ABSTRACT
NO synthase 2 (NOS2) is induced in airway epithelium by influenza virus infection. NOS2 induction late in the course of viral infection may occur in response to IFN-gamma, but early in infection gene expression may be induced by the viral replicative intermediate dsRNA through the dsRNA-activated protein kinase (PKR). Since PKR activates signaling pathways important in NOS2 gene induction, we determined whether PKR is a component in the signal transduction pathway leading to NOS2 gene expression after viral infection of airway epithelium. We show that NOS2 gene expression in human airway epithelial cells occurs in response to influenza A virus or synthetic dsRNA. Furthermore, dsRNA leads to rapid activation of PKR, followed by activation of signaling components including NF-kappaB and IFN regulatory factor 1. NOS2 expression is markedly diminished and IFN regulatory factor 1 and NF-kappaB activation are substantially impaired in PKR null cells. Strikingly, NOS2 induction in response to LPS is abolished in PKR null cells, confirming a central role for PKR in the general signaling pathway to NOS2.
Subject(s)
Influenza A virus/physiology , Nitric Oxide Synthase/biosynthesis , RNA, Double-Stranded/physiology , eIF-2 Kinase/physiology , Animals , Bronchi/cytology , Bronchi/enzymology , Bronchi/metabolism , Bronchi/virology , Cells, Cultured , DNA-Binding Proteins/biosynthesis , Enzyme Activation , Enzyme Induction , Epithelial Cells/enzymology , Epithelial Cells/metabolism , Epithelial Cells/virology , Humans , Interferon Regulatory Factor-1 , Mice , Mice, Inbred C57BL , Mice, Knockout , NF-kappa B/metabolism , Nitric Oxide Synthase Type II , Phosphoproteins/biosynthesis , RNA, Double-Stranded/chemical synthesis , RNA, Viral/chemical synthesis , RNA, Viral/pharmacology , eIF-2 Kinase/deficiency , eIF-2 Kinase/geneticsABSTRACT
We have made a human thymus cell hybridoma that secretes an immunosuppressive monoclonal lymphokine, referred to as hybridoma suppressor factor (HSF). This factor modulates the function of CD4+ cells suppressing their IL-2 production and suppressing PWM-induced B cell differentiation into Ig producing cells. Here we have examined the effect of HSF on the generation of T cell-derived lymphokines that regulate B cell growth and differentiation as well as the expression of other proteins involved in the control of T cell growth i.e., the p55 chain of the IL-2R and the transferrin receptor (TFR). HSF suppressed IFN-gamma activity produced by mitogen-stimulated PBMC without affecting the generation of lymphokines responsible for BCGF and BCDF activities. Additionally, HSF did not inhibit the expression of either IL-2R (p55) or TFR by activated T cells in spite of causing the suppression of IL-2 production. This evidence was further supported by experiments in which HSF selectively suppressed the accumulation of IL-2 mRNA without affecting IL-2R (p55) mRNA expression in mitogen-stimulated PBMC. The selective action of HSF may help to clarify the regulatory mechanisms involved in lymphokine gene expression as well as provide a way by which immune responses involved in autoimmunity and transplant rejection may be interrupted.
Subject(s)
Hybridomas/drug effects , Lymphokines/biosynthesis , Lymphokines/pharmacology , Antibody Formation/drug effects , Depression, Chemical , Gene Expression Regulation/drug effects , Humans , Hybridomas/metabolism , Interferon-gamma/biosynthesis , Interferon-gamma/pharmacology , Interleukin-2/biosynthesis , Interleukin-4/biosynthesis , Receptors, Interleukin-2/biosynthesis , Receptors, Transferrin/biosynthesisABSTRACT
Immune (gamma) interferon is a substance produced by immunologically activated mononuclear cells. Besides its antiviral activity, interferon gamma has a crucial role in immunoregulation, by acting directly upon lymphocytes and monocytes, and interacting with other soluble mediators of the immune response. Studies of the interferons system in inflammatory bowel disease have been limited, and little information is available on the generation of interferon during immunological events occurring in the human gut. To investigate the capacity of intestinal mucosal mononuclear cells to produce interferon gamma, lamina proprial mononuclear cells, isolated from Crohn's disease, ulcerative colitis, and control patients, were incubated with interleukin 2 or phytohemagglutinin, and the amounts of interferon gamma present in the culture supernatants were measured by a virus cytopathic effect inhibition assay. Under identical stimulatory conditions, culture supernatants of cells derived from actively involved mucosa of inflammatory bowel disease specimens contained two- to fivefold less interferon gamma than those of cells from control tissue. However, the amount of interferon gamma present in supernatants of cells from uninvolved inflammatory bowel disease mucosa was similar to that found in control supernatants. These results indicate that, in patients with active Crohn's disease and ulcerative colitis, mononuclear cells produce decreased amounts of interferon gamma in the intestinal mucosa. The exact significance of these findings is unclear, but because of the importance of interferon gamma in a variety of cell-mediated immune phenomena, its impaired availability might be relevant to the pathogenesis of inflammatory bowel disease.
