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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha) and Fas-ligand can mediate potent inhibitory signals in haemopoietic cells. Clinical and laboratory studies have suggested the involvement of these cytokines in the regulation of normal haemopoiesis and in the pathophysiology of bone marrow (BM) failure syndromes. As the effects of cytokines may also be regulated at the cellular receptor level, we studied the expression and modulation of TNF receptor (TNFR), IFN-gammaR and Fas-R on haemopoietic progenitor cells. In freshly isolated BM, using flow cytometry, TNFR1 (p55), TNFR2 (p75), IFN-gammaR, and Fas-R were detected on 5-12% of mononuclear cells. Two-colour staining showed comparable receptor expression on a CD34+ population, which includes haemopoietic progenitor and stem cells. Using reverse transcriptase-PCR (RT-PCR) transcription of mRNA coding for these receptors was demonstrated in fresh, highly purified CD34+ cells. These findings indicate that the effects of these factors on progenitor cells may be directly mediated. In cultured BM cells, expression of TNFR1 was not influenced by IFN-gamma, TNF-alpha or apoptosis-inducing anti-Fas monoclonal antibody (mAb). IFN-gamma decreased CD34+ cell TNFR2 expression. CD34+ cell Fas-R expression was increased by IFN-gamma and TNF-alpha. IFN-gammaR expression was enhanced by anti-Fas mAb and to lesser degree with TNF-alpha. Similar results were obtained with RT-PCR analysis in cultured CD34+ cells. Potentiation of anti-Fas mAb-mediated inhibition of haemopoietic colony formation by IFN-gamma and TNF-alpha was observed. Similarly, anti-Fas mAb enhanced the inhibitory effects of IFN-gamma. These results suggest that, in addition to interacting at the level of intracellular signalling pathways, IFN-gamma, TNF-alpha or Fas-ligand may potentiate or antagonize their effects through modulation of cytokine receptor expression.
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PMID:Expression and modulation of cellular receptors for interferon-gamma, tumour necrosis factor, and Fas on human bone marrow CD34+ cells. 916 2

Exposure of hematopoietic progenitor cells (HPC) from mice and humans with Fanconi anemia group C (FAC) to interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF-alpha) at doses too low to inhibit growth of normal HPC induces profound apoptotic responses. Because the IFN-gamma hypersensitivity of cells lacking the FAC protein is mediated, in part, through priming of the Fas pathway, and because several other members of this family are capable of inducing apoptosis either alone or in concert with each other, we tested the hypothesis that IFN-gamma induces increased expression of members of the TNF receptor (TNFR) superfamily in cells nullizygous for the FAC gene. Using isogenic human Epstein-Barr virus-transformed lymphoblast cell lines and c-kit+ bone marrow cells from mice with inactivating mutations of the FAC locus, we quantified mRNA levels by reverse transcriptase polymerase chain reaction and surface expression of the gene products by flow cytometry of TNFR1, TNFR2, Fas, CD30, CD40, and nerve growth factor receptor. We found that neither constitutive nor IFN-gamma-induced expression of these receptors was influenced by the absence of a functional FAC gene product, and expression of these receptors was not suppressed in nullizygous cells complemented with the normal FAC cDNA. We conclude that, although exaggerated apoptotic responses in FAC-deficient cells are at least partially mediated through activation of members of the TNFR superfamily, the normal FAC protein does not function as a direct suppressor of this family of molecules and inactivation of FAC does not augment expression of these proteins.
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PMID:The Fanconi anemia group C gene product modulates apoptotic responses to tumor necrosis factor-alpha and Fas ligand but does not suppress expression of receptors of the tumor necrosis factor receptor superfamily. 992 38

Cytokines induce apoptosis in pancreatic beta cells, but the exact mechanisms and sequence of events are not clear. Here, we investigate a role for tumor necrosis factor alpha (TNF-alpha) in the apoptosis of beta cells. Using the ribonuclease (RNase) protection assay and the reverse transcriptase-polymerase chain reaction (RT-PCR) method, we confirmed that TNF receptor 1 (TNFR1), TNFR1-associated death domain protein (TRADD), Fas receptor-associated intracellular protein with death domain (FADD), and FADD-like interleukin-1beta-converting enzyme (FLICE) were expressed in the pancreatic beta cell line, MIN6 cells. Fluorescent microscopic examination using Hoechst 33342 dye (Sigma, St Louis, MO) demonstrated that TNF-alpha induced time- and dose-dependent apoptotic nuclear changes in these beta cells. In situ end-labeling (ISEL) DNA analysis revealed that 10 nmol/L TNF-alpha generated new 3'-OH DNA strand breaks. Moreover, qualitative assessment of the induced DNA damage on agarose gels showed that 10 nmol/L TNF-alpha produced characteristic apoptotic patterns of DNA fragments formed by internucleosomal hydrolysis of static chromatin. In addition, C2-ceramides and natural ceramides dispersed in a solvent mixture of ethanol and dodecane induced characteristic features of apoptosis in MIN6 cells, mimicking TNF-induced DNA damage. We also determined endosomal ceramide production after TNF-alpha (10 nmol/L) treatment in MIN6 cells using the diacylglycerol kinase assay. These results suggest that TNF-alpha can cause apoptosis in pancreatic beta cells through TNFR1-linked apoptotic factors, TRADD, FADD, and FLICE, and TNF-induced ceramide production may be involved in the pathways.
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PMID:Tumor necrosis factor alpha signaling pathway and apoptosis in pancreatic beta cells. 1059 77

Membrane-associated tumor necrosis factor (mTNF) has recently been shown to induce inflammatory cellular responses previously attributed to the soluble form. The present study measures for the first time the expression and function of mTNF on the surface of alveolar macrophages (AMs) to determine whether it is associated with the development of acute respiratory distress syndrome (ARDS). TNF expression was determined by flow cytometry, and the function of mTNF on the surface of AMs was determined by an in vitro cytotoxicity assay. Tumor necrosis factor (TNF)-alpha bioactivity was measured by bioassay. Soluble TNF receptor (TNFR) protein and messenger RNA (mRNA) expression were measured by enzyme-linked immunosorbent assay and reverse transcriptase/polymerase chain reaction, respectively. Increased detection of mTNF was observed on the surface of AMs derived from subjects with ARDS (mean percentage increase in fluorescence 22.30 +/- 3.50% for subjects with ARDS compared with 7.09 +/- 1.70% for At Risk subjects [P < 0.003]). mTNF cytotoxicity in the bioassay positively correlated with the mTNF expression determined by flow cytometry (r(2) = 0.97). Although there was increased mTNF expression and cytotoxic function in ARDS, there was no significant increase in soluble TNF expression in the bronchoalveolar lavage fluid or the AM supernatants. Lower levels of CD120b-soluble TNFR were detected in the AM supernatants derived from subjects with ARDS compared with At Risk (mean 0.264 +/- 0.058 versus 0.593 +/- 0.143 ng/ml, respectively [P < 0.05]). By contrast, there was increased CD120b mRNA expression in AMs derived from subjects with ARDS (P < 0.03), suggesting that increased surface expression of this receptor may be important in mediating the signal of mTNF. These data demonstrate for the first time the presence of functionally active mTNF on the surface of AMs in ARDS and highlight a potential mechanism for TNF-mediated lung injury.
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PMID:Increased expression of functionally active membrane-associated tumor necrosis factor in acute respiratory distress syndrome. 1061 67

Keratinocyte-derived cytokines have been implicated in the pathogenesis of a number of skin diseases. In this study we examined the possible role of keratinocyte-derived cytokines in the development of acantholysis in pemphigus vulgaris. Nineteen patients with pemphigus vulgaris, demonstrating the characteristic clinical, pathologic, and immunopathologic findings were studied. In situ immunolabeling demonstrated the presence of two cytokines interleukin-1alpha and tumor necrosis factor-alpha, in lesional and perilesional areas. Results were confirmed by reverse transcriptase-polymerase chain reaction, demonstrating overexpression of both cytokines in vivo. To study the role of these cytokines in the pathogenesis of pemphigus vulgaris both in vitro and in vivo studies were performed. The results of the in vitro study demonstrated that pemphigus vulgaris IgG induced interleukin-1alpha and tumor necrosis factor-alpha mRNA in the skin. The potential pathogenic role of these mediators was demonstrated by a blocking study using antibodies against human interleukin-1alpha and tumor necrosis factor-alpha in keratinocytes cultures. A combination of anti-interleukin-1alpha and anti-tumor necrosis factor-alpha antibodies inhibited in vitro pemphigus vulgaris IgG induced acantholysis. To confirm the role of interleukin-1 and tumor necrosis factor-alpha in pemphigus, we utilized passive transfer studies using interleukin-1 deficient mice (ICE-/-, interleukin-1beta-/-) and tumor necrosis factor-alpha receptor deficient mice (TNFR1R2-/-). Both groups demonstrated a decreased susceptibility to the passive transfer of pemphigus. Our data support the role of cytokines interleukin-1 and tumor necrosis factor-alpha in the pathogenesis of pemphigus vulgaris.
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PMID:In vitro and in vivo expression of interleukin-1alpha and tumor necrosis factor-alpha mRNA in pemphigus vulgaris: interleukin-1alpha and tumor necrosis factor-alpha are involved in acantholysis. 1062 Jan 18

We investigated the effects of TNF receptor 1 (TNFR1) and 2 (TNFR2) modulation on the death of human aortic endothelial cells (HAECs) resistant to TNF-alpha-induced cell death. Alteration of the transcription of anti-apoptotic proteins, including inhibitor of apoptosis protein 1, 2 (cIAP1, 2), TNF receptor-associated factor 1 (TRAF1), nuclear factor kappa B1 protein (NFkappaB1), and FLICE-inhibitory protein (FLIP) was assessed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). TNF-alpha (200 ng/ml) or actinomycin D (ActD) (5 ng/ml) did not kill cells, while 5 ng/ml of TNF-alpha and 5 ng/ml of ActD increased expression of Fas (CD95) and FasL (CD95L), and 45% of cells died. TNFR2 blockade suppressed NFkappaB1 and FLIP expression and increased cell death. TNFR1 blockade enhanced FLIP expression and decreased cell death. Cells insensitive to TNF-alpha may respond to TNF-alpha through TNFR that induces transcription of NFkappaB1 and FLIP. Down-regulation of these proteins may facilitate death of cells insensitive to TNF-alpha-induced cell death.
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PMID:Reduced expression of flice-inhibitory protein (FLIP) and NFkappaB is associated with death receptor-induced cell death in human aortic endothelial cells (HAECs). 1150 81

The osteoclast (OCL) is the primary bone resorbing cell. OCL formation and activity is regulated by local factors produced in the bone microenvironment. We recently identified OCL inhibitory peptide-1 (OIP-1/ hSca) as a novel inhibitor of OCL formation and bone resorption that is produced by OCLs. OIP-1 is a glycosylphosphatidyl-inositol (GPI)-linked membrane protein (16 kDa) related to the mouse Ly-6 family of hematopoietic proteins. OIP-1 mRNA is expressed in human OCL precursors, granulocyte-macrophage colony-forming unit (GM-CFU), bone marrow cells, and osteoblast cells. We used cycle-dependent reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, which further demonstrated that interferon-gamma (IFN-gamma) strongly enhanced OIP-1/hSca mRNA expression in bone marrow cells and GM-CFU. Similarly, interleukin (IL)-1beta also enhanced OIP-1 mRNA expression in GM-CFU. To determine the participation of OIP-1 in IFN-gamma inhibition of OCL formation, we tested the capacity of a neutralizing antibody specific to OIP-1 c-peptide to inhibit IFN-gamma's effects on OCL-like cell differentiation of mouse macrophages, RAW 264.7 cells. Anti-OIP-1 c-peptide specific antibody partially neutralized IFN-gamma inhibition of OCL differentiation. Furthermore, OIP-1 inhibited phospho-c-Jun (p-c-Jun) kinase activity in RAW 264.7 cells. However, OIP-1/hSca did not affect NF-kappaB activation in these cells. Western blot analysis further demonstrated that OIP-1 significantly decreased TNF receptor associated factor 2 (TRAF-2) expression in RAW 264.7 cells. However, OIP-1 had no effect on TRAF-6 expression in these cells. These data show that IFN-gamma enhances OIP-1/hSca expression in OCL precursors, GM-CFU, and that OIP-1 inhibits OCL formation through suppression of TRAF-2 and p-c-Jun kinase activity.
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PMID:CytokineRegulation and the signaling mechanism of osteoclast inhibitory peptide-1 (OIP-1/hSca) to inhibit osteoclast formation. 1261 30

Myelodysplastic syndromes (MDS) are characterized by peripheral blood cytopenias despite hypercellularity of the bone marrow regarded as the result of ineffective hematopoiesis mainly caused by apoptosis. In this study, we examined the role of tumor necrosis factor (TNF)-induced apoptosis in the bone marrow cells of MDS patients. The constitutive expression of mRNA for TNF receptors (TNFR), including TNFRI and TNFRII, and the adapter molecules, such as the TNF receptor-associated death domain protein (TRADD), Fas-associated death domain protein (FADD), receptor interacting protein (RIP) and TNF receptor-associated factor 2 (TRAF-2) were analyzed by reverse transcriptase (RT)-PCR in bone marrow samples from control, MDS and AML cases. In bone marrow cells from refractory anemia (RA) patients, there was a significant increase in TNFRI expression as compared with control subjects. The expression of TNFRII was also up-regulated in RA cases. In contrast, RA with excess of blasts (RAEB), RAEB in transformation (RAEB-T) and AML cases revealed increased expression of TNFRII, whereas the expression of TNFRI was comparable to control subjects. Immunohistochemical staining revealed that the TNFRI, as well as TNFRII of MDS bone marrow was expressed mainly in hematopoietic cells, but not in macrophage-lineage stromal cells at the protein level. An increased constitutive expression of mRNA for TRADD, FADD and RIP and decreased expression of mRNA for TRAF-2 were observed in bone marrow cells from MDS patients, especially from RA patients, as compared with controls, although the differences were not significant. In many of the AML bone marrow samples, strong expression of TRAF-2 mRNA was marked, while expression levels of other proteins were similar to those in control subjects. These data suggested enhanced signaling by the TNFRI-TRADD-FADD pathway and suppressed signaling by the TRAF-2 pathway in RA. Thus, TNF-alpha-induced apoptosis may play a role in ineffective hematopoiesis in "early stage MDS" bone marrow, although the regulatory mechanisms for TNF-alpha-induced signaling would be complicated and not be simply explained only by these pathways.
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PMID:Expression of TNF receptors and related signaling molecules in the bone marrow from patients with myelodysplastic syndromes. 1268 57

Apoptosis-inducing tumor necrosis factor (TNF) ligands and receptors have been reported in human placentas, but the expression patterns of family members lacking this function [a proliferation-inducing ligand (APRIL), B lymphocyte stimulator (BLyS), CD30L/CD153, CD40L/CD154, TNF-related activation-induced cytokine, CD27L/CD70, OX40L, activation-inducible TNF receptor ligand (AITRL)] are incompletely documented or unknown. We therefore investigated expression of these eight ligands and nine of their receptors (B cell maturation antigen, transmembrane activator and calcium-modulator and cyclophilin ligand-interactor, CD30, CD40, receptor activator of nuclear factor-kappaB, osteoprotegerin, CD27, OX40/CD134, AITR). Analysis by reverse transcriptase-polymerase chain reaction revealed mRNAs encoding only three of the ligands (APRIL, BLyS, CD30L/CD153). Immunoblots demonstrated all three proteins in first-trimester and term placentas, and immunohistochemical experiments showed that expression was cell-specific and gestation-related. Although mRNAs encoding receptors for the three expressed ligands were absent, those encoding receptors for all of the unexpressed ligands were detectable. Collectively, the results are consistent with the postulate that nonapoptosis-inducing, placenta-derived TNF superfamily cytokines contribute to the T helper cell type 2 bias required for successful pregnancy. Patterns of placental expression of receptors suggest bidirectional maternal-fetal cytokine communication.
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PMID:Cell-specific expression of B lymphocyte (APRIL, BLyS)- and Th2 (CD30L/CD153)-promoting tumor necrosis factor superfamily ligands in human placentas. 1283 45

Tumor necrosis factor-alpha (TNF-alpha) is a cytokine considered to play a key role in beta-cell destruction in insulin-dependent diabetes mellitus (IDDM). Serum thymic factor (Facteur thymique serique; FTS) is a nonapeptide thymus hormone known to inhibit IDDM in a mouse model. In this study, the effect of TNF-alpha on the murine pancreatic beta-cell line MIN6 was examined. Cell shrinkage and detachment were seen in cells treated with 0-50 ng/ml TNF-alpha for 12h. Oligonucleosomal DNA fragmentation was determined from non-adherent cells, indicating that the TNF-alpha-induced cell destruction was attributed to apoptosis. Fragmented DNA was quantified by enzyme-linked immunosorbent assay to measure the amount of histone-bound oligonucleosomes. FTS was treated with TNF-alpha and the percentage of fragmented DNA was analyzed. The data indicate a distinct reduction of fragmented DNA at a concentration of 1 ng/ml FTS. Expression of TNF receptor I, inducible form of nitric oxide synthase (iNOS), interleukin-1 beta-converting enzyme (ICE), Bcl-2, and nuclear factor kappa B (NF-kappa B) was analyzed by reverse transcriptase-polymerase chain reaction to investigate the suppressor mechanism of FTS on TNF-alpha-induced apoptosis. FTS treatment suppressed the expression of iNOS and Bcl-2 mRNA in TNF-alpha-treated cells. The expression of NF-kappa B mRNA in TNF-alpha-treated cells was enhanced after FTS treatment, while that of ICE mRNA did not change in TNF-alpha-treated cells with or without FTS treatment. These results suggest that the inhibition of MIN6 cell death by FTS on TNF-alpha-induced apoptosis is caused by a negative feedback mechanism involving the inhibition of iNOS induction.
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PMID:Suppressor mechanism of serum thymic factor on tumor necrosis factor-alpha-induced apoptosis in the mouse pancreatic beta-cell line. 1459 44


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