Search PubMed⌕ Search

Biomedical subjects

R W Tucker

Publications and source records attributed to R W Tucker.

At least 37 records · Page 2Linked to original sources
PubMed1989

Intracellular calcium alterations in response to increased external calcium in normal and neoplastic keratinocytes.

Normal keratinocytes proliferate when cultured in medium with 0.02-0.10 mM calcium and terminally differentiate when medium calcium is increased to greater than 0.1 mM. In contrast, neoplastic keratinocyte cell lines maintain the potential for continued cell renewal and survive when external calcium is increased. In order to determine whether elevation of extracellular calcium produced changes in intracellular free calcium (Cai) levels, Cai was measured in individual living keratinocytes by use of the fluorescent calcium probe fura-2. Most normal keratinocytes responded to increased extracellular calcium by a gradual 2- to 3-fold increase in Cai lasting for at least 28 min. A subpopulation displayed a sharp peak of Cai at 2 min. In contrast, the Cai level in neoplastic cells in either low or high calcium medium was 2- to 3-fold higher than that in normal cells, and all cells in the population showed a transient 4- to 9-fold elevation of Cai 2 min after external calcium was increased. Thus normal and neoplastic keratinocytes differ in the level of Cai under low calcium conditions and in their response to elevated external calcium. The regulation of Cai in keratinocytes may be important in determining their potential for terminal differentiation.

Animals↗
PubMed1989

A two-step timed sequential treatment for acute myelocytic leukemia.

Since 1980, adults with acute myelocytic leukemia (AML) have been treated on two clinical studies using intensive timed sequential therapy. All patients ages 16 to 80, including those with secondary AML (SAML) and those with AML preceded by a hematologic disorder (AHD), were treated, regardless of medical complications at the time of diagnosis. The first study combined high doses of cytarabine (ara-C, AC) and daunorubicin (DRN, D) in sequence (Ac2-D-Ac) and resulted in a complete remission rate of 55%. A group of these patients selected by functional status was able to receive a second course of therapy in remission, which resulted in a disease-free survival (DFS) of greater than 40% at 7 years. Because of toxicity in that study, 114 patients were entered on a second trial initiated 4 years ago, using a less aggressive first course, with amsacrine, to achieve a stable remission (Ac2-D-Amsa). This first treatment was followed by a more intensive second course (Ac6-D-Ac). With this two-step approach, a higher complete remission (CR) rate (76% for de novo AML and 54% for SAML-AHD) was achieved, and more patients were able to receive the second course of therapy. At the current median follow-up of 26 months, the median duration of DFS and overall survival are 11 and 14 months for patients with de novo AML. Age less than or equal to 55 is the most significant prognostic factor for both prolonged DFS and overall survival, with median durations of 17 and 18 months, respectively, for these younger patients. Patients with SAML-AHD remain relatively refractory to treatment despite aggressive chemotherapy, with median durations of DFS and overall survival of 9 months and 5 months, respectively.

Adolescent↗
PubMed1988

Microtubule changes and cytotoxicity in leukemic cell lines treated with taxol.

Taxol, a diterpenoid plant product that enhances the polymerization of tubulin, is currently entering clinical trials in the treatment of human leukemia. In order to develop an in vitro assay to predict tumor sensitivity to taxol, human leukemic cell lines were exposed to clinically achievable concentrations of taxol for relevant exposure periods. Changes in microtubules visualized by indirect immunofluorescence were compared to drug sensitivity measured by a clonogenic assay. Taxol produced either multiple mitotic asters in G2/M or microtubule bundling throughout the cell cycle. In cells that were relatively resistant to taxol, microtubule bundling was reversible while microtubule bundling in relatively sensitive cells persisted in the presence or absence of taxol. In contrast, aster formation was unrelated to cytotoxicity in any cell line. In the future, these microtubule effects may be useful in predicting the chemotherapeutic efficacy of taxol.

Alkaloids↗
PubMed1988

Signal transduction for proliferation and differentiation in keratinocytes.

In mouse and human epidermis, the Ca2+ environment of the basal cell layer is substantially below serum Ca2+, while that of the granular cell layer is unusually high. Reduction of extracellular Ca2+ concentration (Cao) in the medium of keratinocyte cultures maintains a basal cell phenotype while serum Ca2+ concentrations induce terminal differentiation. Measurements of intracellular Ca2+ (Cai) by the use of Fura 2 and digital imaging technology reveal that Cai increases 10-20-fold in response to an increase in Cao and remains elevated. Concomitant with the rise in Cai is an increase in the metabolism of phosphatidylinositol (PI) to yield inositol phosphates and diacylglycerol. PI metabolism is also stimulated by calcium ionophores suggesting that a rise in Cai is directly responsible. The consequent increase in diacylglycerol and Cai would activate protein kinase C, an event known to trigger epidermal differentiation. Specific Cao and Cai determine the expression of individual markers of keratinocyte differentiation in vitro. These findings may account for the importance of the Ca2+ gradient for maintaining regulated growth and differentiation of the epidermis in vivo.

Animals↗
PubMed1986

Cytosolic free calcium and DNA synthesis in BALB/c 3T3 cells: aequorin luminescence studies.

In order to investigate how growth factors stimulate DNA synthesis, we studied the relationship between early increases in the intracellular concentration of free calcium (Cai) and early mitogenic events (competence) in quiescent BALB/c 3T3 cells. Cai was measured by aequorin luminescence, and DNA synthesis was quantitated by autoradiography. While both serum (2%) and platelet-derived growth factor (2 U/ml) stimulated large increases in Cai without inducing competence, fibroblast growth factor (100-400 ng/ml) produced a large number of competent cells without causing a detectable increase in Cai. Thus, there was no correlation between the magnitude of Cai increase and subsequent competence for DNA synthesis.

Aequorin↗
PubMed1986

Deceased numbers of spindle and cytoplasmic microtubules in hamster embryo cells treated with a carcinogen, diethylstilbestrol.

Diethylstilbestrol (DES) has been shown to induce neoplastic transformation in the absence of measurable mutations at specific loci in Syrian hamster embryo cells. It has been proposed that DES induces cell transformation via the production of aneuploidy. In the present study we document that concentrations of DES that cause aneuploidy also produce abnormal or arrested mitotic spindles. Thus, DES may disrupt spindle microtubules and produce aneuploidy that results in disordered gene expression and eventual neoplastic transformation.

Aneuploidy↗
PubMed1985

Intracellular type A retrovirus movement associated with an intact microtubule system.

Intracytoplasmic type A particles known to be precursors to type B retroviruses in murine, hamster and marsupial cells are closely associated with microtubules and microtubule organizing centres. In this publication, the active participation of microtubules in the intracellular transport of the particles to the cell surface has been examined in NIH 3T3 cells infected with M432 virus using vincristine sulphate (VCR) as inhibitor of microtubule polymerization. The release of virus at different times after exposure to VCR was quantified by reverse transcriptase determinations of cell supernatants and by electron microscopic quantification of the number of virions at the cell surface using freeze-dried whole cell replicas. These studies indicate that VCR inhibits both microtubule polymerization and virus release, and thus suggest that intact cytoplasmic microtubules are necessary for intracellular transport and release of virus.

Animals↗
PubMed1983

Centrioles, primary cilia and calcium in the growth of Balb/c 3T3 cells.

In every mammalian cell the interphase centriole forms a primary cilium during some part of the cell cycle. Many sensory receptors are modifications of primary cilia, but a sensory role for primary cilia in mammalian cells has not been proven. Nevertheless, we have found that both growth factors and calcium ionophore (A23187) induce calcium fluxes and shortening of the primary cilium. This evidence raises the question of whether the primary cilium is involved in calcium fluxes that are necessary for growth stimulation in mammalian cells.

Animals↗
PubMed1981

Interaction of serum and cell spreading affects the growth of neoplastic and non-neoplastic fibroblasts.

Both growth factor availability and cell-to-cell contact have been mechanisms used to explain cell growth regulation at high cell density. Recently Folkman and colleagues have shown that changes in cell shape, rather than cell-to-cell contact, can regulate the growth of fibroblasts. However, in those studies the relation between serum and shape regulation of growth was not studied, nor were neoplastic and non-neoplastic cells compared. In this report we have studied these aspects by varying cell spreading and serum concentration independently for 2 non-neoplastic and 3 neoplastic cell lines. Cell spreading (projected cell area) was controlled by decreasing the adhesiveness of tissue culture growth was measured as the increase in cell number/day. We have found that more spreading increased net growth of both neoplastic and non-neoplastic cells, while less spreading (toward rounded configuration) depressed growth. There were also quantitative differences between neoplastic and non-neoplastic cells. Neoplastic cells continued to grow under conditions of cell rounding, which completely prevented the growth of their non-neoplastic counterparts. Some neoplastic cells also tended to show little or no increase in net cell number for serum concentrations above 10% as cells became more spread; in contrast, all non-neoplastic cells grew more with increasing concentrations of serum as they became well spread. Thus, in normal cells, it appears that the sensitivity of cells to humoral factors is governed by cell spreading. This interaction between serum and cell shape is less prominent in some neoplastic cells.

Animals↗
PubMed1980

Transformed and neoplastic phenotype: reversibility during culture by cell density and cell shape.

The ability of parental and transformed mouse C3H (10T 1/2) cells to grow when held at a graded series of cell shapes (flat to round) was studied using substrata of decreasing adhesivity. The parental cells showed a decrease in growth when they reached the most rounded configurations. In contrast, transformed cells proliferated at the same rate regardless of cell shape. In addition, when transformed cells were serially passaged at low density on highly adhesive plastic, which maintained cells in a flat configuration, a reversion from the transformed phenotype to a non-transformed phenotype occurred with the concomitant return of growth control by cell shape. However, when transformed cells were passaged at low density on a substratum which prevented cell spreading, the reversion to the parental phenotype did not occur and the cells escaped the growth control of cell shape and remained tumorigenic. Thus, in this cell system it appears that a change in cell configuration can dictate whether or not the transformed and neoplastic phenotype will be expressed.

Animals↗
PubMed1980

Regulation of the Balb/c-3T3 cell cycle-effects of growth factors.

The platelet-derived growth factor (PDGF), which is found in serum but not in plasma, has been purified to homogeneity; it stimulates replication at a concentration of 10(-10) M. Brief treatment with PDGF causes density-inhibited Balb/c-3T3 cells to become competent to synthesize DNA; pituitary fibroblast growth factor (FGF) or precipitates of calcium phosphate also induce competence. Continuous treatment with plasma allows competent, but not incompetent, cells to synthesize DNA. A critical component of plasma is somatomedin, a group of hormones with insulin-like activity; multiplication-stimulating activity (MSA) or insulin replace plasma somatomedin in promoting DNA synthesis. We have studied the molecular correlates of competence and the role of SV40 gene A products in regulating DNA synthesis. Treatment of quiescent cells with pure PDGF or FGF causes the preferential synthesis of five cytoplasmic proteins (approximate molecular weight 29,000, 35,000, 45,000, 60,000, and 72,000 detected by SDS-PAGE under reducing conditions). Two of these competence-associated proteins (29,000 and 35,000 daltons) are insulin, or epidermal growth factor (EGF). PDGF, FGF, or calcium phosphate induce an ultrastructure change within the centriole of 3T3 cells; this ultrastructural modification of the centriole is detectable by immunofluorescence within 2 h or PDGF treatment. Plasma, EGF, or MSA do not modify the centriole. SV40 induces replicative DNA synthesis in growth-arrested 3T3 cells but does not cause this alteration in centriole structure. Gene A variants of SV40, including a mutant with temperature-sensitive (ts) T-antigen (ts A209), a deletion in t-antigen (dl 884), and several ts A209 strains containing t-antigen deletions were used to induce DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells is not t and is not temperature sensitive. This SV40 gene function overrides the cellular requirement for hormonal growth factors. It does not induce transient centriole deciliation, a hormonally regulated event.

Animals↗
PubMed1980

The cytoskeleton in Chediak-Higashi syndrome fibroblasts.

The Chediak-Higashi syndrome (CHS) trait is expressed in cultured human skin fibroblasts as an abnormal perinuclear concentration of moderately enlarged lysosomes. The cytoskeleton of CHS fibroblasts appears intact. Microtubules are normal in number and morphology, as assessed by colchicine binding studies, antitubulin immunofluorescence, and electron microscopy. Deformability by shear force is unaltered and microfilaments are abundant. However, CHS lysosomes appear to interact abnormally with the cytoskeleton, since the perinculear aggregation partially disperses after depolymerization of cell microtubules with colchicine. These results suggest that CHS is associated with a defect of either the lysosomal membrane itself or of lysosomal membrane-microtubule interaction.

Acridine Orange↗