Metastatic breast carcinoma in the pelvis.
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Adenocarcinoma, Scirrhous↗
Biomedical subjects
W G Taylor
Publications and source records attributed to W G Taylor.
Nuclear magnetic resonance spectra and mosquito repellent properties of some oxazolidine derivatives of 5-methyl-4-hexenal.
Seven novel oxazolidines from 5-methyl-4-hexenal were synthesized, characterized, and evaluated in a cloth test system as mosquito repellents. These heterocycles were a homologous series of 3-alkanoyl-2-(4'-methyl-3'-pentenyl)oxazolidines, with substituents at N-3 ranging from acetyl to n-nonanoyl groups. Carbon-13 NMR spectroscopy at 100.6 MHz revealed that these molecules existed in deuterochloroform solution as unequal mixtures of cis (syn)- and trans (anti)-rotational isomers. At 25 degrees C, this isomerism at the amide group resulted in two detectable 13C signals for all assigned carbon atoms, except for the methyl carbons of the isopentenyl side chain. Differences in 13C chemical shifts for the methyl group at N-3 were observed up to the n-butanoyl homologue. The 13C signal assignments were aided by the distortionless enhancement by polarization transfer (DEPT) spectral editing technique. Two-dimensional carbon-hydrogen chemical shift correlation experiments aided in partially interpreting the complex proton spectra of these molecules. Repellent activity of the acetyl, trideuteroacetyl, and propionyl derivatives was observed in bioassays with three mosquito species, but the long-chain homologues were practically ineffective as mosquito repellents.
Aedes↗
Purification and characterization of a newly identified growth factor specific for epithelial cells.
A growth factor specific for epithelial cells was identified in conditioned medium of a human embryonic lung fibroblast cell line. The factor, provisionally termed keratinocyte growth factor (KGF) because of its predominant activity on this cell type, was purified to homogeneity by a combination of ultrafiltration, heparin-Sepharose affinity chromatography, and hydrophobic chromatography on a C4 reversed-phase HPLC column. KGF was both acid and heat labile and consisted of a single polypeptide chain of approximately 28 kDa. Purified KGF was a potent mitogen for epithelial cells, capable of stimulating DNA synthesis in quiescent BALB/MK epidermal keratinocytes by greater than 500-fold with activity detectable at 0.1 nM and maximal at 1.0 nM. Lack of mitogenic activity on either fibroblasts or endothelial cells indicated that KGF possessed a target cell specificity distinct from any previously characterized growth factor. Microsequencing revealed an amino-terminal sequence containing no significant homology to any known protein. The release of this growth factor by human embryonic fibroblasts raises the possibility that KGF may play a role in mesenchymal stimulation of normal epithelial cell proliferation.
Animals↗
Pathways in which growth factors and oncogenes interact in epithelial cell mitogenic signal transduction.
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Animals↗
Nuclear war and plastic surgery.
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Nuclear Warfare↗
Interactions of growth factors and retroviral oncogenes with mitogenic signal transduction pathways of Balb/MK keratinocytes.
Balb/MK epidermal keratinocytes require epidermal growth factor (EGF) for growth in serum-containing medium and terminally differentiate in response to high Ca++ concentration. Several oncogenic retroviruses have been shown to relieve the EGF requirement and to block calcium-induced terminal differentiation. We developed a chemically defined medium to investigate the minimum growth factor requirements for Balb/MK cells, as well as how such requirements might be altered by retroviral oncogenes. In this medium insulin, apparently acting as IGF-1, and EGF supported cell growth in a manner comparable to serum and EGF. Acidic as well as basic fibroblast growth factor (FGF) substituted for EGF but not insulin in supporting Balb/MK proliferation. Infection with retroviruses containing v-ras oncogenes (v-H-ras, v-Ki-ras), oncogenes derived from growth factor receptors (v-erbB, v-fms) or the v-mos oncogene permitted growth in defined medium containing insulin but lacking EGF. The v-fgr oncogene, a member of the src subfamily, was unique in conferring independence from both insulin and EGF. Our findings establish the applicability of this system for biologic assay of epithelial cell growth factors as well as identification of specific growth factor requirements that can be altered or complemented by the actions of specific oncogenes.
Animals↗
Chiral-phase capillary gas chromatography and mosquito repellent activity of some oxazolidine derivatives of (+)- and (-)-citronellol.
Fourteen stereoisomeric mixtures of six oxazolidine heterocycles and one amino alcohol were obtained from commercial samples of (+)-, (-), or (+/-)-citronellol and evaluated in a cloth test system as mosquito repellents. A technical sample of 3-acetyl-2-(2',6'-dimethyl-5'-heptenyl)oxazolidine, evaluated initially as a mixture of two pairs of enantiomers, was an effective repellent against two of the three species of mosquitoes. To study repellency-stereochemistry relationships, 1 was isolated as fractions G-1, G-2, I-1, and I-2, corresponding to representative samples of the four possible stereoisomers. The enantiomeric composition of 1, but not of the other derivatives, could be determined by chiral-phase capillary gas chromatography with Chirasil-Val. Two chiral phases showed that experimental samples of 1, and hence the commercial samples of 2, were composed of not more than 90% of one enantiomer. Differences in repellency of isomerically enriched samples were generally small. Fraction G, a mixture of diastereoisomers readily prepared from (+)-2, was considerably more effective than standard insect repellents in bioassays with three mosquito species.
Acyclic Monoterpenes↗
Repair of DNA-protein cross-links in an excision repair-deficient human cell line and its simian virus 40-transformed derivative.
DNA-protein cross-links are induced in mammalian cells by X-rays, ultraviolet light, fluorescent light, and numerous chemical carcinogens. Others have shown that these cross-links are repaired by normal cells but that excision repair-deficient xeroderma pigmentosum (XP) Group A cells, XP12BE, are deficient in repair of these bulky adducts. This paper compares the DNA-protein cross-link repair competency of another XP Group A strain, XP20S, with its more rapidly proliferating simian virus 40-transformed derivative line and with normal human skin fibroblasts. DNA-protein cross-links were induced with 20 microM transplatinum(II)diamminedichloride and assayed by the membrane alkaline elution procedure of Kohn. Treated and untreated cells are lysed on a polycarbonate membrane filter, and the coelution rates of the DNA at pH 12.2 are compared; DNA-protein cross-links retard elution of DNA. The repair competency of XP20S cells for trans-platinum(II)diamminedichloride-induced DNA-protein cross-links was similar to that of XP12BE cells, but the competency of the simian virus 40-transformed XP20S cells was nearly equal to that of normal human skin fibroblasts. These results suggest that either cell cycling compensates for the genetic deficiency present in the nucleotide excision process of XP Group A cells or that a process other than nucleotide excision can repair these lesions; this process requires cell cycling or activation by the virus.
Cell Line↗
Serum-independent modulation of hemicyst formation by dissolved oxygen in postconfluent epithelial monolayers.
Hemicyst formation is considered a manifestation of either transepithelial solute and fluid movement or secretory activity in culture. This study shows that hemicyst formation in postconfluent monolayers of rhesus monkey kidney (LLC-MK2) cells is modulated by the dissolved oxygen concentration (PO2) of the culture medium. Either daily replacement of serum-free medium or displacement of the gas phase with 18% vol/vol O2 (initial medium PO2 = 125 to 135 mm Hg) enhances formation of hemicysts. Use of 30% O2 (medium PO2 approximately equal to 175 mm Hg) does not further increase the incidence, but neither 10% O2 (medium PO2 = 90 to 95 mm Hg) nor 1% O2 (medium PO2 = 35 to 50 mm Hg), the approximate range of dissolved oxygen values in blood, supports hemicyst formation unless cultures are gently rocked to disrupt diffusion gradients. Phase photomicrography of living cultures shows that the surface of a turgid hemicyst is furrowed, and cinephotomicrography reveals that the walls vibrate subtly. When hypoxic conditions (0 to 1% O2) are introduced this vibration ceases within 2 to 3 h, whereas collapse and disappearance of turgid hemicysts requires 18 to 20 h, seems virtually synchronous, and is reversible. Hemicysts form in a broad osmotic range, and increased electrolyte concentration increases the incidence. Hemicysts persist in locally dense areas when cell-free strips are etched in the postconfluent monolayer; no DNA synthesis is detected under these conditions, but two-dimensional cell spreading into the denuded area is seen along the edge of the wound. We conclude that the dissolved oxygen supply in the cellular microenvironment modulates functional expression by differentiated kidney epithelial cells in culture and that increased electrolyte concentration also enhances expression of this phenotypic marker.
Animals↗
Approaches to enhance proliferation of human epidermal keratinocytes in mass culture.
Because of interest in mechanisms of carcinogenesis in human epithelial cells, quantitative procedures were developed for the mass culture of human epidermal keratinocytes in the absence of feeder cells. Several approaches were used to enhance proliferation since target cells are considered most susceptible to transformation if they are treated with carcinogenic agents during DNA synthesis. Mass cultures of enzymatically dispersed human foreskin were initiated in collagen-coated flasks containing medium NCTC 168 with 10% Chelex 100-treated horse serum. Under these conditions, human keratinocytes required a higher calcium ion concentration ([Ca2+]) than that reported for suspensions plated at low cell density. Neither the cohesiveness of the epidermal sheet nor continued proliferation was maintained by 0.15 mM Ca2+; 0.3 mM Ca2+ maintained these properties in primary culture only. A concentration of 1.0 mM Ca2+ provided the highest cell yield for prolonged growth as determined by the enumeration of cell nuclei isolated by citric acid. Reproducibility of successful initiation was achieved by inoculation of cells into a medium designed for clonal growth followed by culture in medium NCTC 168. Thus the balance of nutrients and electrolytes must be adjusted to satisfy the requirements of a dynamically expanding keratinocyte population.
Calcium↗
Modulation of epithelial cell proliferation in culture by dissolved oxygen.
Modulation of epithelial cell proliferation by the dissolved oxygen concentration (PO2) of the growth medium was assessed with primary human foreskin epithelium and a continuous monkey kidney epithelial cell line (LLC-MK2). Direct measurement of the growth medium PO2 provides the first quantitative evaluation of epithelial cell proliferation as a function of PO2. Sustained proliferation of LLC-MK2 cells occurs in serum-free medium equilibrated with a gas phase containing 18% or 30% O2 v/v. Mid-logarithmic phase cultures rapidly consume dissolved oxygen; this results in a 60-70 mm Hg decline in PO2, and leads to a stable growth medium PO2 between 70 and 100 mm Hg, well above anoxic values. In contrast, if culture medium is equilibrated with a gass phase containing 0% or 1% O2 v/v to yield a growth medium PO2 - approximately 20-40 mm Hg, proliferation of LLC-MK2 and primary foreskin epithelial cells is retarded, and LLC-MK2 cells use little dissolved oxygen. Gentle, continuous rocking to prevent diffusion gradient formation enhances proliferation slightly at the higher PO2, but neither periodic fluid renewals nor continued rocking stimulates cells retarded by a lowered oxygen concentration to resume proliferation. The data collectively demonstrate that epithelial cell proliferation requires a PO2 greater than 40 mm Hg, and threshold requirements are probably closer to 70 mm Hg. Glycolysis continues at a PO2 insufficient for proliferation, but more lactic acid accumulates in actively proliferating cultures than in cultures equilibrated with 0% oxygen. We conclude that epithelial cells in vitro both consume more oxygen and require a higher PO2 for continued proliferation, and that the oxygen requirement for epithelial cell proliferation exceeds that of a comparable population of fibroblasts for which low oxygen may enhance survival and proliferation.
Animals↗
Effect of visible light on benzo(a)pyrene binding to DNA of cultured human skin epithelial cells.
The ubiquity of the photosensitive carcinogen benzo(a)pyrene (BP) and visible light in the environment suggests that their interaction might lead to photoproducts harmful to humans. To test the combined impact of these two agents on human epithelial cells, binding of BP to cellular DNA was assessed following treatment of cultures with BP and low-intensity (4.6 watts/sq m) intermittent (12 hr daily, 3 to 5 days) cool white fluorescent light. Light exposure reduced the formation of covalent BP adducts 20-fold (from 150 to 7 pmol BP per mg DNA) in cells treated with 1 microgram BP per ml and completely inhibited cytotoxicity; even with 10 microgram BP per ml, light exposure markedly inhibited cytotoxicity. However, at low BP dosage (0.1 microgram/ml), covalent adducts (2 pmol/mg DNA) to cellular DNA are produced and their formation is not influenced by light. These adducts persisted for at least 7 days following treatment; this observation suggests that chronic low-level exposure of human epithelium to BP may lead to an accumulation of DNA damage.
Benzo(a)pyrene↗
A new culture medium for human skin epithelial cells.
A new culture medium, NCTC 168, has been designed for human skin epithelial cells. This medium formulation was developed, by combining and testing at various concentrations, components of media NCTC 135 and 163, since a 1:1 mixture of these two media with 10% horse serum supplement was found to promote epithelial cell outgrowth from human skin explants. The buffer system in NCTC 168 maintains the pH of the medium between 7.0 and 7.2. In contrast to other media tested, NCTC 168 with 10% horse serum is capable of initiating and sustaining larger epithelial cell outgrowths. Explants in serum-supplemented NCTC 168 in the absence of feeder cells reproducibly yield confluent epithelial cell sheets apparently free of fibroblasts after only 19 to 28 days as compared with 5 weeks or longer for the other media tested. NCTC 168 also supports passage of human epithelial cells to the sixth subculture generation without feeder cells. Electron microscopy has shown the presence of desmosomes and tonofilaments in the passaged cells indicating the epithelial nature of the cells. The addition of epithelial growth factor, hydrocortisone and insulin at 5 ng per ml, 4 microgram per ml and 5 microgram per ml, respectively, did not appreciably enhance the growth of the epithelial cells.
Blood↗
Fluorescent light-induced chromosome damage in human IMR-90 fibroblasts. Role of hydrogen peroxide and related free radicals.
Exposure of human fibroblasts (IMR-90) to cool-white fluorescent light causes chromatid breaks and exchanges. This chromatid damage is caused largely by the production of hydrogen peroxide (H2O2) since it can be prevented almost completely by the addition of catalase. In support of this conclusion, exogenous H2O2 is shown to induce chromatid breaks. The clastogenic amounts of H2O2 generated during light exposure are formed within the cell since cells illuminated in saline showed the same extent of damage as cells in culture medium. Addition of selenite to the cultures during light exposure significantly decreases the chromatid damage in a dose-related manner and may be necessary to maintain sufficient activity of glutathione peroxidase. The free hydroxyl radical, . OH, appears to be partially responsible for the light-induced chromatid damage. Of the free-radical scavengers tested, i.e., mannitol, vitamin E, and dimethyl sulfoxide, only mannitol, which scavenges . OH, significantly decreases the light-induced chromatid damage. Thus, both . OH and H2O2 formed within the cell during light exposure are agents that directly or indirectly cause chromatid damage.
Catalase↗
Effect of intensity and wavelength of fluorescent light on chromosome damage in cultured mouse cells.
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Animals↗
Density-dependent effects of oxygen on the growth of mammalian fibroblasts in culture.
Various concentrations of oxygen were used to determine the optimum culture medium PO2 for survival and proliferation of attached human and mouse fibroblasts grown from different inoculum sizes. When T-15 flasks were seeded with less than or equal to 2 X 10(4) cells (less than or equal to 1.3 X 10(3) cells/cm2), the highest plating efficiencies and cell yields were obtained with a culture medium PO2 of 40-60 mm Hg. At higher inoculum sizes (10(5) cells per T-15) used routinely for mass cultured, no difference in cell yield or glycolytic activity was observed between cultures gassed with atmospheric, i.e., 18% O2 (growth medium PO2 approximately equal to 125-135 mm Hg) and those gassed with 1% O2 (growth medium PO2 approximately euqal to 40-60 mm Hg). The enhanced clonal growth observed at the latter PO2 results from an increased proliferation rate rather than more efficient attachment and survival of inoculated cells. Glucose uptake and lactic acid accumulation were increased in sparse cultures sparged with 1% O2. A slight extension of lifespan was observed in WI-38 cells serially subcultured with a gas phase of 1% O2.
Animals↗
Herpesvirus salmonis: Characterization of a New Pathogen of Rainbow Trout.
A new agent, provisionally designated Herpesvirus salmonis, was isolated from post-spawning rainbow trout (Salmo gairdneri) and studied primarily in the RTG-2 rainbow trout cell line. Infection of RTG-2 cells resulted in the formation of syncytia and Cowdry type A intranuclear inclusions. Replication occurred regularly at 5 and 10 degrees C, but was inconsistent at 15 degrees C, largely inhibited at 0 degrees C, and completely inhibited at 20 degrees C or higher. The virus was acid, heat, ether, and chloroform labile, but stable to freezing and thawing. It did not hemagglutinate. Viral DNA had a buoyant density of 1.709 g/cm(3) and a guanine-cytosine value of 50%. Hexagonal nucleocapsids had a diameter of 90 nm and were first seen in nuclei at 36 h. Enveloped forms measured about 150 nm and occurred both cytoplasmically and extracellularly. At 10 degrees C, a one-step growth culture required about 96 h; cell-associated virus peaked at about 10(5) PFU/ml and exceeded released virus by a factor of about 10.
Journal Article↗