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)

Similar to interleukin-3 (IL-3), IL-5, and granulocyte macrophage colony-stimulating factor (GM-CSF), IL-4 can be secreted by several cell types involved in allergic inflammatory reactions, and therefore can affect eosinophil function similarly. In this study, we investigated the presence of an IL-4 receptor (IL-4R) on human eosinophils. When two different monoclonal antibodies (mAbs) against the IL-4R alpha-chain (IL-4Ralpha) were used, fluorescent-activated cell sorter analysis revealed the presence of an IL-4Ralpha on both eosinophils of normal donors and atopic dermatitis patients. In addition, the expression of the IL-2R gamma-chain, a functional component of the IL-4R in some cell types, was demonstrated. The IL-4Ralpha appeared to be expressed constitutively, and stimulation with cytokines IL-2, IL-3, IL-5, GM-CSF, and interferon-gamma did not further increase IL-4Ralpha expression. Evidence for an IL-4Ralpha was further substantiated by mRNA analysis. Both Northern blot analysis and reverse transcriptase/polymerase chain reaction revealed the presence of mRNA for the IL-4Ralpha in eosinophils from normal individuals and AD patients. Furthermore, we demonstrated that both IL-4 and IL-13 were capable of inducing PI-3 kinase activity in human eosinophils. Because this activation could be inhibited by an IL-4Ralpha mAb, we conclude that both cytokines can activate human eosinophils through binding to a receptor complex comprising the IL-4Ralpha and-yet to be identified-associated proteins. In addition, the involvement of IL-4 in functional responses was studied. IL-4 appeared to "prime" eosinophils to respond chemotactically toward regulated on activation, normal T cells expressed and secreted, but did not affect platelet-activating factor-induced chemotaxis. Taken together, these data show the presence of a functional IL-4R on human eosinophils.
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PMID:Human eosinophils constitutively express a functional interleukin-4 receptor: interleukin-4 -induced priming of chemotactic responses and induction of PI-3 kinase activity. 976 67

The cyclin-dependent kinase inhibitor (CDKI) p15INK4B (p15) induces cell cycle arrest in G0/G1 phase. Several studies report deletion or transcriptional loss of the p15 gene in myeloid and lymphoid hematological malignancies, and a possible role as a tumor suppressor gene has been proposed for this CDKI. In this study we evaluated the expression of p15 by cytofluorometric, immunohistochemical, and reverse transcriptase-polymerase chain reaction (RT-PCR) methods in CD34+ progenitors (both during steady state and after chemotherapy and/or granulocyte-colony stimulating factor [G-CSF] administration) and in cells belonging to different hematopoietic differentiative lineages. We found that p15 is not expressed in normal G0/G1-arrested peripheral blood (PB)- or bone marrow (BM)-CD34+ cells. Moreover, p15 is expressed in G0/G1-blocked CD34+ cells mobilized by chemotherapy and G-CSF but not in CD34+ cells mobilized by G-CSF alone. To clarify the role of p15 in normal hematopoiesis, we used flow cytometry to investigate its expression in normal differentiating BM and PB cells. We found that p15 was expressed in cells belonging to the granulocyte-monocyte lineage and in B and T lymphocytes, whereas erythroid and megakaryocytic cells were p15 negative. These findings, which were confirmed both by immunohistochemical and RT-PCR analysis, definitely establish a linkage between p15 expression and granulocyte-monocyte differentiation.
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PMID:Expression of p15INK4B in normal hematopoiesis. 980 52

The products of the human Arg gene and human, mouse, Drosophila, and nematode Abl genes characterize the Abelson family of nonreceptor tyrosine protein kinase. The Arg gene, expressed as a 12-kb transcript, codes a protein highly related to c-abl in the tyrosine kinase, SH2, and SH3 domains, and both proteins have a myristoylated isoform. The C-terminal domains of Arg and c-abl, poorly similar to each other, may account for their different functions. Arg is cytoplasmic, c-abl also has nuclear localization, and their products have different transforming activity. To gain insight about the role of Arg in myeloid differentiation we investigated Arg gene expression in HL-60 cells differentiated with all-trans retinoic acid and 12-O-tetradecanoyl-phorbol-13-acetate. With a semiquantitative reverse transcriptase-polymerase chain reaction assay it was evident that the Arg transcript level in HL-60 cells differentiated toward granulocyte and macrophage-like lineage was, respectively, 3.5- and 2.8-fold the Arg level evidenced in undifferentiated HL-60 cells. In the HL-60 cells, under the same differentiating conditions, the c-abl RNA level did not change significantly, showing that Arg and c-abl responded in a different way to the inducers of differentiation used.
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PMID:Increment of nonreceptor tyrosine kinase Arg RNA as evaluated by semiquantitative RT-PCR in granulocyte and macrophage-like differentiation of HL-60 cells. 982 10

Indoor air pollutants may cause inflammatory changes of the airways and adjacent pulmonary tissue. After phagocytosis of inhaled particles alveolar macrophages (AM) release chemotactic mediators capable of attracting inflammatory cells into the lung tissue. To evaluate these mechanisms further peripheral blood mononuclear cells (PBMNC) and human AM (freshly recovered from the lower respiratory tract) were exposed to the indoor particles Soot FR 101 and Printex 90, the asbestos fiber Chrysotile B, and titanium dioxide (TiO2) at concentrations of 10 or 50 microg/10(6) cells for up to 8 h. The migration of granulocytes into the conditioned supernatants of AM and PBMNC was quantified by chemotaxis assay in a Boyden chamber. Granulocyte migration increased by 42.3 +/- 25.8% (Chrysotile B), 64. 6 +/- 18.3% (FR 101), 74.2 +/- 16.5% (P 90), and 86.7 +/- 25.6% (TiO2) in AM-conditioned supernatants (p < 0.05). Qualitative, Interleukin (IL)-8 specific reverse transcriptase-polymerase chain reaction was performed after exposure of AM or PBMNC to Chrysotile B, FR 101, P 90, and TiO2 at concentrations of 10 and 50 microg/10(6) cells for 90 min. Each of the tested particles caused an increase in IL-8-specific mRNA expression of AM or PBMNC after particle exposure compared with the unexposed control. To find out if IL-8, the most powerful granulocyte chemokine, is involved, supernatants were preincubated with anti-IL-8. Granulocyte migration decreased by up to 35 +/- 15% (50 ng/ml anti-IL-8) and 41.5 +/- 16% (100 ng/ml anti-IL-8) (p < 0.0625) in AM-conditioned supernatants. Pretreatment of the granulocytes with human IL-8 decreased by up to 59 +/- 18% (10 ng/ml) (p < 0.0625) in AM-conditioned supernatants. Similar reaction patterns were observed using anti-IL-8-pretreated supernatants of particle-exposed PBMNC. In conclusion, indoor air pollutants may promote inflammatory changes in the lung via IL-8 release by alveolar macrophages.
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PMID:Indoor air pollutants stimulate interleukin-8-specific mRNA expression and protein secretion of alveolar macrophages. 983 30

There is considerable interest in an autologous transplantation (AT) programme for patients with high-risk breast cancer; however, the issue of the incidence of occult bone marrow (BM) micrometastasis at diagnosis, and the cancer contamination of peripheral blood stem cell (PBSC) collections used for haematological rescue, is still debated. The presence of BM micrometastasis was evaluated in bilateral BM biopsies obtained at diagnosis of 33 patients with stage II/IIIA breast cancer using: (i) a 'nested' reverse transcriptase-polymerase chain reaction (RT-PCR) assay for cytokeratin 19 (K19) mRNA, (ii) histology, and (iii) immunohistochemistry (IHC) analysis with a panel of three monoclonal antibodies. The RT-PCR assay only was used to determine contamination of PBSC collections obtained after priming with recombinant human granulocyte-colony stimulating factor (rhG-CSF). K19 transcripts in one or both BM samples were detected in 48% of patients at diagnosis, with an overall 85% concordance with the results of IHC analysis. On the other hand, 56% of PCR- and IHC-positive BM samples were diagnosed as 'normal' on histological analysis. 57% of patients showed K19 mRNA in at least one PBSC collection; the possibility to have contaminated PBSC collections was significantly higher in patients with K19 positivity in BM at diagnosis. In four patients who had shown K19 positivity in BM and in PBSC collections, immunoselected CD34+ cells used for haematological rescue were K19-negative. There was a trend towards longer relapse free survival (RFS) in patients transplanted with K19-negative PBSC collections as compared to the others. In conclusion, a substantial proportion of patients with high-risk non-metastatic breast cancer present occult BM micrometastasis at diagnosis and also show cancer contamination of PBSC collections used for AT. These might represent a category of patients with poorer prognosis after AT, and possible candidates for more intensive and/or alternative therapeutic regimens, including AT with purged PBSCs.
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PMID:Evaluation of breast tumour cell contamination in the bone marrow and leukapheresis collections by RT-PCR for cytokeratin-19 mRNA. 985 8

Human leukemia (HL) 60 cells were differentiated by dimethylsulfoxide (DMSO) treatment to granulocyte-like cells, leukotriene (LT) synthesizing activity of which was increased in response to the differentiation of the cells. Four synthesizing enzymes, cytosolic phospholipase A2 (cPLA2), 5-lipoxygenase (5-LO), LTA4 hydrolase and LTC4 synthase, and an enzyme associated protein, 5-lipoxygenase activating protein (FLAP) are involved in the generation of LTC4 and LTB4. We examined the expression of messenger RNA (mRNA) for these LT synthesizing enzymes and an associated protein in DMSO differentiated HL-60 cells by reverse transcriptase polymerase chain reaction (RT-PCR). The production of LTC4 and LTB4, measured by radioimmunoassay (RIA), was increased after the incubation with DMSO for more than 3 days. Messenger RNA abundance for 5-LO, LTC4 synthase and LTA4 hydrolase was increased, that for FLAP was stable, but that for cPLA2 was decreased. These results indicate that DMSO induced increase of LT synthesis is associated with the increase of mRNA expression of 5-LO, LTC4 synthase and LTA4 hydrolase, although the precise regulatory mechanisms of the increased mRNA expression are not determined. We also investigated an action of dexamethasone (DEX) on DMSO-induced enhancement of LT synthesis. DEX suppressed DMSO induced increase of LTC4 synthesis, but rather enhanced DMSO induced LTB4 production. The DEX attenuated the DMSO-induced increase of mRNA expression for LTC4 synthase, but showed no effect on that for LTA4 hydrolase. The inhibition of LTC4 synthesis is associated with the suppression of mRNA expression for LTC4 synthase. However, increased LTB4 synthesis by DEX is regulated by the mechanisms which are independent from mRNA level of LTA4 hydrolase.
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PMID:Effect of dexamethasone on leukotriene synthesis in DMSO-stimulated HL-60 cells. 1010 84

The influence of the haematopoietic growth factor Flt-3 ligand (FL) on the incidence and function of donor major histocompatibility complex (MHC) class II+ cells in the lymphoid tissues of noncytoablated recipients of heart allografts and donor bone marrow (BM) cells was investigated. C3H (H2k) mice received a nonvascularized B10 (H2b) heart allograft in the dorsal ear pinna, followed by an i.v. infusion of 50 x 10(6) donor BM cells. They were given FL (10 microg/day i.p., x7 days), tacrolimus (2mg/kg/day i.p., x13 days) or both agents immediately following heart transplantation (HTx) and were killed 10 or 21 days later. Their BM cells were propagated in vitro in granulocyte macrophage colony stimulating factor (GM-CSF) and IL-4 for 5 days to promote the growth of dendritic cells (DC). Donor DC were identified by immunocytochemical staining. Spleens were harvested, and donor (IAb+) cells enumerated by immunohistochemical analysis. Donor MHC class II DNA was detected in spleens and cultured BM-derived cells by reverse transcriptase-polymerase chain reaction (RT-PCR). A striking increase in donor MHC class II+ cells was noted in both the spleen and BM of the BM + tacrolimus-treated group compared to either the BM alone, or BM + FL-treated groups. Addition of FL treatment to BM + tacrolimus led to a further increase in donor cells in spleen (three-fold at 10 days, and two-fold at 21 days). The increase in donor cells at 10 days was almost 140-fold compared to that with donor BM alone. PCR analysis at this time revealed enhanced donor DNA in the BM + FL + tacrolimus group compared to that in the BM + tacrolimus group. FL treatment augmented mixed leucocyte reactions (MLR) and cytotoxic T lymphocyte (CTL) activity of host spleen cells against donor alloantigens. These effects were reversed by tacrolimus administration. Histopathology of heart grafts from tacrolimus-treated animals at 10 and 21 days showed absence or substantial reduction in cellular infiltration, and the preservation of viable myocardium. By contrast, in untreated mice, or animals given BM or BM + FL alone, there was marked cellular infiltration, and features of accelerated rejection. Donor-derived DC could be propagated in vitro from the BM of heart transplant recipients given donor BM, especially from mice that also received tacrolimus +/- FL. At day 21, donor-derived cells could only be propagated from the BM + FL + tacrolimus-treated group. These findings show that numbers of donor antigen presenting cells (APC) or their progenitors can be markedly increased in conventionally immunosuppressed organ allograft recipients given donor BM + a potent haematopoietic and DC-growth promoting cytokine. Although withdrawal of systemic immunosuppression appears to allow exhibition of the potential allostimulatory activity of these donor APC leading to rejection, the model provides a useful basis for further evaluation of the persistence and manipulation of donor haematopoietic cells and in particular, donor-derived APC, on the outcome of organ transplantation.
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PMID:Impact of Flt-3 ligand on donor-derived antigen presenting cells and alloimmune reactivity in heart graft recipients given adjuvant donor bone marrow. 1034 36

We report the cellular characteristics of cells from three patients with de novo acute myelocytic leukaemia (AML) with t(16;21)(p11;q22), two M4 and one M5a according to the FAB classification, and two permanent cell lines with t(16;21)(p11;q22), TSU1621MT and YNH-1. The FUS/ERG fusion mRNA was demonstrated in all cases by reverse transcriptase-polymerase chain reaction (RT-PCR). The immunophenotypes of the AML cells, and YNH-1 and TSU1621MT cell lines with t(16;21) were characterized as CD34+CD33+CD13+CD11b+CD18+CD56+ HLA-DR-/+. Cells from all samples strongly expressed c-kit, granulocyte colony-stimulating factor receptor (G-CSFR), c-fms (macrophage colony-stimulating factor receptor), interleukin-3 receptor alpha chain (IL-3Ralpha), and granulocyte macrophage colony-stimulating factor receptor alpha chain (GM-CSFRalpha), and these data corresponded well to the growth responsiveness to the cytokines. IL-2Ralpha expression was also found in all t(16;21) samples, but IL-2 did not act on the proliferation of the leukaemic cells in in vitro cultures. G-CSF distinctly promoted the proliferation of leukaemic cells of t(16;21) AML, but did not enhance the expression of MPO and neutrophil differentiation of these cells. Our findings indicate that AML cells with t(16;21) preserve stem cell properties such as CD34 and c-kit expression, and suggest that they have the potential to differentiate into a monocytic lineage. The relationship between the unique cellular characteristics (especially CD56 and IL-2Ralpha expression) and FUS/ERG protein remains undetermined.
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PMID:Myeloid differentiation antigen and cytokine receptor expression on acute myelocytic leukaemia cells with t(16;21)(p11;q22): frequent expression of CD56 and interleukin-2 receptor alpha chain. 1035 36

We have previously demonstrated that short-term exposure to diesel exhaust (DE) for 1 h induced a marked leukocytic infiltration in the airways of healthy human volunteers involving neutrophils, lymphocytes, and mast cells along with increases in several inflammatory mediators. We hypothesized that the leukocyte infiltration and the various inflammatory responses induced by DE were mediated by enhanced chemokine and cytokine production by resident cells of the airway tissue and lumen. To investigate this, 15 healthy human volunteers were exposed to diluted DE and air on two separate occasions for 1 h each in an exposure chamber. Fiberoptic bronchoscopy was performed 6 h after each exposure to obtain endobronchial biopsies and bronchial wash (BW) cells. Using reverse transcriptase/polymerase chain reaction enzyme-linked immunosorbent assay (RT-PCR ELISA), a novel and sensitive technique to quantify relative amounts of cytokine mRNA gene transcripts, and immunohistochemical staining with computer-assisted image analysis to quantify expression of cytokine protein in the bronchial tissue, we have demonstrated that DE enhanced gene transcription of interleukin-8 (IL-8) in the bronchial tissue and BW cells along with increases in IL-8 and growth-regulated oncogene-alpha (GRO-alpha) protein expression in the bronchial epithelium, and an accompanying trend toward an increase in IL-5 mRNA gene transcripts in the bronchial tissue. There were no significant changes in the gene transcript levels of interleukin-1B (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), interferon gamma (IFN-gamma), and granulocyte macrophage colony-stimulating factor (GM-CSF) either in the bronchial tissue or BW cells after DE exposure at this time point. These observations suggest an underlying mechanism for DE-induced airway leukocyte infiltration and offer a possible explanation for the association observed between ambient levels of particulate matter and various respiratory health outcome indices noted in epidemiological studies.
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PMID:Acute exposure to diesel exhaust increases IL-8 and GRO-alpha production in healthy human airways. 1067 99

An in vitro culture system to generate human osteoclasts (OC) was recently described in which OC precursors in the human peripheral blood mononuclear cell (PBMC) population differentiate in the presence of murine ST-2 stromal cells. We used this culture system to define the cytokine environment in which human OC form and to determine the separate contributions of the stromal and hematopoietic elements. We designed a panel of reverse transcriptase-polymerase chain reaction (RT-PCR) primers that specifically amplify the respective murine or human mRNA species that correspond to cytokines and their cognate receptors previously shown to promote or inhibit OC differentiation. ST-2 cells were cocultured with human PBMC for up to 21 days in the presence of 1alpha,25(OH)(2) vitamin D(3), dexamethasone, and recombinant human macrophage-colony stimulating factor (M-CSF). OC formation was monitored by the appearance of cells that were positive for tartrate-resistant acid phosphatase (TRAP) and able to form resorption lacunae on slices of dentine. We found that the ST-2 cells in these cultures express messenger RNA (mRNA) encoding a repertoire of many of the reported osteoclastogenic factors (interleukins [IL]-1/IL-1R1, IL-11, IL-6/IL-6R, and IL-17 transforming growth factor [TGF]-beta), as well as the recently described OC differentiation factor (ODF/TRANCE/RANKL). The stromal cells also expressed mRNA encoding two molecules shown to be inhibitory to osteoclastogenesis, osteoprotegerin (OPG) and IL-18. OPG, IL-1, IL-1R1, IL-6, IL-6R, IL-11R, IL-17, IL-18, IL-18R, TGF-beta, and M-CSF were expressed by both the stromal cells and the PBMC. Expression of mRNA encoding RANK, IL-1R2, and c-fms, was specific for the PBMC. In addition, PBMC were found to express sIL-6R, granulocyte macrophage (GM)-CSF, GM-CSFRalpha, and tumor necrosis factor (TNF)-alpha. Whereas this indicated that human OC formation occurs in a complex environment of many positive and negative influences, we identified three apparent features of the cytokine environment that may be a characteristic of normal osteoclast formation. First, the ratio of mouse ODF:OPG mRNA was found to increase during the cocultures, consistent with a key role for ODF in the promotion by stromal cells of OC formation. Second, we found that mRNA encoding IL-1 and IL-17, as well as IL-6 and sIL-6R, were coordinately expressed by the PBMC. Third, analysis of the culture medium showed that the PBMC secreted IL-1, IL-6, and TNF-alpha protein only in coculture with ST-2 cells during the first few days of osteoclast development. We conclude that human OC formation occurs in a complex environment of many positive and negative influences; however, these are likely to be strictly regulated by a coordinated cytokine response of both stromal and hematopoietic cells.
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PMID:Coordinated cytokine expression by stromal and hematopoietic cells during human osteoclast formation. 1083 38


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