Cytokine reporter mice in immunological research: perspectives and lessons learnedCroxford, Andrew L.; Buch, Thorsten
doi: 10.1111/j.1365-2567.2010.03372.xpmid: 21070235
Cytokines are soluble messenger molecules with important regulatory functions throughout the immune system. ‘Cytokine reporter’ strains express marker molecules under control of elements from cytokine genes allowing for easy identification of their cellular sources. Such systems are well‐accepted tools for research of cytokine function. The value of these strains lies in the ability to perform experiments relying on identification and isolation of live cytokine‐expressing cells, provided that the reporter faithfully reflects the proper cytokine mRNA and protein production. As more diverse cell subsets are defined by their cytokine expression, the field has adapted with the generation of more sophisticated strains. In this review we summarize the evolution of cytokine detection methods and give examples of knowledge gained using cytokine reporter mice for cell types expressing interferon‐γ and interleukin‐4, ‐10 and ‐17. We also discuss current options for generating such reporter strains and their potential pitfalls.
Transgenic modelling of cytokine polarization in the lungDela Cruz, Charles S.; Kang, Min‐Jong; Cho, Won‐Kyung; Lee, Chun Geun
doi: 10.1111/j.1365-2567.2010.03376.xpmid: 21091906
The lung is one of the commonest sites of exposure to environmental allergen or pathogen, so the expression of a variety of cytokines in the lung is dynamically regulated by inflammatory or structural cells in the lung. In the last decades, characterization of the local lung cytokine milieu in allergic or injury models has identified a collective role of certain cytokines, such as type 1 or type 2 cytokines, driving polarized inflammatory and tissue phenotypes. With the development of transgenic mouse modelling systems, the effector function of individual cytokine and the pathophysiological consequences of cytokine polarization in the lung have been effectively evaluated. Here, we present an overview of the transgenic systems currently used to assess the biological function of cytokine or other mediators in the lung. We discuss the inflammatory and tissue phenotypes detected in the lungs of transgenic mice over‐expressing representative T helper type 1 (interferon‐γ, interleukin‐12), T helper type 2 (interleukins ‐4, ‐5, ‐9, ‐10 and ‐13), and T helper type 17 cytokines. The effects of genetic modification of cytokine receptors or transcriptional factors such as GATA‐3 and T‐bet in pulmonary inflammation and remodelling tissue responses are also discussed because these transcription factors are regarded as essential regulators of cytokine polarization. Finally, we discuss the limitations and future application of transgenic approaches in the studies of human lung diseases characterized by cytokine polarization.
Evolution of CD33‐related siglecs: regulating host immune functions and escaping pathogen exploitation?Cao, Huan; Crocker, Paul R.
doi: 10.1111/j.1365-2567.2010.03368.xpmid: 21070233
Sialic‐acid‐binding immunoglobulin‐like lectins, siglecs, are important immune receptors expressed widely in mammals. A unique feature of siglecs is their ability to bind sialylated glycans and transmit signals to immune cells. The CD33‐related siglecs (CD33rSiglecs) form a major subfamily of the siglecs, containing a large, rapidly evolving group of genes that expanded in mammals through an inverse duplication event involving a primordial cluster of siglec genes over 180 million years ago. Humans express a much larger set of CD33rSiglecs than mice and rats, a feature that can be explained by a dramatic loss of CD33rSiglec genes in rodents. Most CD33rSiglecs have immune receptor tyrosine‐based inhibitory motifs and signal negatively. Interestingly, novel DAP‐12‐coupled ‘activating’ CD33rSiglecs have been identified, such as siglec‐14 and siglec‐16, which are paired with the inhibitory receptors, siglec‐5 and siglec‐11, respectively. The evolution of these activating receptors may have been driven in part by pathogen exploitation of inhibitory siglecs, thereby providing the host with additional pathways by which to combat these pathogens. Inhibitory siglecs seem to play important and varied roles in the regulation of host immune responses. For example, several CD33rSiglecs have been implicated in the negative regulation of Toll‐like receptor signalling during innate responses; siglec‐G functions as a negative regulator of B1‐cell expansion and appears to suppress inflammatory responses to host‐derived ‘danger‐associated molecular patterns’. Recent work has also shown that engagement of neutrophil‐expressed siglec‐9 by certain strains of sialylated Group B streptococci can suppress killing responses, thereby providing experimental support for pathogen exploitation of host CD33rSiglecs.
Histamine H4 receptor activation on human slan‐dendritic cells down‐regulates their pro‐inflammatory capacityGschwandtner, Maria; Schäkel, Knut; Werfel, Thomas; Gutzmer, Ralf
doi: 10.1111/j.1365-2567.2010.03336.xpmid: 20722760
6‐Sulpho LacNAc dendritic cells (slanDC) are a major population of human blood DC that are highly pro‐inflammatory, as characterized by their outstanding capacity to produce tumour necrosis factor‐α and interleukin‐12 (IL‐12) and to prime antigen‐specific T‐cell responses. SlanDC were found to be present in inflamed tissue such as atopic dermatitis, where high levels of histamine are also present. As histamine is an important regulator of allergic inflammation we investigated the role of histamine receptors, particularly the most recently identified histamine H4 receptor (H4R), in modulating the pro‐inflammatory function of slanDC. The expression of H4R was evaluated by real‐time PCR and flow cytometry. Cytokine production in response to H4R stimulation was assessed by intracellular flow cytometric staining and enzyme‐linked immunosorbent assay. We show that slanDC express the H1R, H2R and H4R on mRNA and the H4R on protein level. No differences were observed in basal H4R expression in patients with atopic dermatitis and psoriasis, but in atopic dermatitis patients the H4R was up‐regulated by interferon‐γ. When stimulated with lipopolysaccharide in the presence of histamine, slanDC produced substantially lower levels of the pro‐inflammatory cytokines tumour necrosis factor‐α and IL‐12, mediated solely via the H4R and via the combined action of H2R and H4R, respectively. In contrast, the production of IL‐10 was not affected by histamine receptor activation on slanDC. The slanDC express the H4R and its stimulation leads to reduced pro‐inflammatory capacity of slanDC. Hence, H4R agonists might have therapeutic potential to down‐regulate immune reactions, e.g. in allergic inflammatory skin diseases.
Porcine circovirus type 2 DNA influences cytoskeleton rearrangements in plasmacytoid and monocyte‐derived dendritic cellsBalmelli, Carole; Steiner, Esther; Moulin, Hervé; Peduto, Nadja; Herrmann, Brigitte; Summerfield, Artur; McCullough, Kenneth
doi: 10.1111/j.1365-2567.2010.03339.xpmid: 20840632
Functional disruption of dendritic cells (DC) is an important strategy for viral pathogens to evade host defences. In this context, porcine circovirus type 2 (PCV2), a single‐stranded DNA virus, impairs plasmacytoid DC (pDC) and conventional DC activation by certain viruses or Toll‐like receptor (TLR) ligands. This inhibitory capacity is associated with the viral DNA, but the impairment does not affect all signalling cascades; TLR7 ligation by small chemical molecules will still induce interleukin‐6 (IL‐6) and tumour necrosis factor‐α secretion, but not interferon‐α or IL‐12. In this study, the molecular mechanisms by which silencing occurs were investigated. PP2, a potent inhibitor of the Lyn and Hck kinases, produced a similar profile to the PCV2 DNA interference with cytokine secretion by pDC, efficiently inhibiting cell activation induced through TLR9, but not TLR7, ligation. Confocal microscopy and cytometry analysis strongly suggested that PCV2 DNA impairs actin polymerization and endocytosis in pDC and monocyte‐derived DC, respectively. Altogether, this study delineates for the first time particular molecular mechanisms involved in PCV2 interference with DC danger recognition, which may be responsible for the virus‐induced immunosuppression observed in infected pigs.
Interleukin‐25 fails to activate STAT6 and induce alternatively activated macrophagesStolfi, Carmine; Caruso, Roberta; Franzè, Eleonora; Sarra, Massimiliano; De Nitto, Daniela; Rizzo, Angelamaria; Pallone, Francesco; Monteleone, Giovanni
doi: 10.1111/j.1365-2567.2010.03340.xpmid: 20840631
Interleukin‐25 (IL‐25), a T helper type 2 (Th2) ‐related factor, inhibits the production of inflammatory cytokines by monocytes/macrophages. Since Th2 cytokines antagonize classically activated monocytes/macrophages by inducing alternatively activated macrophages (AAMs), we here assessed the effect of IL‐25 on the alternative activation of human monocytes/macrophages. The interleukins IL‐25, IL‐4 and IL‐13 were effective in reducing the expression of inflammatory chemokines in monocytes. This effect was paralleled by induction of AAMs in cultures added with IL‐4 or IL‐13 but not with IL‐25, regardless of whether cells were stimulated with lipopolysaccharide or interferon‐γ. Moreover, pre‐incubation of cells with IL‐25 did not alter the ability of both IL‐4 and IL‐13 to induce AAMs. Both IL‐4 and IL‐13 activated signal transducer and activator of transcription 6 (STAT6), and silencing of this transcription factor markedly reduced the IL‐4/IL‐13‐driven induction of AAMs. In contrast, IL‐25 failed to trigger STAT6 activation. Among Th2 cytokines, only IL‐25 and IL‐10 were able to activate p38 mitogen‐activated protein kinase. These results collectively indicate that IL‐25 fails to induce AAMs and that Th2‐type cytokines suppress inflammatory responses in human monocytes by activating different intracellular signalling pathways.
Varying expression of four genes sharing a common regulatory sequence may differentiate rheumatoid arthritis from ageing effects on the CD4+ lymphocytesSoroczyńska‐Cybula, Monika; Bryl, Ewa; Smoleńska, Żaneta; Witkowski, Jacek M.
doi: 10.1111/j.1365-2567.2010.03341.xpmid: 20738421
The CD28 gene is similarly down‐regulated in CD4+ lymphocytes from both healthy elderly people and patients with rheumatoid arthritis (RA) because of impaired protein‐binding activity of the ‘α’ sequence in its promoter region. Other genes important for the CD4+ cell function may share that sequence and may be similarly regulated and affected. We searched GenBank for possible ‘α’ homologues and then compared transcriptional activities of the respective genes in the CD4+ cells of young and older healthy individuals and those with RA by real‐time PCR. We show here that genes encoding one of the zinc finger proteins (ZNF334), the ‘aging hormone’ Klotho, the retinoid acid receptor β2 (RARβ2) and the T‐cell adapter protein GRAP‐2, contain sequences with various (exceeding 70%) degrees of homology to the ‘α’ sequence near their promoters. These genes are transcribed in human CD4+ lymphocytes; the expressions of RARβ2, KLOTHO and ZNF334 are significantly decreased in a correlated manner in the cells of patients with RA compared with those of healthy individuals. In RA patients, the extremely reduced expression of ZNF334 does not depend on the individual’s age, apparently constituting a disease‐related phenomenon; whereas that of RARβ2 and KLOTHO occurs mostly in the cells of relatively younger patients, making them similar to the lymphocytes of healthy elderly in this aspect.