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Biomedical subjects

D E Williams

Publications and source records attributed to D E Williams.

At least 163 records · Page 9Linked to original sources
PubMed1992

Stromal support enhances cell-free retroviral vector transduction of human bone marrow long-term culture-initiating cells.

Gene transfer into hematopoietic stem cells by cell-free virions is a goal for gene therapy of hematolymphoid disorders. Because the hematopoietic microenvironment provided by the stroma is required for stem cell maintenance both in vivo and in vitro, we reasoned that cell-free transduction of bone marrow cells (BMC) may be aided by stromal support. We used two high-titer replication-defective retroviral vectors to differentially mark progenitor cells. The transducing vector was shown to be a specific DNA fragment by polymerase chain reaction of colony-forming cells derived from progenitors maintained in long-term culture (LTC). BMC were infected separately by cell-free virions with or without pre-established, irradiated, allogeneic stromal layers, and in the presence or absence of exogenous growth factors (GF). The GF assessed were interleukin-3 (IL-3) and IL-6 in combination, leukemia inhibitory factor (LIF), mast cell growth factor (MGF), and LIF and MGF in combination. In addition, we developed a competitive LTC system to directly assess the effect of infection conditions on the transduction of clonogenic progenitors as reflected by the presence of a predominate provirus after maintenance in the same microenvironment. The results show gene transfer into human LTC-initiating cells by cell-free retroviral vector and a beneficial effect of stromal support allowing a transduction efficiency of 64.6% in contrast to 15.8% without a supporting stromal layer. A high transduction rate was achieved independent of stimulation with exogenous GF. We propose that autologous marrow stromal support during the transduction period may have application in clinical gene therapy protocols.

Base Sequence↗
PubMed1992

Cytokine stimulation of multilineage hematopoiesis from immature human cells engrafted in SCID mice.

Severe combined immunodeficient (SCID) mice transplanted with human bone marrow were treated with human mast cell growth factor, a fusion of interleukin-3 and granulocyte-macrophage colony-stimulating factor (PIXY321), or both, starting immediately or 1 month later. Immature human cells repopulated the mouse bone marrow with differentiated human cells of multiple myeloid and lymphoid lineages; inclusion of erythropoietin resulted in human red cells in the peripheral blood. The bone marrow of growth factor-treated mice contained both multipotential and committed myeloid and erythroid progenitors, whereas mice not given growth factors had few human cells and only granulocyte-macrophage progenitors. Thus, this system allows the detection of immature human cells, identification of the growth factors that regulate them, and the establishment of animal models of human hematopoietic diseases.

Animals↗
PubMed1992

Recombinant murine steel factor stimulates in vitro production of granulocyte-macrophage progenitor cells.

The ability of murine Steel factor to promote the in vitro production of granulocyte-macrophage progenitor cells (CFU-GM) was examined in short-term liquid cultures. Bone marrow from C57BL/6J or Sl/Sld mice was placed in culture for seven days with either Steel factor alone or in the presence of IL-3. CFU-GM responsive to GM-CSF, IL-3, and CSF-1 were measured in the input population and again after 3 or 7 days in culture. Steel factor alone increased the number of all CFU-GM types as early as 3 days after culture initiation, with further increases at day 7. This effect was potentiated by the addition of IL-3. Production of CFU-GM by C57BL/6J or Sl/Sld marrow was comparable except for enhanced production of CSF-1 responsive progenitors by Sl/Sld marrow. A recombinant Sld protein was also shown to be equivalent to the wild-type protein in its capacity to promote CFU-GM production from normal bone marrow.

Animals↗
PubMed1992

Steel factor is required for maintenance, but not differentiation, of melanocyte precursors in the neural crest.

Skin melanocytes are derived from neural crest cells that migrate into the dermis during embryogenesis. Two mouse mutants, Steel and White dominant-spotting, which have defects in melanocyte production, have recently been shown to have deletions in the genes that code for a new growth factor, steel factor (SLF), and its receptor, respectively. Here, we have investigated the role that SLF plays in melanogenesis using cultures of mouse neural crest and found that its primary action is the maintenance of melanocyte precursors. It has no effect on the final stage of melanocyte differentiation, the production of melanin, which appears to require an additional factor whose action is mimicked by the phorbol ester TPA (12-O-tetradecanoyl-phorbol-13-acetate).

Animals↗
PubMed1992

The Steel factor.

Steel factor (SLF) is a recently identified growth factor which is the gene product of the murine Steel locus and a ligand for the c-kit tyrosine kinase receptor, the product of the dominant white spotting locus (W). Defects at these genetic loci result in aberrant melanocyte, germ cell, and hematopoietic development. Both the receptor (c-kit) and the ligand (SLF) have been shown to undergo tissue-specific mRNA splicing to produce distinct isoforms which have unique biological functions. As predicted by the phenotype of these mutations, SLF influences the growth and differentiation of melanocytes, primordial germ cells, and a broad spectrum of cell types in the hematopoietic progenitor and stem cell hierarchy. SLF has also been shown to have effects on hematopoietic lineages not predicted by defects seen in the Steel mouse.

Animals↗
PubMed1992

Alterations in lipid peroxidation, antioxidant enzymes, and carcinogen metabolism in liver microsomes of vitamin E-deficient trout and rat.

Feeding rainbow trout for 16 weeks a diet in which the levels of vitamin E were reduced 70-fold resulted in marked depletion (18-fold) of vitamin E levels in liver microsomes from these fish. The susceptibility of hepatic microsomes to lipid peroxidation in vitro and the levels of plasma and liver microsomal lipid hydroperoxides generated in vivo were markedly elevated in vitamin E-depleted trout. No appreciable alterations were observed in the liver microsomal cytochrome P450-dependent mixed-function oxidase system or in the fatty acid composition of trout liver microsomal membranes. Livers from rats fed a vitamin E-deficient diet for 10 weeks also had significantly lower levels of microsomal vitamin E. In addition, total cytochrome P450 levels were depressed (15%) and cytosolic glutathione was enhanced (40%) in livers from rats fed the vitamin E-depleted diet. Covalent binding of [3H]-(+)-benzo[a]pyrene-7,8-dihydrodiol to exogenous DNA in vitro was enhanced with liver microsomes from vitamin E-deficient trout and these fish were much more sensitive to the acute toxicity of this carcinogenic polycyclic aromatic hydrocarbon. These results indicate that trout may be a useful model for studying the significance of peroxidative pathways in carcinogenesis and their manipulation by dietary antioxidants.

Animals↗
PubMed1992

Letter matching: effects of age, Alzheimer's disease, and major depression.

We compared Alzheimer's disease (AD) patients and Major Depressive Disorder (MDD), Aged normal, and Young normal controls on a letter-matching task designed to measure the time needed to access overlearned linguistic information in long-term memory. Name identity (NI) and physical identity (PI) reaction time and the NI-PI difference were compared for ADs, MDDs, and Aged normals and separately for Aged and Young normal groups. AD subjects had slower NI and PI reaction times and a bigger NI-PI difference than Aged normal and MDD subjects, suggesting that speed of access to overlearned letter-name information in long-term memory is slowed for ADs. There were no reliable differences between Aged normal and MDD subjects. Aged normals had slower NI and PI reaction times and a bigger NI-PI difference than Young normals, suggesting that the highly practiced operations needed to access letter-name information slow with age. A discriminant analysis was used to evaluate the usefulness of the "easy to perform" letter-matching task for diagnostic purposes. Ninety percent of normal and MDD subjects but only 68% of AD subjects were classified correctly.

Adult↗
PubMed1992

c-Kit-kinase induces a cascade of protein tyrosine phosphorylation in normal human melanocytes in response to mast cell growth factor and stimulates mitogen-activated protein kinase but is down-regulated in melanomas.

The proto-oncogene c-Kit, a transmembrane receptor tyrosine kinase, is an important regulator of cell growth whose constitutively active oncogenic counterpart, v-kit, induces sarcomas in cats. Mutations in murine c-kit that reduce the receptor tyrosine kinase activity cause deficiencies in the migration and proliferation of melanoblasts, hematopoietic stem cells, and primordial germ cells. We therefore investigated whether c-Kit regulates normal human melanocyte proliferation and plays a role in melanomas. We show that normal human melanocytes respond to mast cell growth factor (MGF), the Kit-ligand that stimulates phosphorylation of tyrosyl residues in c-Kit and induces sequential phosphorylation of tyrosyl residues in several other proteins. One of the phosphorylated intermediates in the signal transduction pathway was identified as an early response kinase (mitogen-activated protein [MAP] kinase). Dephosphorylation of a prominent 180-kDa protein suggests that MGF also activates a phosphotyrosine phosphatase. In contrast, MGF did not induce proliferation, the cascade of protein phosphorylations, or MAP kinase activation in the majority of cells cultured from primary nodular and metastatic melanomas that grow independently of exogenous factors. In the five out of eight human melanoma lines expressing c-kit mRNAs, c-Kit was not constitutively activated. Therefore, although c-Kit-kinase is a potent growth regulator of normal human melanocytes, its activity is not positively associated with malignant transformation.

Cells, Cultured↗
PubMed1992

Dietary hydrogen peroxide enhances hepatocarcinogenesis in trout: correlation with 8-hydroxy-2'-deoxyguanosine levels in liver DNA.

The tumor-enhancing effect of hydrogen peroxide (H2O2) in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-initiated rainbow trout hepatocarcinogenesis was investigated and correlated with the levels of the mutagenic DNA adduct 8-hydroxy-2'-deoxyguanosine (oh8dG). In addition, the protective role of vitamin E was examined in relation to tumor enhancement and oh8dG levels in liver DNA. Trout were fed diets containing two levels of vitamin E (1000 or 20 mg/kg wet wt), each of which were made up to contain three levels of H2O2 (0, 600 or 3000 p.p.m.). Dietary vitamin E levels had no significant effect on tumor incidence or levels of oh8dG in liver DNA. On the other hand, dietary H2O2 enhanced liver tumors in a dose-dependent manner. Liver tumor incidence correlated significantly with the mean level of liver DNA oh8dG content (r = 0.87). We conclude that the H2O2 tumor-enhancing effect coincides with higher levels of oh8dG in the trout liver genome. Thus, rainbow trout may be a useful model for the study of the relationship of oh8dG levels in vivo to enhancement or promotion of carcinogenesis and its modulation by dietary enhancers and inhibitors of oxidative stress.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1992

Biological activities and potential therapeutic uses of steel factor. A new growth factor active on multiple hematopoietic lineages.

Recently, a novel growth factor has been cloned that has growth promoting activities on a wide variety of hematopoietic cell lineages. This factor has been referred to as mast cell growth factor, stem cell factor, or kit ligand, and will be referred to here as steel factor. Steel factor stimulates the growth of cells via its interaction with the c-kit proto-oncogene, which is a tyrosine kinase receptor that is expressed on the surface of a number of different cell types. In addition to its effects on hematopoiesis, this factor also plays a role in the development of melanocytes and germ cells. The discovery of this growth factor provided the final piece of the puzzle to explain the molecular defects associated with several well known genetic mutations in mice, and has opened the door to understanding the role of this factor in development. Similar genetic defects may exist in humans as well. The aim of this paper is to review the biological structure and activities of this new growth factor, and to discuss its potential applications in clinical medicine.

Animals↗
PubMed1992

Developmental abnormalities in Steel17H mice result from a splicing defect in the steel factor cytoplasmic tail.

The murine dominant White spotting (W) and Steel (Sl) loci encode the c-kit tyrosine kinase receptor and its cognate ligand steel factor (SLF), respectively. Mutations at either locus produce deficiencies in the same three migratory cell populations--those giving rise to pigment cells, germ cells, and blood cells. The identification of the gene products of these two loci combined with the plethora of W and Sl mutations available for molecular analysis offers a unique opportunity to dissect the role of a tyrosine kinase receptor and its cognate ligand during development in a fashion not possible for most other mammalian genes. Among the most interesting Sl mutations available for study are those that induce sterility in only one sex. In studies described here, we show that one of these alleles, Sl17H, which in the homozygous condition induces sterility in males but not females, is the result of a splicing defect in the SLF cytoplasmic tail. We also characterize the nature of the germ cell defects in male and female Sl17H mice and show that both sexes are affected equally during embryonic but not postnatal development. These studies provide new insights into the role of SLF in germ cell development and indicate that the cytoplasmic domain of SLF is important for its normal biological function.

Amino Acid Sequence↗
PubMed1992

Efficacies of Novel N-Halamine Disinfectants against Salmonella and Pseudomonas Species.

Six novel N-halamine compounds of potential importance as disinfectants to the food-processing industry were tested against Salmonella enteritidis, Salmonella gallinarum, Salmonella typhimurium, and Pseudomonas fluorescens in aqueous solution. Inactivation times for 10-fold reductions were determined as a function of water quality at pH 6.5 and 25 degrees C. Phenol coefficients for the efficacies of the compounds against S. enteritidis have been reported also. When both stability and efficacy data are considered, as well as cost of production, two compounds, 1,3-dichloro-2,2,5,5-tetramethylimidazolidin-4-one and 1-chloro-2,2,5,5-tetramethylimidazolidin-4-one, offer the greatest potential as biocides for the food-processing industry.

Journal Article↗
PubMed1992

Pharmacokinetic parameters of recombinant mast cell growth factor (rMGF).

The gene product of the murine Steel (Sl) locus encodes an early-acting hematopoietic growth factor that is a ligand for the c-kit protooncogene. Several cDNAs for the Sl gene product, known as mast cell growth factor (MGF), stem cell factor (SCF), or kit ligand (KL), have recently been isolated, and both soluble and membrane-associated versions have been shown to be biologically active. The potential for therapeutic usage of recombinant MGF (rMGF) indicated a need for determining the biodistribution and elimination parameters of this cytokine. Pharmacokinetic studies demonstrated that radiolabeled rMGF had a distribution half-life of 2 min and an elimination half-life of 2.1 h in wild-type mice following iv injection, during which a striking localization of labeled rMGF in the lungs was noted. When administered by subcutaneous injection the elimination half-life was prolonged to 8.4 h. The primary sites of rMGF elimination appeared to be the kidneys and the liver. Pharmacokinetic analysis of labeled rMGF in mutant Sl/Sld mice, which are mast cell deficient, demonstrated similar distribution and elimination half-lives compared to wild-type mice (1.4 min and 1.8 h, respectively). In addition, the biodistribution pattern of the labeled rMGF in Sl/Sld mice was similar to that observed in wild-type mice, including the striking localization to the lungs. Binding of radiolabeled rMGF to lungs in vivo subsequent to iv injection was completely inhibited by excess unlabeled rMGF. Interestingly, mice that received an iv injection of the higher doses of rMGF (15 micrograms) demonstrated profound respiratory distress and hypotension within minutes of administration. Histologic analysis of lungs from such mice revealed extensive mast cell degranulation, which was associated with vasodilatation and pronounced hyperemia of virtually all pulmonary vessels. The respiratory distress in normal mice was probably a consequence of mast cell degranulation induced by rMGF since similar findings were not observed in Sl/Sld mice injected with identical concentrations of rMGF.

Animals↗
PubMed1992

Retroviral vector-mediated gene transfer into primitive human hematopoietic progenitor cells: effects of mast cell growth factor (MGF) combined with other cytokines.

Retroviral vector-mediated gene transfer into human hematopoietic stem cells may permit gene therapy of numerous genetic diseases. Stimulation of marrow with hematopoietic growth factors (HGFs) has been shown to increase the level of retroviral transduction. We have examined the effects of recombinant human mast cell growth factor (MGF), alone and in combination with other HGFs, on the efficiency of gene transfer into human hematopoietic progenitor cells. MGF acts in concert with interleukin 3 (IL-3) and interleukin 6 (IL-6) to increase the percentage of CD34+ progenitors transduced with a retroviral vector expressing the neo gene. The most potent combination of growth factors that we examined, interleukin 1 (IL-1)/IL-3/IL-6/MGF, resulted in the conferral of G418 resistance to 45% of progenitors and long-term culture-initiating cells. Extending the time of cocultivation of the marrow cells with the vector-producing cells did not further increase gene transfer frequency, suggesting that the amount of available vector is not limiting. To analyze the effects of the HGF on gene transfer into more primitive hematopoietic progenitors, CD34+ cells were isolated from marrow samples that were purged of committed progenitor cells by treatment with 4-hydroperoxycyclophosphamide (4-HC). Preculturing the CD34+ 4-HC-treated cells with the combination of four HGF (IL-1/IL-3/IL-6/MGF) permitted transduction of 20%-28% of the progenitors that formed colonies after 30 days in culture. These results demonstrate that MGF in combination with other HGFs enhances gene transduction of human hematopoietic progenitor cells.

Antigens, CD↗
PubMed1992

Ex vivo expansion of peripheral blood progenitor cells with recombinant cytokines.

Peripheral blood mononuclear cells were isolated and cultured in the presence of Steel factor and/or PIXY321, a GM-CSF/IL-3 fusion protein. We compared the number of colony forming cells (CFC) per culture on day zero to the number of CFC after liquid culture in the presence of these cytokines. After a four day incubation with PIXY321 and Steel factor the number of CFU-GM increased 5.6-fold and the number of BFU-E increased 2.2-fold in four separate experiments. The expansion on day 8 post incubation was 16.1-fold for myeloid colony formation and 9.7-fold for erythroid colony formation. These studies demonstrate the potential to expand CFC from peripheral blood with a simple ex vivo culture procedure.

Cell Count↗
PubMed1992

Effects of timolol, betaxolol, and levobunolol on human tenon's fibroblasts in tissue culture.

Evidence has been found suggesting that long-term therapy with topical antiglaucoma medications may decrease the success of glaucoma filtering surgery. To investigate this question further, the antiproliferative effects of the preservative benzalkonium chloride and three pure and commercially available beta-adrenergic antagonist preparations (timolol, betaxolol, and levobunolol) were studied on tissue cultures of human Tenon's capsule fibroblasts. Each drug preparation was tested on three different cell lines. Fibroblast growth was measured with tritiated thymidine uptake and hexosaminidase assays. Trypan blue uptake was used to assess cell viability microscopically. The commercially available preparations containing benzalkonium chloride and those of betaxolol and levobunolol without the preservative had similar inhibitory doses for 50% of cells. The timolol preparation without preservative was significantly less toxic than its commercially available one. The three tested beta-adrenergic blockers did not stimulate fibroblast proliferation directly in this in vitro model. Even when the cultures were washed free of the drugs, growth continued to be suppressed, suggesting that the inhibition was not reversible. An increase in fibroblasts and inflammatory cells after long-term antiglaucoma medical therapy thus may be caused not by a direct stimulation of cell proliferation but by chronic inflammation from the irritating effects of antiglaucoma medications and/or their preservatives.

Benzalkonium Compounds↗