Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A biologically relevant mechanism of generating peripheral tolerance in T cells that have escaped thymic deletion is by the oral administration of soluble antigens. Oral tolerance occurs by two distinct mechanisms. Feeding a single high dose of antigen induces anergy of antigen-specific TH1 cells, while multiple low doses of antigen induce regulatory T cells that mediate suppression by producing immunosuppressive cytokines. Although cytokine production orally tolerant animals has been well studied utilizing in vitro assay systems, semi-quantitative characterization of cytokine production in oral tolerance in vivo has not been carried out. In this paper we have developed a system using semi-quantitative RNA polymerase chain reaction to characterize cytokine gene expression in vivo in mice orally tolerized by feeding either a single high dose or multiple low dosages of antigen. We find that measurable differences in interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) gene expression occurred between the tolerized and non-tolerized groups. Qualitatively, IL-4 mRNA was increased in both orally tolerized groups. However, significant differences in IL-4 gene expression between the two groups in both magnitude and kinetics were found. A large but short-lived increase in IL-4 was produced in mice fed a single high dose, while a relatively more moderate, longer-lived increase was produced in mice fed multiple low doses. The increase in IL-4 gene expression was specific only to the draining lymph node following antigen administration. Expression of IFN-gamma was decreased in both orally tolerant groups. These results indicate that tolerance in TH1 cells is induced by both of these feeding regimens while TH2 responses are intact and amplified upon reexposure to antigen.
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PMID:Different kinetic patterns of cytokine gene expression in vivo in orally tolerant mice. 795 64

Because activated human macrophages can be potent sources of IL-10, and because immunosuppressive tolerance-inducing macrophages populate the skin after UV exposure, we determined whether IL-10 is induced after UV exposure of human skin and whether it is related to the immigrating macrophages. Keratomes were obtained from control skin or from skin obtained 72 h after a single exposure to four minimal erythemal doses of UVB. Quantitative reverse-transcriptase PCR on total RNA extracted immediately from skin keratomes showed that IL-10 mRNA was elevated in UV-exposed skin. Epidermal cell suspensions from non-UV-exposed keratomes (C-EC) and UV-exposed keratomes (UV-EC) were fractionated by sequential immunobead selection. IL-10 mRNA was reproducibly 200- to 400-fold higher in CD11b+ UV-EC (macrophages) relative to CD11b- UV-EC (keratinocytes). IL-10 mRNA was not detected in C-EC that contained the CD1a+ population (Langerhans cells) nor in CD1a- C-EC keratinocytes from normal skin. As determined by ELISA, CD11b+ UV-EC IL-10 cell-associated protein was fivefold higher than that of CD11b- UV-EC; this was confirmed by flow cytometric visualization of IL-10 protein in permeabilized cells. CD11b+ UV-EC macrophages secreted IL-10 protein into the supernatant at a level of 333 +/- 51 pg/10(6) cells, whereas UV-EC keratinocytes did not secrete detectable levels of IL-10 (n = 3), although UV did induce low levels of IL-10 mRNA and cell-associated protein in keratinocytes. Therefore, although human keratinocytes accumulate intracellular IL-10 after in vivo UV exposure, the most potent production and secretion of IL-10 in the epidermis seems to be that of UV-induced macrophages. Skin-infiltrating macrophage secretion of such a potent immunoregulatory cytokine may account for the delayed immunosuppressive environment of sunburned skin and the altered APC activity of the infiltrating macrophages.
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PMID:CD11b+ macrophages that infiltrate human epidermis after in vivo ultraviolet exposure potently produce IL-10 and represent the major secretory source of epidermal IL-10 protein. 796 79

High viral burden and replication persist during all phases of human immunodeficiency virus (HIV) disease. Although monotherapy has yielded considerable benefits, these benefits are neither absolute nor durable. Combination therapy has multiple goals: to reduce viral replication and burden; to relieve drug toxicity; to attenuate viral mutations leading to resistance and possibly to conversion from non-syncytium-inducing to syncytium-inducing virus; and to broaden the spectrum of specific cells and tissues in which antiretroviral agents are active. At present, zidovudine remains the cornerstone of antiretroviral monotherapy and combination therapy. A partial list of agents tried in combinations with and without zidovudine includes the nucleoside analogues zalcitabine and didanosine; non-nucleoside reverse-transcriptase inhibitors (nevirapine, delavirdine, atevirdine, pyridinones, TIBO derivatives); protease inhibitors; inhibitors of viral regulatory functions (tat inhibitors); cytokine antagonists; acyclovir; and colony-stimulating factors. The rationales, the regimens, and the results all vary. We usually recommend combination therapy for treatment-naive patients who are asymptomatic with < 200 CD4+ cells/mm3 or who are symptomatic, and for patients who have been receiving zidovudine monotherapy and who are stable but whose CD4+ counts have fallen to < 300 cells/mm3, or who are progressing. In the absence of definitive results from clinical trials of combination therapy, the decision to embark on this route remains to be made between each individual patient and the practitioner.
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PMID:Issues in combination antiretroviral therapy: a review. 796 49

A novel immunomodulator, imiquimod, has been shown to be an effective topical antiviral and antitumor agent in animal models. Imiquimod has been reported to induce interferon-alpha and other cytokines in animals and humans, but its precise role as an immunomodulator at skin sites has not been determined. We investigated its effect on cytokine gene expression in the human epidermal carcinoma cell line COLO-16 and human keratinocytes. COLO-16 cells were incubated with imiquimod (1 and 10 micrograms/ml) and human keratinocytes with 5 micrograms/ml for 6 or 24 h. Cytokine gene expression was analyzed by reverse-transcriptase PCR. In COLO-16 cells, imiquimod stimulated IL-6 mRNA levels 2.3- and 4.4-fold at 1 and 10 micrograms/ml after 6 h. IL-8 mRNA increased 4-fold at both 1 and 10 micrograms/ml. At 24 h, though IL-6 mRNA level at 1 micrograms/ml was further stimulated, enhanced expressions of IL-8 at 1 micrograms/ml and both IL-6 and IL-8 at 10 micrograms/ml were down-regulated. In human keratinocytes, 5 micrograms/ml of imiquimod stimulated IL-6 mRNA levels 1.4-fold at 6 h and 2.1-fold at 24 h, and IL-8 mRNA levels 1.7- and 2.0-fold at 6 and 24 h. IL-1 alpha mRNA levels in COLO-16 or keratinocytes were unchanged by either dose or incubation time. These results suggest that stimulation of IL-6 and IL-8 expression may be involved in the immunomodulating action of imiquimod.
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PMID:Effects of a novel topical immunomodulator, imiquimod, on keratinocyte cytokine gene expression. 806 Nov 17

Cytokine gene expression in peripheral blood mononuclear cells during the development of graft-versus-host disease (GVHD) in patients who underwent allogeneic bone marrow transplantation (allo BMT) was analysed using a semiquantitative reverse-transcriptase polymerase chain reaction (RT-PCR). The expression of interleukin (IL)-1 beta, IL-6, and tumour necrosis factor (TNF)-alpha mRNA was increased during the development of GVHD and the degree of this increment depended on the severity of the disease. IL-2 expression was not detected at all and interferon-gamma expression was not much changed during GVHD. In patients with hepatic veno-occlusive disease (VOD), another transplantation-related complication, the expression of IL-1 beta and TNF-alpha mRNA was increased but IL-6 mRNA expression showed little increase. These findings suggest that IL-1 beta, IL-6 and TNF-alpha produced by peripheral blood mononuclear cells play an important role in the development of GVHD. Furthermore, liver dysfunction due to GVHD or VOD may be distinguishable by this type of cytokine analysis. Analysis of cytokine mRNA expression in peripheral blood mononuclear cells after allogeneic bone marrow transplantation may provide important information concerning the immune response and the cytokine network system in marrow transplant patients.
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PMID:Cytokine gene expression in peripheral blood mononuclear cells during graft-versus-host disease after allogeneic bone marrow transplantation. 813 79

We assessed the origin of peripheral blood cells and bone marrow cells obtained from 15 patients after allogeneic bone marrow transplantation (allo BMT) by sensitive two-step polymerase chain reaction (PCR) amplification of MCT118, a variable number of tandem repeats regions (VNTR), that can be used to detect the DNA pattern of a minor cell population of only 1% without using radioisotopes. Mixed chimerism(MC) was detected in the haematopoietic cells of 3 patients. Two patients developed relapse of leukaemia after the detection of MC and one patient died of bone marrow hypoplasia 7 months after BMT. These findings indicate the clinical usefulness of this method to monitor patients with MC. Also, we analyzed cytokine gene expression in peripheral blood mononuclear cells during the development of graft-versus-host disease (GVHD) in patients who underwent allo BMT using a semiquantitative reverse-transcriptase polymerase chain reaction (RT-PCR). The expression of interleukin(IL)-1 beta, IL-6, and tumor necrosis factor (TNF)-alpha mRNA was increased during the development of GVHD and the degree of this increment depended on the severity of the disease. These findings suggest that IL-1 beta, IL-6, and TNF-alpha produced by peripheral blood mononuclear cells play an important role in the development of GVHD. Therefore, analysis of MC and cytokine mRNA expression using the PCR technique after allogeneic bone marrow transplantation provide important information for treatment and monitoring of marrow transplant patients.
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PMID:[Clinical application of gene technology to monitor bone marrow transplantation]. 815 60

Human interleukin-10 (h-IL-10) is a pleiotropic cytokine with stimulatory activity on B-lymphocytes. Recent evidence indicates that infection with Epstein-Barr virus (EBV) induces h-IL-10 production in B-cells and that this cytokine may contribute to EBV-induced B-cell transformation. It is not known whether h-IL-10 induction by EBV correlates with distinct phenotypic features of the infected cells or with the expression of particular viral genes. We have approached these questions by investigating the expression of h-IL-10 mRNA in a panel of B-cell lines including: in vitro EBV-transformed lymphoblastoid cell lines (LCLs), EBV-carrying Burkitt lymphoma (BL) lines, EBV-negative BL lines and their sublines infected with different EBV strains, or transfected with the transformation-associated viral gene. h-IL-10 mRNA was detected by reverse-transcriptase-assisted (RT)-PCR in a subset of EBV-negative BLs and in all EBV-positive BL lines and LCLs investigated except Daudi. This cell line carries an EBV nuclear antigen (EBNA)-2 gene-defective virus strain. h-IL-10 mRNA was induced by conversion of 3 EBV-negative and h-IL-10-negative BL lines (BL41, BL47 and BL49) with the transforming, B95.8-derived EBV strain. P3HR-I virus convertants that do not express the viral EBNA-2 and the EBV latent membrane protein (LMP)-1, and fail to progress towards a LCL-like cell phenotype, showed no evidence of h-IL-10 up-regulation. Expression of LMP1 was sufficient to induce h-IL-10 mRNA in transfected sublines of the EBV-negative DG75 and BL41 cell lines, whereas expression of EBNA1, 2, 5, or 6 had no effect. h-IL-10 was detected in the culture supernatants of the LMP1 transfectants by specific ELISA assays. The present findings confirm the role of LMP1 in the transactivation of a wide variety of cellular genes which may be involved in EBV-induced B-cell transformation.
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PMID:The Epstein-Barr virus latent membrane protein-1 (LMP1) induces interleukin-10 production in Burkitt lymphoma lines. 815 62

Cytokine production by T cells in the cerebrospinal fluid (CSF) and central nervous system (CNS) of SJL/J mice during myelin basic protein (MBP)-induced experimental allergic encephalomyelitis (EAE) was examined. Reverse transcriptase/polymerase chain reaction (RT/PCR) was used to measure interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) mRNA levels from perfused CNS tissue (brain and spinal cord) and from cells isolated from CSF. Animals were grouped according to EAE severity, ranging from asymptomatic (adjuvant only) to severe disease (paralysis or severe paresis). Cytokine signals, normalized to actin, were almost undetectable in control tissues, and only slightly elevated in whole CNS tissue from animals with mild EAE. Both cytokine messages were strongly upregulated in CNS tissues derived from severely affected animals, consistent with previous observations correlating disease progression with infiltration by memory/effector CD4+ T cells, the major source of these cytokines. This cytokine upregulation was specific to the CNS, since other organs from the same animals did not express significant levels of IL-2 and IFN-gamma. CSF was obtained from the cisterna magna of unperfused mice and verified as such by absence of red blood cells (RBCs) and by immunoglobulin concentration orders of magnitude lower than in serum. Cytokine message was measured in RNA isolated from cells in CSF. Levels of IL-2 and IFN-gamma mRNA in CSF cells were significantly elevated in mild EAE and strongly upregulated in severe disease, correlating with those in total CNS tissue. These results confirm the CSF as representative of the immune status of the CNS and indicate a role for IL-2 and IFN-gamma in inflammatory CNS disease.
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PMID:Cytokine production by cells in cerebrospinal fluid during experimental allergic encephalomyelitis in SJL/J mice. 829 48

In situ hybridization is a technique with widespread application. However, its usefulness has been limited by the need for radioactive materials and the requirement for the DNA to be cloned onto an appropriate vector. We have utilized the polymerase chain reaction to directly incorporate a T7 RNA polymerase promoter sequence onto the cDNA for interleukin-2. Digoxigenin-labelled riboprobes were then synthesized using this PCR product as a template. The digoxigenin-labelled riboprobes were then used in non-radioactive in situ hybridization to detect messenger RNA for interleukin-2 in mitogen stimulated peripheral blood mononuclear cells. This methodology has the potential for widespread application in immunology and cytokine research.
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PMID:Rapid nonradioactive in situ hybridization for interleukin-2 mRNA with riboprobes generated using the polymerase chain reaction. 830 89

Local cytokine gene expression in vivo was analyzed by direct analysis of RNA obtained from salivary gland tissues of MRL/lpr mice with autoimmune sialadenitis. The expression of cytokine genes were assessed by the reverse-transcriptase polymerase chain reaction, and by immunohistochemical analysis. The expression of interleukin-1(IL-1)beta and tumor necrosis factor was detected before the onset of inflammatory lesions in the salivary glands of mice of 1 or 2 months of age, and IL-6 mRNA expression was clearly detected at the time of onset of typical autoimmune sialadenitis at 3 months of age in MRL/lpr mice, and was up-regulated with advancing age. These results suggest that the overexpression of these inflammatory cytokine genes is involved in the development and progression of organ-localized autoimmunity in the salivary glands of MRL/lpr mice.
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PMID:Expressions of cytokine genes during development of autoimmune sialadenitis in MRL/lpr mice. 840 38


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