vitro. IFN‐λ and IFN‐α exhibited comparable anti‐HCV activity and gene expression profiles in Huh7.5 cells. While the majority of genes induced by IFN‐α and IFN‐λ were similar, IFN‐λ exhibits profound, but delayed kinetics of IFN‐stimulated genes (ISG) induction, while IFN‐α induced more rapid induction of ISGs. Furthermore, the increased induction of ISG expression by IFN‐λ correlated with up‐regulation of IFN‐λ receptor (IL‐28RA) expression and more prolonged activation of the Jak‐STAT signalling pathway. The findings from our comparative analysis of IFN‐α and IFN‐λ in HCV‐infected and noninfected cells support the clinical use of IFN‐λ as a potential alternative to IFN‐α in the treatment of chronic hepatitis C.

Induction of Uteroglobin-Related Protein 2 (Ugrp2) Gene Expression by the Th2 Cytokines IL-4 and IL-13

Uteroglobin-related proteins 1 and 2 (UGRP1 and -2) are thought to play important roles in inflammation and immunologic responses in the lung. In this study we demonstrate that IL-4 and IL-13 enhance Ugrp2 gene expression in the mouse transformed Clara cell line, mtCC, in a time- and dose-dependent manner. Addition of actinomycin D abrogated the IL-4- and IL-13-induced increase of Ugrp2 expression, demonstrating that this increase occurs at the transcriptional level. When mtCC cells were pretreated with IFN-γ before the addition of IL-4 or IL-13, IL-4- and 13-induced Ugrp2 mRNA increase was markedly decreased. IL-4 and IL-13 induced phosphorylation of STAT6 in mtCC cells, which binds to the proximal STAT-binding element (SBE) in the Ugrp2 gene promoter, leading to transcriptional activation of this gene. Mutations of the proximal SBE abrogated the binding of activated STAT6 to this site and the IL-4-induced increase in Ugrp2 gene promoter activity. IFN-γ-activated STAT1 binds to the same SBE in the Ugrp2 gene promoter to which STAT6 binds and decreases the binding of STAT6 to this site. Furthermore, an IL-4-induced increase in Ugrp2 expression was not observed in primary cultures of lung cells derived from STAT6-deficient mice. These results indicate that Ugrp2 expression is enhanced by IL-4 and IL-13 through STAT6 binding to the proximal SBE located in the Ugrp2 gene promoter.

IL-4 engagement of the type I IL-4 receptor complex enhances mouse eosinophil migration to eotaxin-1 in vitro.

Background Previous work from our laboratory demonstrated that IL-4Rα expression on a myeloid cell type was responsible for enhancement of Th2-driven eosinophilic inflammation in a mouse model of allergic lung inflammation. Subsequently, we have shown that IL-4 signaling through type I IL-4 receptors on monocytes/macrophages strongly induced activation of the IRS-2 pathway and a subset of genes characteristic of alternatively activated macrophages. The direct effect(s) of IL-4 and IL-13 on mouse eosinophils are not clear. The goal of this study was determine the effect of IL-4 and IL-13 on mouse eosinophil function. Methods Standard Transwell chemotaxis assay was used to assay migration of mouse eosinophils and signal transduction was assessed by Western blotting. Results Here we determined that (i) mouse eosinophils express both type I and type II IL-4 receptors, (ii) in contrast to human eosinophils, mouse eosinophils do not chemotax to IL-4 or IL-13 although (iii) pre-treatment with IL-4 but not IL-13 enhanced migration to eotaxin-1. This IL-4-mediated enhancement was dependent on type I IL-4 receptor expression: γC-deficient eosinophils did not show enhancement of migratory capacity when pre-treated with IL-4. In addition, mouse eosinophils responded to IL-4 with the robust tyrosine phosphorylation of STAT6 and IRS-2, while IL-13-induced responses were considerably weaker. Conclusions The presence of IL-4 in combination with eotaxin-1 in the allergic inflammatory milieu could potentiate infiltration of eosinophils into the lungs. Therapies that block IL-4 and chemokine receptors on eosinophils might be more effective clinically in reducing eosinophilic lung inflammation.