Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interleukin (IL)-12 is a heterodimeric cytokine consisting of 35 and 40 kDa subunits, produced primarily by phagocytic cells in response to bacteria or bacterial products. IL-12 is important in the regulation of both innate and antigen-specific immunity through its stimulatory effects on NK cells and cytotoxic lymphocytes. Reverse transcriptase-polymerase chain reaction with primers derived from human sequence was used to clone the p35 and p40 subunits of porcine IL-12. Predicted amino acid sequences for both subunits are approximately 85% homologous to their human cognates but contain a 3aa addition and a 4aa deletion in p35 and p40 subunits, respectively. The high degree of similarity indicates the proteins may be cross reactive, an important consideration in pig-human xenotransplantation. Both subunits of pIL-12 are constitutively expressed in a variety of porcine tissues. Highest levels of the p40 subunit were found in lymphoid tissues including inguinal and mesenteric lymph nodes, Peyer's patches, spleen and thymus. The p35 subunit was also detected in these tissues. Levels of mRNA encoding the p40 subunit, but not the p35 subunit, were rapidly increased in alveolar macrophages stimulated with lipopolysaccharide or killed Staphylococcus aureus. Thus, the heterodimeric subunits appear to be differentially regulated at the transcriptional level. Since p40 also self-associates to form inactive homodimers, differential expression may be a mechanism for regulating IL-12 activity.
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PMID:Molecular cloning and mRNA expression of porcine interleukin-12. 923 44

IL-12, a 75-kDa heterodimeric cytokine composed of two chains (p35 and p40), is a central regulator of immune responses and may be implicated in the pathogenesis of certain inflammatory diseases of the central nervous system (CNS). We have examined the capacity of two CNS APC, microglia and astrocytes, to produce IL-12 upon stimulation with cytokines, LPS, or a neurotropic virus. In purified microglial cultures from neonatal mouse brains, expression of IL-12 p35 and p40 mRNA is induced by LPS and is stimulated maximally by combined IFN-gamma/LPS treatment, as detected by semiquantitative reverse-transcriptase PCR. LPS induces secretion of IL-12 p40, but not of IL-12 p75, as detected by specific ELISA. Combined stimulation with IFN-gamma/LPS enhances IL-12 p40 secretion and induces IL-12 p75 secretion by microglia. Conversely, mouse astrocytes do not express IL-12 p35 mRNA and do not secrete IL-12 p75 under any condition tested. IL-12 production by activated microglia is inhibited by IL-10, PGE2, and cAMP-elevating agents. Coculture of microglia with astrocytes or exposure of microglia to astrocyte-conditioned medium also results in marked reduction of IL-12 p75 and p40 secretion by IFN-gamma/LPS-stimulated microglia, indicating a regulatory role of astrocytes on IL-12 production. This novel mechanism of IL-12 regulation may play an important role in the control of immune responses during infection or in Th1 cell-mediated autoimmune diseases of the CNS.
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PMID:IL-12 production by central nervous system microglia is inhibited by astrocytes. 925 19

In the present study, we have analyzed the pattern of cytokines expressed by two independent dendritic cell (DC) subpopulations generated in vitro from human cord blood CD34+ progenitors cultured with granulocyte-macrophage CSF and TNF-alpha. Molecularly, we confirmed the phenotypic differences discriminating the two subsets: E-cadherin mRNA was only detected in CD1a+-derived DC, whereas CD68 and factor XIIIa mRNAs were observed exclusively in CD14+-derived DC. Semiquantitative reverse-transcriptase PCR analysis revealed that both DC subpopulations spontaneously expressed IL-1alpha, IL-1beta, IL-6, IL-7, IL-12 (p35 and p40), IL-15, IL-18, TNF-alpha, TGF-beta, macrophage CSF, and granulocyte-macrophage CSF, but not IL-2, IL-3, IL-4, IL-5, IL-9, and IFN-gamma transcripts. Both subpopulations were shown to secrete IL-12 after CD40 triggering. Interestingly, only the CD14+-derived DC secreted IL-10 after CD40 activation, strengthening the notion that the two DC subpopulations indeed represent two independent pathways of DC development. Furthermore, both DC subpopulations expressed IL-13 mRNA and protein following activation with PMA-ionomycin, but not with CD40 ligand, in contrast to IL-12 and IL-10, revealing the existence of different pathways for DC activation. Finally, we confirmed the expression of IL-7, IL-10, and IL-13 mRNA by CD4+ CD11c+ CD3- DC isolated ex vivo from tonsillar germinal centers. Thus, CD14+-derived DC expressing IL-10 and factor XIIIa seemed more closely related to germinal center dendritic cellsGCDC than to Langerhans cells.
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PMID:The cytokine profile expressed by human dendritic cells is dependent on cell subtype and mode of activation. 946 23

Epidermal cytokines such as interleukin (IL)-1beta, tumor necrosis factor-alpha, and IL-12 have been described to play a crucial role in the induction and elicitation phase of allergic contact dermatitis upon exposure to haptens. In this study we asked whether these cytokines may also play a role in the epidermis of patients with atopic dermatitis after the application of house dust mite antigens (HDM) to their skin. Epidermal samples were collected by scraping healthy appearing skin of atopic patients and healthy individuals 8 h after the application of an extract of HDM. Sodium lauryl sulfate and saline served as controls. Reverse transcriptase-polymerase chain reaction was performed for IL-1beta, tumor necrosis factor-alpha, IL-12 p35, and IL-12 p40. Exposure to HDM led to a significant upregulation of mRNA of these cytokines in atopic patients only. Whereas IL-1beta and tumor necrosis factor-alpha also showed an upregulation in part of these patients after exposure to the irritant sodium lauryl sulfate, IL-12 p40 mRNA was exclusively enhanced by the application of the allergen. In contrast to IL-12 p40, IL-12 p35 mRNA was not detectable in significant amounts. Interestingly, also in untreated, normal appearing skin of atopic individuals (n = 16), the levels of these cytokines were higher than in normal individuals (n = 8), possibly explaining the increased skin irritability of atopic individuals. Finally, comparing epidermal cytokines in the skin of patients who developed a positive allergen patch test to those who stayed negative, suggests that only expression of IL-1beta mRNA may be a predictive marker for the development of a positive patch test reaction to HDM.
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PMID:Epidermal cytokines IL-1beta, TNF-alpha, and IL-12 in patients with atopic dermatitis: response to application of house dust mite antigens. 985 37

The genome of Sapovirus (SaV), a causative agent of gastroenteritis in humans and swine, contains either two or three open reading frames (ORFs). Functional motifs characteristic to the 2C-like NTPase (NTPase), VPg, 3C-like protease (Pro), 3D-like RNA-dependent RNA polymerase (Pol), and capsid protein (VP1) are encoded in the ORF1 polyprotein, which is afterwards cleaved into the nonstructural and structural proteins. We recently determined the complete genome sequence of a novel human SaV strain, Mc10, which has two ORFs. To investigate the proteolytic cleavage of SaV ORF1 and the function of protease on the cleavage, both full-length and truncated forms of the ORF1 polyprotein either with or without mutation in (1171)Cys to Ala of the GDCG motif were expressed in an in vitro coupled transcription-translation system. The translation products were analyzed directly by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or by immunoprecipitation with region-specific antibodies. The ORF1 polyprotein was processed into at least 10 major proteins: p11, p28, p35, p32, p14, p70, p60, p66, p46, and p120. Seven of these products were arranged in the following order: NH(2)-p11-p28-p35(NTPase)-p32-p14(VPg)-p70(Pro-Pol)-p60(VP1)-COOH. p66, p46 and p120 were precursors of p28-p35 (NTPase), p32-p14 (VPg), and p32-p14 (VPg)-p70 (Pro-Pol), respectively. Mutagenesis in the 3C-like protease motif fully abolished the proteolytic activity. The cleavage map of SaV ORF1 is similar to those of other heretofore known members of the family Caliciviridae, especially to rabbit hemorrhagic disease virus, a member of the genus Lagovirus.
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PMID:Proteolytic processing of sapovirus ORF1 polyprotein. 1591 82

We evaluated the effect of interleukin-12 (IL-12) gene therapy using an Ewing's sarcoma animal model in T-cell-deficient nude mice. Subcutaneous injection of TC71 cells resulted in tumor development by day 5. Mice were treated with a single intratumor injection of adenovirus beta-galactosidase (Ad.beta-gal) or adenovirus murine IL-12 (Ad.mIL-12) (2 x 10(9) PFU) and killed 1-7 days later. Reverse transcriptase-polymerase chain reaction analysis of tumor tissue demonstrated peak expression of IL-12 p35 and p40 at 48 h, which persisted up to 7 days. For in vivo therapy, mice received intratumor Ad.beta-gal or Ad.mIL-12 twice weekly for 2.5 weeks starting on day 6. Ad.mIL-12-treated tumors were significantly smaller (median volume, 19.7 mm3; range, 3.41-159.5 mm3) than Ad.beta-gal-treated tumors (median volume, 3214.9 mm3; range 1679.9-5909.8 mm3, P<0.003) on day 31. The weight of Ad.mIL-12-treated tumors was also lighter than the Ad.beta-gal-treated tumors (median, 2 mg; range, 1-5 mg versus median, 1960 mg; range 1640-5230 mg, P<0.01). Ad.mIL-12 therapy significantly prolonged the survival time and also inhibited the growth of an untreated tumor on the contralateral side. Immunohistochemistry analysis of the IL-12-treated tumors demonstrated IL-12 expression with increased Fas, Fas ligand and tumor cell apoptosis. CD31 and vascular endothelial growth factor expression were decreased. These data suggest that IL-12 gene therapy may be useful in the treatment of Ewing's sarcoma.
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PMID:Intratumor murine interleukin-12 gene therapy suppressed the growth of local and distant Ewing's sarcoma. 1676 9

SKIN(2) ZK1300 is a three-dimensional human skin model consisting of multilayered dermal fibroblasts and well-differentiated epidermal keratinocyte layers, including a stratum corneum. To characterize this model better, constitutive levels of cytokine gene expression were determined. Reverse transcriptase-polymerase chain reaction (RT-PCR), followed by liquid hybridization to labelled internal probes, demonstrated that interleukin (IL)-1I, IL-1beta, IL-6, IL-8, IL-10, tumour necrosis factor (TNF)I, granulocyte macrophage-colony stimulating factor (GM-CSF), transforming growth factor (TGFbeta1) and IL-12 p35 mRNAs were constitutively expressed whereas IL-12 p40 was not. The contribution of the dermal component of this human skin model (Model ZK1100) was further characterized by determining constitutive cytokines expressed and their modulation by phorbol 12-myristate, 13-acetate (PMA). The dermal component, consisting of multilayered human dermal fibroblasts, constitutively expressed message for IL-1I, 1L-1beta, IL-6, IL-8, TGFbeta1, GM-CSF and IL-12 p35. Message was not detected for IL-10, TNFI or IL-12 p40. PMA treatment of the multilayered dermal fibroblasts increased steady-state mRNA levels of IL-1I, IL-1beta, IL-6, IL-8, GM-CSF and TGFbeta1, but did not induce IL-10, TNFI or IL-12 p40 expression at the dose and times tested. In summary, these studies demonstrate that the SKIN(2) three-dimensional human skin cultures, and their dermal component, constitutively express mRNA for an array of inflammatory and immunomodulatory cytokines, and that PMA exposure modulates mRNA levels of the dermal cytokines.
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PMID:Cytokine mRNA expression in an in vitro human skin model, SKIN(2). 2065 Feb 32