Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Eicosanoid release during multilineage hematopoiesis was assessed using freshly isolated mouse bone marrow cells cultured in the presence of interleukin-3 (IL-3) (10% WEHI-3 culture-conditioned medium). Cells that could release prostaglandin E2 (PGE2) when stimulated with calcium ionophore A23187, but not with phorbol ester (PMA), appeared within 4 days. The cells harvested on day 10 released 42 ng of PGE2/10(6) cells/mL after A23187 stimulation. Leukotriene B4 (LTB4) (4 ng/mL) was also detected after A23187 stimulation, but there was no detectable LTC4 (less than 0.5 ng/mL). Nonadherent bone marrow cells were isolated from 28-day cultures and cloned. All clones were strongly IL-3-dependent. Although other growth factors such as granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage CSF (GM-CSF), and CSF-1 failed to promote survival or support proliferation of the cells, three clones (11-1-A6, 3-2-D5, and 11-1-A1) showed significant increases in 3H-thymidine incorporation, respectively, after PMA treatment for 24 hours. Surviving cells displayed dominantly myeloid type morphology and phenotypic characteristics. The data suggest that IL-3 is important in the formation of PGE2-producing cells. In contrast to many macrophages (MO), neither the IL-3-dependent cell lines nor the IL-3-cultured bone marrow cells released significant amounts of PGE2 when stimulated with PMA or IL-3, although PMA and IL-3 both induced translocation of protein kinase C (PKC) to the membrane fraction. The lack of production of PGE2 and other eicosanoids by the PMA- and IL-3-stimulated cell lines was confirmed by measuring the release of 3H-arachidonic acid. The data suggest that in IL-3-dependent bone marrow cell lines the activation of eicosanoid metabolism requires elevated cellular Ca2+; PKC activation alone does not appear to be a sufficient stimulus.
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PMID:Calcium ionophore but not phorbol ester promotes eicosanoids release by proliferating interleukin-3-dependent bone marrow cells. 216 25

We have prepared a population of bone marrow cells that is highly enriched in neutrophil/macrophage progenitor cells (GM-CFC). Four distinct haemopoietic growth factors can stimulate the formation of mature cells from this population, although the proportions of neutrophils and/or macrophages produced varied depending on the growth factor employed: interleukin 3 (IL-3) and granulocyte/macrophage colony-stimulating factor (GM-CSF) stimulated the formation of colonies containing both neutrophils and macrophages; macrophage colony-stimulating factor (M-CSF) produced predominantly macrophage colonies; and granulocyte colony-stimulating factor (G-CSF) promoted neutrophil colony formation. Combinations of these four growth factors did not lead to any additive or synergistic effect on the number of colonies produced in clonal soft agar assays, indicating the presence of a common set of cells responsive to all four haemopoietic growth factors. These enriched progenitor cells therefore represent an ideal population to study myeloid growth-factor-stimulated survival, proliferation and development. Using this population we have examined the molecular signalling mechanisms associated with progenitor cell proliferation. We have shown that modulation of cyclic AMP levels has no apparent role in GM-CFC proliferation, whereas phorbol esters and/or Ca2+ ionophore can stimulate DNA synthesis, indicating a possible role for protein kinase C activation and increased cytosolic Ca2+ levels in the proliferation of these cells. The lack of ability of all four myeloid growth factors to mobilize intracellular Ca2+ infers that these effects are not achieved via inositol lipid hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Identification of a common signal associated with cellular proliferation stimulated by four haemopoietic growth factors in a highly enriched population of granulocyte/macrophage colony-forming cells. 255 52

We studied the ability of the recombinant human-active hemopoietic growth factors granulocyte-macrophage colony-stimulating factor (GM-CSFrh) and granulocyte colony-stimulating factor (G-CSFrh) to activate receptor-mediated transduction pathways which have been implicated in the stimulation of cytotoxic functions in granulocytes. With the use of a panel of fluorescent probes, we found that these two growth factors exerted no detectable immediate effect on the resting transmembrane electrical potential, the intracellular concentration of free calcium ions, or the cytosolic pH of isolated, mature granulocytes. However, when granulocytes were "primed" by preincubation for 90 min with GM-CSFrh or G-CSFrh, the rate of membrane depolarization induced by 10(-7) M N-formyl-methionyl-leucyl-phenylalanine, but not the rate of rise in free calcium ions, was greatly accelerated. In examining potential mechanisms to account for the priming effect of these growth factors, we found that although they did not induce translocation of protein kinase C or stimulate significant degranulation, they each directly caused prompt release of arachidonic acid from plasma membrane phospholipids. Our data indicate that although GM-CSFrh and G-CSFrh do not activate the transduction signals that have most clearly been implicated in receptor-mediated activation of cytotoxic functions in granulocytes--namely, those coupled to membrane depolarization or release of intracellular calcium ions--they appear directly to induce the release of arachidonic acid esterified to membrane phospholipids, an event which may represent the receptor-mediated activation of membrane phospholipases and which may contribute to the "priming" of the cells for enhancement of their functional responsiveness.
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PMID:Effects of recombinant human granulocyte and macrophage colony-stimulating factors on signal transduction pathways in human granulocytes. 311 8

The haematopoietic growth factors multi-colony-stimulating factor, granulocyte/macrophage colony-stimulating factor, granulocyte colony-stimulating factor and interleukin 2 specifically control the production and proliferation of distinct leucocyte series. Each growth factor acts on a unique surface receptor associated with an appropriate signal-transduction apparatus. In this report we identify a 68 kDa substrate which is phosphorylated after stimulation of different cell types with multi-colony-stimulating factor, granulocyte colony-stimulating factor and interleukin 2. The 68 kDa substrate is also phosphorylated in each cell line stimulated with synthetic diacylglycerol, a direct activator of protein kinase C. Interestingly, granulocyte/macrophage colony-stimulating factor does not induce phosphorylation of the 68 kDa molecule. The 68 kDa molecule that is phosphorylated after stimulation with each ligand yielded similar peptide maps after chymotryptic digestion; furthermore, the substrate was always phosphorylated on threonine residues. Phosphorylation of the same residues in the 68 kDa substrate suggests that activation of protein kinase C is one common signal-transduction event associated with the action of multi-colony-stimulating factor, granulocyte colony-stimulating factor and interleukin 2.
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PMID:Identification of a signal-transduction pathway shared by haematopoietic growth factors with diverse biological specificity. 350 46

N-Acetylglucosaminyl-1,6-anhydro-N-acetylmuramyl-L-alanyl-D-isoglutam yl-m- diaminopimelyl-D-alanine [G (Anh)MTetra], a naturally occurring breakdown product of peptidoglycan from bacterial cell walls, was studied for its ability to induce granulocyte colony-stimulating factor (G-CSF) mRNA and protein expression in human adherent monocytes. Resting monocytes did not express G-CSF mRNA or secrete G-CSF protein. In contrast, monocytes exposed to G(Anh)MTetra showed a dose-dependent increase in G-CSF mRNA accumulation, which correlates with the secretion of G-CSF protein. Maximal levels of G-CSF mRNA were reached within 2 h of activation. Expression of G-CSF was mediated by an increase in the stability of G-CSF transcripts rather than by an increase in the transcription rate of the G-CSF gene. Experiments with the protein synthesis inhibitor cycloheximide revealed that G(Anh)MTetra-induced G-CSF mRNA expression was independent of new protein synthesis. Furthermore, it was shown that the effect of G(Anh)MTetra was regulated by a protein kinase C-dependent pathway, whereas protein kinase A and tyrosine kinases were not involved. Finally, it was shown that G(Anh)MTetra also induced G-CSF mRNA expression in human endothelial cells. The data indicate that, besides lipopolysaccharide, other naturally occurring bacterial cell wall components are able to induce G-CSF expression in different hematopoietic cells.
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PMID:G(AnH)MTetra, a naturally occurring 1,6-anhydro muramyl dipeptide, induces granulocyte colony-stimulating factor expression in human monocytes: a molecular analysis. 751 14

Chronic exposure of humans to benzene (BZ) causes acute myelogenous leukemia. These studies determined whether BZ, or its reactive metabolite, hydroquinone (HQ), affect differentiation of myeloblasts. BZ or HQ administered to C57BL/6J mice specifically induced terminal granulocytic differentiation of myeloblasts. The ability of the compounds to induce differentiation of the myeloblast was tested directly using the murine interleukin 3 (IL-3)-dependent myeloblastic cell line, 32D.3 (G) and the human HL-60 promyelocytic leukemic cell line. Treatment of HL-60 myeloblasts with BZ activated protein kinase C and upregulated the 5-lipoxygenase (LPO) pathway for the production of leukotriene D4 (LTD4), an essential effector of granulocytic differentiation. Differentiation was prevented by sphinganine, a kinase C inhibitor, as well as by LPO inhibitors and LTD4 receptor antagonists. BZ and HQ also induced differentiation in 32D.3 (G) myeloblasts. Both compounds interact with cellular signaling pathways activated by granulocyte colony-stimulating factor (G-CSF) and thus replace the requirement for G-CSF. IL-3 induces a growth response, whereas G-CSF provides both growth and differentiation signals. BZ does not induce growth in the absence of IL-3, but provides a differentiation signal. Both HQ and LTD4 induce differentiation and synergize with IL-3 for growth, however, neither support growth in the absence of IL-3. BZ-induced 32D cells showed a gradual progression of progenitor differentiation to granulocytes similar to that seen with G-CSF or LTD4. HQ blocks differentiation at the myelocyte stage; only a small percentage of progenitors proceed to granulocytes. BZ, like G-CSF, upregulates LTD4 production, whereas HQ obviates the requirement for LTD4 by activating the LTD4 receptor.
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PMID:Benzene and its metabolite, hydroquinone, induce granulocytic differentiation in myeloblasts by interacting with cellular signaling pathways activated by granulocyte colony-stimulating factor. 754 15

Severe congenital neutropenia (SCN) can be corrected in vivo by treatment with pharmacological dosages of recombinant human granulocyte colony-stimulating factor (rhG-CSF). In order to analyze the decreased chemotaxis of neutrophils from SCN patients receiving rhG-CSF, neutrophil functions essential for chemotaxis were investigated. The mobilization of cytosolic calcium ([Ca2+]i) and the functional state of cytoskeletal proteins in neutrophils from SCN patients were compared with either neutrophils from healthy donors (or, in selected experiments, from patients with cyclic neutropenia) and neutrophils from patients with chemotherapy-induced neutropenia also receiving rhG-CSF. Using flow cytometric analysis, two neutrophil subpopulations were detected in SCN patients in response to N-formylmethionine leucyl-phenylalanine (FMLP) (10(-9) M to 10(-7) M), one of which was unable to respond to this stimulus with an increase in [Ca2+]i. Whereas a homogeneous increase in [Ca2+]i in normal neutrophils occurred at 10(-9) M FMLP, neutrophils from SCN patients required 10(-6) M FMLP to respond homogeneously with an increase in [Ca2+]i. In contrast, G-CSF induced neutrophils from patients with cyclic neutropenia and from patients with chemotherapy-induced neutropenia showed a normal increase in [Ca2+]i after stimulation. The [Ca2+]i-dependent superoxide anion (O2-) generation in response to FMLP was also significantly diminished in neutrophils from SCN patients compared to normal neutrophils. However, O2- generation elicited by phorbolester (PMA), which directly activates protein kinase C (PKC), was not affected in SCN neutrophils. The total immunoreactive actin content and basal F-actin content in neutrophils from SCN patients were elevated as compared to normal neutrophils and neutrophils from patients with chemotherapy-induced neutropenia. The increase in F-actin content following FMLP activation was much lower in neutrophils from SCN patients as compared with normal neutrophils. These data suggest a defect in the signal transduction pathway in neutrophils from SCN patients between FMLP ligand-receptor interaction and Ca2+ mobilization, whereas upstream of PKC, triggered events seem to be unaffected. Therefore, [Ca2+]i-dependent neutrophil function in response to FMLP, such as actin disassembly, chemotaxis and O2- generation are diminished in SCN neutrophils. The pathomechanism responsible for the defective [Ca2+]i increase might be an initial step in understanding the underlying pathophysiology of SCN.
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PMID:Abnormal regulation in the signal transduction in neutrophils from patients with severe congenital neutropenia: relation of impaired mobilization of cytosolic free calcium to altered chemotaxis, superoxide anion generation and F-actin content. 767 87

To study the receptors involved in the interaction between extracellular matrix proteins and hematopoietic progenitor cells, we analyzed the expression of beta 1 integrins on CD34+ bone marrow cells by means of immunoflowcytometry. Alpha 4 beta 1 and alpha 5 beta 1 were expressed, whereas alpha 1 beta 1, alpha 2 beta 1, alpha 3 beta 1, alpha 6 beta 1, and alpha v beta 1 were virtually absent. Furthermore, we assessed the alpha 4 and alpha 5 expression on committed myeloid progenitor cells. These colony-forming cells were detected in the alpha 4 dull fraction and the alpha 5 dull fraction. During myeloid differentiation, both in vivo and in vitro, a differential expression of alpha 4 beta 1 and alpha 5 beta 1 was observed. alpha 5 beta 1 was found to be lost at the myelocytic-metamyelocytic stage, before the loss of alpha 4 beta 1, at the band stage. Functional studies showed no binding of erythroid progenitor-depleted, CD34+ bone marrow cells to fibronectin. However, protein kinase C activation strongly induced fibronectin binding (68% of the cells). Inhibition experiments with specific antibodies and peptides showed the binding to be mediated by both alpha 4 beta 1 and alpha 5 beta 1. Also, colony-forming cells of granulocytes and macrophages were demonstrated to adhere to fibronectin in an activation-dependent way. During granulocyte colony-stimulating factor-induced in vitro maturation, the activation-dependent fibronectin binding capacity is gradually lost. We conclude that: (1) CD34+ bone marrow cells express alpha 4 beta 1 and alpha 5 beta 1; (2) the expression of alpha 4 beta 1 and alpha 5 beta 1 is differentially expressed during myeloid differentiation; and (3) binding of CD34+ bone marrow cells to fibronectin is activation dependent.
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PMID:Alpha 4 beta 1 and alpha 5 beta 1 are differentially expressed during myelopoiesis and mediate the adherence of human CD34+ cells to fibronectin in an activation-dependent way. 767 11

The effects of direct activators of protein kinase C (PKC) (the phorbol ester tetradecanoyl phorbol myristic acid [TPA] or bryostatin) on the ability of a highly enriched population of granulocyte-macrophage colony-forming cells (GM-CFC) to proliferate and develop in soft agar was assessed. In the absence of colony stimulating factors, the PKC activators did not stimulate colony formation. However, in the presence of optimal concentrations of granulocyte colony-stimulating factor (G-CSF) or interleukin-6 (IL-6), TPA or bryostatin markedly elevated the number of colonies formed from the GM-CFC. In the absence of TPA, IL-6, and G-CSF, respectively, both stimulated the formation of about 3% of the colonies observed when IL-3 was present. When TPA plus G-CSF or IL-6 were added together, this figure increased to 48% and 54%, respectively. In both instances, the types of mature cells formed was altered from colonies of mature neutrophilic cells to a mixture consisting predominantly of macrophages with some neutrophils. Similar results were observed when bryostatin replaced TPA in these assays. When single cell colony-forming assays were performed, the same results were obtained. The presence of G-CSF, or IL-6, and the activator of PKC used (TPA or bryostatin) was required throughout the colony-forming assay for an optimal synergistic effect to be observed. These data indicate that agents that activate PKC can promote the proliferation and development of GM-CFC via a synergistic interaction with G-CSF or IL-6. Furthermore, there is an apparent role for PKC in development and possibly lineage commitment of GM-CFC.
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PMID:Protein kinase C activators can interact synergistically with granulocyte colony-stimulating factor or interleukin-6 to stimulate colony formation from enriched granulocyte-macrophage colony-forming cells. 767 6

We have examined the effect of the macrocyclic lactone protein kinase C (PK-C) activator bryostatin 1 on the proliferative capacity and lineage commitment of CD34+ human bone marrow cells exposed to the granulocyte-macrophage colony-stimulating factor/interleukin-3 (GM-CSF/IL-3) fusion protein pIXY 321. pIXY 321 administered at a dose of 10 ng/mL was as effective as the combination of plateau concentrations of recombinant (r) IL-3 and rGM-CSF (e.g., 50 ng/mL) in stimulating the growth of day-14 committed myeloid progenitors (colony-forming units granulocyte/macrophage [CFU-GM]). In the large majority of samples tested, coadministration of 0.5 to 100 nM bryostatin 1 with either pIXY 321 or the combination of rIL-3 plus rGM-CSF led to modest but significant increases (e.g., 30 to 75%) in the number of CFU-GM, compared to administration of growth factors alone. The degree of bryostatin 1-induced potentiation, however, was considerably less than that previously observed in the case of cells exposed to either rIL-3 or rGM-CSF, where increases of 100 to 150% were regularly noted. While at least 50% of day-14 CFU-GM stimulated by either pIXY 321 or the combination of rIL-3 plus rGM-CSF were of the pure or mixed eosinophilic variety, coadministration of bryostatin 1 resulted in a dramatic inhibition of eosinophilic colonies and a corresponding increase in pure and mixed neutrophil and macrophage colonies. Although coadministration of recombinant granulocyte colony-stimulating factor (rG-CSF) or recombinant colony-stimulating factor-1 (rCSF-1) mimicked the capacity of bryostatin 1 to increase the total number of pIXY 321-induced day-14 CFU-GM, these growth factors, unlike bryostatin 1, were not capable of inhibiting eosinophilic colony formation. Furthermore, whereas addition of neutralizing antibodies to G-CSF or CSF-1 blocked the capacity of these growth factors to potentiate colony formation in the presence of pIXY 321, it did not abrogate the effect of bryostatin 1 on progenitor cell growth or lineage commitment. Finally, in contrast to its effects on committed myeloid progenitors, bryostatin 1 did not increase the growth of erythroid (burst-forming units-erythroid [BFU-E]) and multipotent (multipotent colony-forming units [CFU-GEMM]) progenitors stimulated by pIXY 321, but instead inhibited colony formation at higher concentrations (e.g., 10 to 100 nM).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Modulation of the activity of a human granulocyte-macrophage colony-stimulating factor/interleukin-3 fusion protein (pIXY 321) by the macrocyclic lactone protein kinase C activator bryostatin 1. 768 3


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