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:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously shown that the growth factor FLT3 ligand (FL) is mitogenic for human primary and continuously cultured myeloid leukemia cells. Despite widespread expression of the receptor FLT3 among the leukemia cell lines from certain cell lineages, only two growth factor-dependent myeloid leukemia cell lines showed a significant proliferative response to FL. In the present study, we examined the proliferative effects of FL on a comprehensive set of growth factor-dependent leukemia cell lines. A significant enhancement of cell growth by FL was seen in 10/12 myelomonocytic cell lines, while all cell lines with predominantly megakaryocytic and/or erythroid characteristics did not respond positively, despite the expression of the receptor. The cytokines interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF) and stem cell factor (SCF) could independently enhance the FL-stimulated proliferation in a synergistic fashion. Transforming growth factor-(beta)1 (TGF-(beta)1), in a dose-dependent fashion, partially inhibited the FL-promoted proliferation, but basic fibroblast growth factor (bFGF), on its own augmenting the response to FL, significantly abrogated the inhibitory effects of TGF-(beta)1. TGF-(beta)1 down-regulated mRNA and protein expression of the FLT3 receptor. Taken together these data suggest that the effects of FL on the growth of normal and malignant hematopoietic cells can be positively and negatively modulated by other cytokines.
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PMID:Effects of FLT3 ligand on human leukemia cells. II. Agonistic and antagonistic effects of other cytokines. 863 36

HTK is a receptor tyrosine kinase that belongs to the Eph subfamily. An extensive screening using BIAcore system revealed that a colon cancer cell line, C-1, expressed the ligand for HTK. From the conditioned medium of C-1 cells, a soluble form of ligand was purified by receptor affinity chromatography, and the isolation of full-length cDNA revealed that this ligand is identical to the human HTK ligand (HTKL) previously reported. HTK receptor tyrosine phosphorylation was induced by membrane-bound or clustered soluble HTKL but not by unclustered soluble HTKL, indicating that HTKL requires cell-to-cell interaction for receptor activation. Binding analysis demonstrated that HTKL binds to HTK with a much higher affinity (Kd: 1.23 nM) than the other transmembrane-type ligand for Eph family, LERK-2/ELKL (Kd: 135 nM). The expression of HTK in cord blood cells was upregulated after the culture in the presence of stem cell factor. Clustered soluble HTKL stimulated the proliferation of sorted HTK+ cord blood cells and a hematopoietic cell line, UT-7/EPO from which HTK was isolated. These findings suggest the involvement of HTK-HTKL system in the proliferation of HTK+ hematopoietic progenitor cells in the hematopoietic environment.
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PMID:Characterization of a ligand for receptor protein-tyrosine kinase HTK expressed in immature hematopoietic cells. 876 3

Bone remodeling requires cooperation between osteoclasts and other specialized or accessory bone cell populations by mechanisms that have not been completely elucidated. Here we describe the expression and functional role of the proto-oncogene c-kit and of its specific ligand stem cell factor (SCF) on human osteoclasts, osteoblasts, and stromal cells derived from different sources. Our results indicate that primary osteoclasts in imprints of metaphyseal bone and giant cell tumors (GCTs) of bone, as well as a bone marrow-derived preosteoclast cell line of human origin (FLG 29.1), expressed immunodetectable c-kit protein. In contrast, tissue osteoclasts did not react with anti-SCF antibodies, and barely detectable levels of SCF mRNA and protein were found in FLG 29.1 cells. Conversely, a strong expression of membrane bound-SCF was found in primary cultured bone marrow stromal cells, in a stromal cell line (C433) derived from the mononuclear component of GCT of bone, and in a human cell line with osteoblast features (Saos-2). FLG 29.1 preosteoclast cells displayed about 29,000 binding sites/cell of a single class of high affinity c-kit receptors (Kd 6.12 x 10(-10) mol/L) with a molecular weight of about 140 kDa, along with a structurally normal c-kit mRNA. Proliferation of FLG 29.1 preosteoclast cells was stimulated by exogenous SCF, indicating that c-kit was capable of transducing growth signals. Finally, in vitro adhesion of FLG 29.1 cells to primary bone marrow stromal cells, GCT-derived stromal cells (C433), and Saos-2 osteoblast cells was significantly inhibited by an excess of soluble SCF or by monoclonal antibodies recognizing SCF binding sites on the c-kit receptor. These results indicate that c-kit is constitutively expressed on human osteoclasts and that it may be directly implicated in cell contact-dependent interaction of osteoclasts with other specialized or accessory cell populations of the bone microenvironment. Our observations suggest a role for SCF in human diseases characterized by abnormal bone resorption and remodeling.
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PMID:Human osteoclasts and preosteoclast cells (FLG 29.1) express functional c-kit receptors and interact with osteoblast and stromal cells via membrane-bound stem cell factor. 878 Aug 89

We have evaluated an easy and fast immunomagnetic method for positive selection of cells expressing the CD34 antigen from BM, peripheral blood (PB) and apheresis products (AP) of CML patients and healthy adults (HA) in order to further characterize them by immunophenotypic analysis. From an initial frequency of CD34+ cells in the original sample of 1.8 +/- 1.7%, CD34+ cells were rapidly and efficiently enriched up to 91.5 +/- 6.4% by high-gradient magnetic cell sorting (MACS) (yield 53 +/- 21%). A five-dimensional flow cytometric analysis of the immunomagnetic isolated CD34+ cells demonstrated little overlap between CD34+HLA-DRlo and CD34+CD38lo subpopulations in both BM-HA and in BM-CML. Only 16 and 6% of the CD34+HLA-DRlo and CD34+CD38lo cells respectively, showed lack of expression of both Ag (CD34+HLA-DRloCD38lo) in BM-CML samples. Between 60 and 70% of the CD34+ cells expressed the stem cell factor (SCF) receptor (c-KIT, CD117) and there were no differences between BM-HA and BM-CML patients. Moreover, more than 60% of the CD34+HLA-DRlo cells, co-expressed c-KIT. MACS-enriched BM-CD34+ cells showed normal hematopoietic colony formation in vitro in all the sources analyzed with a higher colony-forming efficiency than the unfractionated sample (MNC).
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PMID:Isolation of CD34+ hematopoietic progenitor cells in chronic myeloid leukemia by magnetic activated cell sorting (MACS). 887 25

The c-KIT proto-oncogene encodes for a transmembrane receptor and is associated with maturation of several cell types, including germ cells. The ligand of the receptor has been identified as stem cell factor (SCF). Loss or alteration of the expression of either of these factors leads to anemia, albinism, and/or sterility in mice. We examined the expression of c-KIT and SCF by immunohistochemistry in specimens from normal and infertile human testis. All specimens were obtained in the evaluation of male subfertility. We were able to demonstrate staining for c-KIT in Leydig cells in all specimens. Normal testis stained for c-KIT in the cytoplasm of early spermatogenic cells, as well as the acrosomal granules of the round spermatids and the acrosome of testicular spermatozoa. However, staining in testis demonstrating maturation arrest failed to demonstrate acrosomal staining, and Sertoli-only specimens demonstrated staining for c-KIT in Leydig cells only. The results for SCF demonstrated an overall uniform staining of Leydig cells in all specimens. The intensity of staining of Sertoli cells increased from normal to maturation arrest to Sertoli-only specimens. Germ cell staining was consistently negative. We hypothesize that these staining patterns for SCF are due to either lack of staining of the receptor-ligand complex or overexpression of the kit ligand in tissue that does not express the kit receptor. It appears that the c-kit receptor is expressed in the acrosome of developing germ cells, as well as in Leydig cells and early spermatogenic cells, suggesting a role in the acrosome reaction, as well as germ cell maturation and differentiation.
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PMID:Expression of c-KIT and its ligand, stem cell factor, in normal and subfertile human testicular tissue. 888 3

FLT3 ligand is a hematopoietic growth factor that plays a key role in growth of primitive hematopoietic cells. FLT3 receptor mRNA is found in early hematopoietic progenitors and in human myeloid leukemia blasts. Much less is known about the surface expression of FLT3 receptor on human hematopoietic cells. Using human 125I-FLT3 ligand, we have identified and characterized surface FLT3 receptors on normal and malignant human hematopoietic cells and cell lines. Our results showed that surface display of FLT3 receptor was greatest in fresh myeloid leukemia blast cells and myeloid leukemia cell lines. Erythroleukemic and megakaryocytic leukemia cell lines (n = 5) bound little to no 125I-FLT3 ligand. Scatchard analysis of 125I-FLT3 ligand binding data shows that three myeloid leukemia cell lines, ML-1, AML-193, and HL-60, as well as normal human marrow mononuclear cells, exhibit high affinity FLT3 receptors. Crosslinking of 125I-FLT3 ligand to FLT3 receptors on the surface of ML-1 myeloid leukemia cells indicates that the FLT3 ligand. The rates of FLT3 ligand internalization and degradation were determined by binding 125I-FLT3 ligand to ML-1 cells and acid stripping to distinguish surface bound from internalized ligand. Internalized 125I-FLT3 ligand was detected within 5 minutes after binding to ML-1 cells. In addition, we evaluated the effect of FLT3 ligand on megakaryocytic colony growth and nuclear endoreduplication, alone or in the presence of thrombopoietin. FLT3 ligand did not promote colony forming unit megakaryocyte (CFU-Meg) colony growth or megakaryocyte nuclear maturation, nor did FLT3 ligand augment the effects of thrombopoietin on these measures of megakaryopoiesis. These data indicate that the FLT3 receptor shares several characteristics with the c-kit receptor including dimerization and rapid internalization. However, the more restricted cellular distribution of the FLT3 receptor may target the effects of FLT3 ligand to primitive hematopoietic cells and to myeloid and lymphoid progenitor cells, in contrast to the pleiotropic effects of the c-kit receptor ligand, stem cell factor.
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PMID:FLT3 receptor expression on the surface of normal and malignant human hematopoietic cells. 889 3

Peripheral blood stem cells (PBSC) are used increasingly for autotransplantation in the treatment of acute leukemia, lymphoma, multiple myeloma, solid tumors such as ovarian and breast carcinoma. They are collected by leukaphereses during rapid hematopoietic recovery, following cytotoxic chemotherapy with or without administration of hematopoietic growth factors. We studied the clonogenic and cytokine-mediated expansion potential of CD34+ cells from mobilized PBSC. Low density mononuclear cells were processed using the CEPRATE LC CD34 KIT (CellPro). CD34+ purified cells, were cultured in suspension with 6 combined hematopoietic growth factors (IL1beta, IL3, IL6 at 100 U/ml and G-CSF, GM-CSF and stem cell factor at 10 ng/ml of each) for up to four weeks. Every week, cells were counted and CFU-GM assay was performed in a methylcellulose based medium. We have analysed the percentage of cells bearing CD34, CD33, CD38, HLA-DR, CD45RA, CD45RO antigens. Our results showed, that CD34+ cells were obtained with a purity of 92 +/- 2.3% and a yield of 71 +/- 10.7%. The majority co-expressed CD33 (57.76 +/- 34.16%) and CD38 (62.2 +/- 34%) antigens. These culture conditions, are necessary to obtain a fold increase of nucleated cells (377 fold at week 4), of CFU-GM progenitors (41.2 fold at week 3) and of CD34+ cell absolute number (10 fold at week 1) with an important differentiation of progenitors in particular myeloid progenitors.
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PMID:Peripheral blood CD34+ cells: method of purification and ex vivo expansion. 890 32

Expression of the tyrosine-kinase receptor encoded by the c-KIT proto-oncogene progressively decreases during local tumor growth and invasion of human melanomas. To provide direct evidence that c-KIT plays a role in metastasis of human melanoma, we transfected the c-KIT gene into the c-KIT negative highly metastatic human melanoma cell line A375SM and subsequently analysed its tumorigenic and metastatic potential. A375SM parental cells, A375SM-NOT (neo, control), and A375SM-KIT-positive cells were injected s.c. and i.v. into nude mice. A375SM-KIT cells produced significantly slower growing s.c. tumors and fewer lung metastases than control cells. Exposure of c-KIT-positive melanoma cells in vitro and in vivo to stem cell factor (SCF), the ligand for c-KIT, triggered apoptosis of these cells but not of c-KIT-negative melanoma cells or normal melanocytes. Since SCF is produced by keratinocytes and other dermal cells in the skin, these results suggest that the loss of c-KIT receptor expression may allow malignant melanoma cells to escape SCF/c-KIT-mediated apoptosis, hence contributing to tumor growth and eventually metastasis. The antitumor and antimetastatic properties of SCF may be useful in treating human melanomas in early stages.
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PMID:Enforced c-KIT expression renders highly metastatic human melanoma cells susceptible to stem cell factor-induced apoptosis and inhibits their tumorigenic and metastatic potential. 895 75

Stromal support is required during retroviral-mediated transduction of human bone marrow-derived CD34+ cells to maintain the clonogenicity of the primitive progenitors. We hypothesized that the cytokine FLT3 ligand (FL) might be able to replace the maintenance role provided by the stroma. CD34+ progenitors from human bone marrow were transduced by the retroviral vector LN with the cytokines interleukin-3 (IL-3), IL-6, and stem cell factor (SCF) present in all cultures. Transductions were performed with or without stromal support and with or without the inclusion of 100 U/mL FL. No significant increase in gene transfer into colony-forming cells was obtained by the addition of FL to the cultures. Transduction and survival of more primitive human hematopoietic cells was determined by growth in immune-deficient mice for 7 to 8 months. Human myeloid cells, T lymphocytes, and colony-forming progenitors were recovered from the marrow of mice that had received human cells transduced on stroma or in suspension culture with IL-3, IL-6, SCF, and FL, but not with IL-3, IL-6, and SCF alone. LN provirus was detected by polymerase chain reaction in the marrow recovered from 9 of 10 mice transplanted with human CD34+ cells transduced with stromal support, 5 of 11 mice that received human cells transduced in suspension culture with FL, but none of the 10 mice that received human cells transduced in suspension culture without FL We conclude that FLT3 ligand, in conjunction with IL-3, IL-6, and SCF, preserves the generative capacity of primitive human hematopoietic cells during in vitro transductions in suspension culture.
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PMID:FLT3 ligand preserves the ability of human CD34+ progenitors to sustain long-term hematopoiesis in immune-deficient mice after ex vivo retroviral-mediated transduction. 900 46

Production of mature erythrocytes requires multiple growth factors, but we do not know how their actions are coordinated. Here we show that erythroid progenitors from erythropoietin receptor (Epo-R)-/- fetal livers, infected in vitro with a retrovirus expressing the wild-type Epo-R, require addition of both Epo and stem cell factor (SCF) to form colony-forming unit erythroid (CFU-E) colonies. Thus, a functional interaction between KIT and the Epo-R, similar to what we reported in cultured cells, is essential for the function of CFU-E progenitors. In contrast, CFU-E colony formation in vitro by normal fetal liver progenitors requires only Epo; the essential interaction between activated KIT and the Epo-R must have occurred in vivo before or at the CFU-E progenitor stage. Using truncated dominant-negative mutant Epo-Rs, we show that KIT does not activate the Epo-R by inducing its dimerization, but presumably does so by phosphorylating tyrosine residue(s) in its cytosolic domain. By expressing mutant Epo-Rs containing only one of eight cytosolic tyrosines, we show that either tyrosine residue Y464 or Y479 suffices for Epo-dependent cell proliferation. However, only Epo-R F7Y479 is capable of supporting erythroid colony formation when expressed in (Epo-R)-/- fetal liver cells, indicating that Y464 either cannot send a differentiation signal or fails to respond to SCF/KIT activation. This work employs a novel experimental system to study the function of growth factors and their receptors in normal hematopoiesis.
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PMID:Functional interaction of erythropoietin and stem cell factor receptors is essential for erythroid colony formation. 905 Aug 60


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