Multifaceted actions of IL-26 in antiviral, antimicrobial, and autoimmune responses.

<p>Upon activation, various immune cells secrete IL-26. IL-26 exerts antiviral and antimicrobial actions through dual action by (A) direct killing of bacteria by forming membrane pores and (B) by priming immune cells, such as neutrophils, NK cells, and plasmacytoid dendritic cells (pDCs). Other Th17 cell-derived molecules might act in synergy with IL-26 to enhance this direct killing of bacteria. (C) IL-26 response requires tight regulation as increased expression of IL-26 has been reported in several autoimmune and inflammatory diseases, including Crohn’s disease and rheumatoid arthritis (RA). This scheme was drawn in part by using pictures from Servier Medical Art.</p

IL-10 family cytokines: source, receptors, and target cells.

<p>IL-10 family cytokines are produced by both immune and non-immune cells and signal through heterodimeric receptors expressed on diverse target cells. However, IL-26 might also mediate heterodimeric IL-20R1/IL-10R2 receptor-independent signaling. This scheme was drawn in part by using pictures from Servier Medical Art. NKT, Natural killer T cells; ILC3, Group 3 innate lymphoid cell; MΦ, Macrophage.</p

Co-treatment of A549 cell with IL-1β and <i>Viscum album</i> inhibits the cytokine-induced COX-2 expression.

<p>A549 cells were treated with IL-1β (10 ng/ml) and two different concentrations of <i>Viscum album</i> Q Spez preparation for 18 hours. Cytosolic COX-2 was measured using flow cytometric analysis. <i>Viscum album</i> is added to the cells either from the beginning of the experiment along with IL-1β (co-treatment) or after 14 hours of IL-1β induction (post-treatment). Percentage COX-2 expression as measured in intracellular staining by flow cytometry (A) and mean fluorescence intensity (MFI) of COX-2 expression (B) is shown. Results are mean ±SEM of 4 independent experiments (**<i>p</i><0.01; ***<i>p</i><0.001).</p

Effect of <i>Viscum album</i> on the stability of COX-2 protein as analyzed by flow cytometry.

<p>A549 cells were stimulated with IL-1β for 90 minutes with or without VA Qu Spez. Cells were harvested at different time intervals after blocking the protein synthesis with cyclohexamide (10 μg/ml) for 90 minutes till 11 hours. Normalised percentage COX-2 expression as measured in intracellular staining by flow cytometry (A) and mean fluorescence intensity (MFI) of COX-2 expression (B) is shown. Data is representative of mean ±SEM of three independent experiments.</p

Fungal hydrophobins.

<p>Fungal hydrophobins are unique amphipathic proteins with multiple roles in the fungal life cycle and in mediating interactions between fungus and host. There is diversity in the primary sequences of hydrophobins but they share a similar core three-dimensional structure and a pattern of four disulfide bonds (shown in amber) that stabilize the structures. Increasingly, these proteins show potential for modification of hydrophobic nanomaterials and in solubilizing lipophilic drugs.</p

Host immune response to pathogens and predisposition to infections due to autoimmunity.

<p>Antigens from invading pathogens are recognized and presented by innate immune cells (A) such as macrophages and dendritic cells to CD4+ and CD8+ T cells (CTL) (B). CD8+ T cells recognize endogenous antigens presented by MHC class I molecules and exert cytotoxic functions upon activation. CD4+ T cells recognize antigens presented in the context of MHC class II molecules, and under the influence of innate cells and cytokine milieu, CD4+ T cells can be polarized into different subsets such as Th1, Th2, Th17, and regulatory T cells (Tregs) that secrete distinct cytokines. CD4+ T cells provide help to B cells to produce antigen-specific antibodies (C). However, due to autoimmunity, neutralizing autoantibodies can be produced against any of these key components of the immune system critical for mounting anti-microbial responses and might either predispose to an increased risk of bacterial, viral, and opportunistic fungal infections or exacerbate the ongoing infectious diseases. Indeed, in patients with infections, the occurrence of neutralizing autoantibodies against several key cytokines such as IFN-γ, IL-6, GM-CSF, IL-17, and IL-22 (highlighted in red boxes) that interfere with the host immune response to pathogens have been demonstrated. In addition, autoantibodies are also reported against type I IFNs and IL-12 that might play role in predisposition to infections (highlighted in blue boxes). CTLA-4, cytotoxic T lymphocyte antigen-4; CTL, cytotoxic T lymphocyte; FasL, Fas ligand; GM-CSF, granulocyte/macrophage–colony stimulating factor.</p

Inhibition of cytokine-induced COX-2 protein expression by <i>Viscum album</i> in A549 cells.

<p>A549 cells were either cultured in medium alone or treated with IL-1β (10 ng/ml) with or without increasing concentration of VA Qu Spez for 18 hours. Using 20 µg of total cellular protein, expression of COX-2 was analyzed by western blotting. <b>A.</b> A representative western-blot depicting the effect of VA on expression of IL-1β-induced COX-2. <b>B.</b> Relative expression (mean ± SEM) of COX-2 protein from four independent experiments as quantified by densitometry (ratio of density of COX-2: that of β-actin). *<i>P</i><0.05 versus control cells and ** <i>p</i><0.05 versus IL-1β-stimulated cells as analyzed by paired Student-t-test. <b>C.</b> Kinetics of COX-2 protein expression upon treatment of A549 cells with various doses of VA. The cells were either cultured in medium alone or stimulated with IL-1β with our without VA for 12, 18, 24 and 36 hours. Expression of COX-2 protein was analyzed by western blotting and relative expression of COX-2 protein was quantified by densitometry (ratio of density of COX-2: that of β-actin). Results are representative of two experiments.</p

Effect of <i>Viscum album</i> on COX-1 expression in A549 cells.

<p><b>A and B.</b> A549 cells were treated with increasing concentrations of VA Qu Spez in presence of IL-1β (10 ng/ml) for 18 hours. Protein expression of COX-1 was analyzed by western blot. A representative western-blot depicting the effect of VA on COX-1 expression under IL-1β-stimulated culture conditions (A). Relative expression (mean ± SEM) of COX-1 protein from three independent experiments as quantified by densitometry (ratio of density of COX-1: that of β-actin). ns, non-significant (B). <b>C.</b> Cells were cultured either in medium alone or treated with increasing concentrations of VA Qu Spez without IL-1β for 18 hours. 20 µg of total cellular protein was separated by 10% SDS-PAGE followed by western blotting to analyze the expression of COX-1. Results are representative of two experiments.</p

Inhibition of PGE2 secretion by <i>Viscum album</i> in A549 cells.

<p>Cells were treated with IL-1β (10 ng/ml) and increasing concentrations of VA Qu Spez for 18 hours. PGE2 was analyzed in culture supernatants by EIA. Results are mean±SEM from 4 independent experiments (*<i>p</i><0.05 versus control cells, **<i>p</i><0.05 versus cells treated with IL-1β, analyzed by paired Student-t-test).</p

Effect of <i>Viscum album</i> on the stability of COX-2 protein as determined by western blot.

<p>Confluent A549 cells were treated with IL-1β in the presence and absence of VA Qu Spez in dose dependent concentrations in μg/ml. Cells were harvested at different time intervals after blocking the protein synthesis with cyclohexamide (10 μg/ml) for 90 minutes till 11 hours. COX-2 expression was measured by western blot using the cytosolic extracts. (A), inhibition of COX2 protein synthesis by VA at 18 hours. (B) (C) (D) are the representative western blots after 90 minutes, 5 hours and 11 hours respectively showing level of COX-2 expression after cyclohexamide treatment with or without <i>Viscum album</i>. β-actin was used as an internal control. All blots are representative of three independent experiments and the densitometry values for each band are mentioned below the representative blots.</p