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D W Barnes

Publications and source records attributed to D W Barnes.

At least 37 records · Page 2Linked to original sources
PubMed1993

Characterization of human plasma growth inhibitory activity on serum-free mouse embryo cells.

Serum-free mouse embryo (SFME) cells are a cell line derived in medium in which serum is replaced with growth factors and other supplements. These cells display unusual properties: a) they do not lose proliferative potential or show gross chromosomal aberration upon extended culture, b) they depend on epidermal growth factor (EGF) for survival, and c) they are reversibly growth inhibited by plasma and serum. Transfection of SFME cells with oncogenes (ras, neu, SV40 T antigen) results in cells that grow in serum-supplemented medium and no longer require EGF for survival. The growth inhibitory activity of human plasma on SFME cells was investigated. The activity was present in delipidated plasma and was not dialyzable against 1 M acetic acid. The activity precipitated in 33% methanol, bound to concanavalin A-agarose and was retarded by Sephadex G-50 in 200 mM acetic acid. A fifty- to one-hundred-fold purification was achieved, although most of the differential inhibition of untransformed vs. transformed cells was lost in the course of the purification.

Animals↗
PubMed1993

Culture of cells from juvenile worms of Schistosoma mansoni.

Tissue disruption methods were developed and serum-free cell culture media formulated for the maintenance in vitro of cells from juvenile worms (day 18 after infection) of Schistosoma mansoni. Cultures maintained viability for up to 6 mo when plated on a feeder layer of irradiated rat liver cells and survived primarily as clusters of small (2.5-4 microns diameter) cells with a high nuclear-to-cytoplasmic ratio and relatively few organelles identified by electron microscopy. Cultures synthesized a protein profile similar to that of intact worms, and the cell clusters maintained a time- and concentration-dependent contractile response to serotonin. Cells synthesizing DNA were detected by precursor incorporation and flow cytometry in cultures initially and also after several weeks in vitro, although the percentage of cells synthesizing DNA decreased with time. Efforts to identify peptide growth factor-responsive tyrosine phosphorylation were negative, and the overall amount of S. mansoni phosphotyrosine-containing proteins identified by western blot with anti-phosphotyrosine monoclonal antibody was much less than that found in a peptide growth factor-responsive mouse cell line.

Animals↗
PubMed1992

Death of serum-free mouse embryo cells caused by transforming growth factor beta 1 and effects of nutritional factors.

Transforming growth factor beta 1 (1 ng/ml) caused death of serum-free mouse embryo cells cultured in a medium consisting of a 1:1 mixture of Dulbecco's Modified Eagle's medium and Ham's F12 medium supplemented with fibronectin, insulin, transferrin, epidermal growth factor, and high density lipoprotein. Cell death occurred in the presence of polyunsaturated fatty acids including linoleic acid in the absence of selenium. The death could be reversed by adding alpha-tocopherol to the culture indicating a mechanism involving fatty acid peroxidation. Butylated hydroxytoluene was a poor suppressor of cell death in contrast to alpha-tocopherol. High density lipoprotein and fatty acid-free albumin also suppressed cell death at the level of 20 micrograms/ml and 1 mg/ml, respectively. Transforming growth factor beta 1 also caused a low rate of cell growth after heat treatment of the cells at 45 degrees C.

Albumins↗
PubMed1992

Functional changes in the pig kidney following irradiation with fractionated doses of fast neutrons (42 MeVd-->Be).

The right kidney of female Large White pigs, approximately 14 weeks old, was irradiated with fractionated doses of fast neutrons (42 MeVd-->Be). The total doses used were 6.6-9.2 Gy. Changes in kidney function, assessed as the functional index (FI, where FI = irradiated kidney function/unirradiated kidney function) or as individual kidney glomerular filtration rate (GFR) and effective renal plasma flow (ERPF), were serially determined up to 104 weeks after irradiation using 99Tcm-DTPA and 131I-hippuran renography. The animals were then euthanized, the kidneys removed and weighed. A dose-dependent reduction in FI was seen within 13 weeks of irradiation. Measuring individual kidney function revealed a hyperaemic response in both irradiated and unirradiated kidney 4 weeks after irradiation. This was followed by a dose-dependent reduction in irradiated kidney GFR and particularly ERPF. The ED50 value for the impairment in ERPF, assessed as the percentage of irradiated kidneys exhibiting a > or = 50% reduction in ERPF, was significantly lower than that for GFR, i.e. 7.20 +/- 0.10 Gy compared with 8.44 +/- 0.07 Gy (p < 0.001). A dose-related reduction in irradiated kidney weight was also observed. These fast neutron-induced changes in renal function and weight are qualitatively similar to those observed following photon irradiation of the pig kidney.

Animals↗
PubMed1991

Serial passage of embryonic human astrocytes in serum-free, hormone-supplemented medium.

We applied serum-free cell culture methods that allow extended proliferation of mouse astrocyte precursor cells to the multipassage culture of embryonic human brain cells. Cells were cultured in nutrient medium supplemented with insulin, transferrin, epidermal growth factor, fibroblast growth factor, heparin, high-density lipoprotein, and fibronectin. Cultures were maintained for a maximum of 70 population doublings before proliferation ceased. The cells synthesized glial fibrillary acidic protein, an astrocyte marker, and expression of this protein was increased by incubation of the cells with transforming growth factor beta or serum. These results identify extracellular factors important for proliferation and differentiation of embryonic human astrocytes and provide a controlled system for multipassage culture.

Astrocytes↗
PubMed1991

Protection of pig epidermis against radiation-induced damage by the infusion of BW12C.

BW12C, which was developed as an agent for the treatment of sickle cell anaemia, increases the binding of oxygen to haemoglobin and hence reduces the availability of oxygen to tissues. Due to these changes in oxygen availability BW12C could act as a protector against radiation-induced injury to normal tissues. In this study the potential value of BW12C, as a radioprotector, was studied in the irradiated epidermis of the pig. The infusion of BW12C caused an instant left shift of the oxygen dissociation curve, an effect that lasted for approximately 1.5 h. This left shift in the oxygen dissociation curves increased with increasing dose of the drug. There appeared to be no long-term systemic effects produced by doses of 20-100 mg/kg of BW12C. In the first 90 min after the infusion of BW12C skin fields were irradiated with single doses of beta-rays from strontium-90 plaques. The incidence of moist desquamation was used as an endpoint for assessing the severity of the radiation response. With animals breathing approximately 70% oxygen in the anaesthetic gas mixture, the ED50 values for moist desquamation were 30-31 Gy after a dose of 30 and 50 mg/kg, and 37-38 Gy for 75 and 100 mg/kg doses of BW12C. These ED50 values were significantly higher than the value of 27.3 Gy for radiation alone. This indicated dose modification factors (DMF) with mean values of approximately 1.13 and approximately 1.40 for irradiation following the infusion of low (30-50 mg/kg) and high (75-100 mg/kg) doses of the drug, respectively. With the animals breathing air (approximately 21% of oxygen) in the 2% halothane anaesthesia gas mixture, irradiation in the presence of 30 and 50 mg/kg of BW12C resulted in ED50 values of approximately 39 Gy for moist desquamation, which was significantly higher than the value of 31.2 Gy for radiation alone. Surprisingly, a higher dose of 75 mg/kg of BW12C resulted in a lower ED50 value for moist desquamation of 34.38 Gy. Irradiation in the presence of a dose of 100 mg/kg of BW12C produced an ED50 value which was not significantly different from that for radiation alone. In the situation where animals were breathing air (approximately 21% oxygen) during irradiation a DMF of 1.14 was obtained for irradiation alone, when the results were compared with those for irradiation alone with approximately 70% oxygen in the anaesthetic gas mixture.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Death of serum-free mouse embryo cells caused by epidermal growth factor deprivation.

Serum-free mouse embryo (SFME) cells, derived in medium in which serum is replaced with growth factors and other supplements, are proastroblasts that are acutely dependent on epidermal growth factor (EGF) for survival. Ultrastructurally, an early change found in SFME cells deprived of EGF was a loss of polysomes which sedimentation analysis confirmed to be a shift from polysomes to monosomes. The ribosomal shift was not accompanied by decreased steady-state level of cytoplasmic actin mRNA examined as an indicator of cellular mRNA level. With time the cells became small and severely degenerate and exhibited nuclear morphology characteristic of apoptosis. Genomic DNA isolated from cultures undergoing EGF deprivation-dependent cell death exhibited a pattern of fragmentation resulting from endonuclease activation characteristic of cells undergoing apoptosis or programmed cell death. Flow cytometric analysis indicated that cultures in the absence of EGF contained almost exclusively G1-phase cells. Some of the phenomena associated with EGF deprivation of SFME cells are similar to those observed upon NGF deprivation of nerve cells in culture, suggesting that these neuroectodermal-derived cell types share common mechanisms of proliferative control involving peptide growth factor-dependent survival.

Animals↗
PubMed1991

The relative biological effectiveness of fractionated doses of fast neutrons (42 MeVd----Be) for normal tissues in the pig. IV. Effects on renal function.

The effects of fractionated doses of fast neutrons (42 MeVd----Be) on the radiation response of the pig kidney have been assessed and compared with those observed after X irradiation. Following X irradiation there was a marked increase in the total dose at which renal function was preserved with decreasing fraction size. The rate of this increase was dependent on the overall treatment time; for fractionated irradiation given over 18 or 39 days the exponents related to fraction number, N, were 0.36 +/- 0.03 and 0.48 +/- 0.003, respectively. In contrast, there was no significant change in the iso-effect dose for renal injury following fractionated irradiation with fast neutrons where there was also little effect of varying the overall treatment time. Analysing these data by means of the linear-quadratic (LQ) model, using both an Fe-plot and the Tucker test, gave alpha/beta ratios of 2.42 +/- 0.06 Gy and 2.99 +/- 0.16 Gy, respectively, for X-ray doses given in 18 days. For fractionated doses of X rays given in 39 days the alpha/beta ratios were 0.40 +/- 0.01 Gy and 0.47 +/- 0.02 Gy, respectively. The alpha/beta ratios for renal tissue following fast neutron irradiation obtained by the two methods were also similar, i.e. 15.00 +/- 0.60 Gy and 15.72 +/- 3.76 Gy, respectively. The pronounced fractionation effect seen with X irradiation, particularly for doses administered over 39 days as opposed to 18 days, coupled with the absence of any such effect with fast neutrons, resulted in a marked increase in relative biological effectiveness (RBE) with decreasing X-ray dose/fraction. The slopes of the resulting regression lines were -0.73 +/- 0.05 and -0.33 +/- 0.02, respectively. The lack of dose sparing associated with fractionation, or variation of the overall treatment time for fast neutron irradiation, suggests that doses administered to tumours adjacent to the kidney can be given as a few relatively large dose/fractions in a short overall treatment time without an increased risk of complications related to renal tissue. This may be of therapeutic advantage in the treatment of rapidly proliferating tumours where dose may be wasted using more conventional protracted fractionated irradiation schedules.

Animals↗
PubMed1990

The role of metabolism in carbon tetrachloride-mediated immunosuppression: in vivo studies.

The role of metabolic bioactivation for carbon tetrachloride-mediated suppression of humoral responses was investigated in B6C3F1 mice. Subchronic studies with CCl4 demonstrated that this chlorinated hydrocarbon markedly suppressed T-dependent antibody responses following 7 consecutive days of administration at doses between 500 and 5000 mg/kg. No significant difference in the magnitude of suppression was observed between the ip and oral routes of exposure. Thirty-day ip administration of CCl4 at doses as low as 25 mg/kg also resulted in a significant inhibition of T-dependent antibody responses. The results from both the 7-day and the 30-day studies indicate that a greater than 50% suppression of antibody responses could not be achieved even at doses of CCl4 as high as 5000 mg/kg. In vivo studies utilized the cytochrome P450 competitive inhibitor, aminoacetonitrile (AAN), in an effort to block the effects of exposure to CCl4. Both the hepatotoxicity, as measured by serum glutamic pyruvic transaminase levels, and the suppression of the T-dependent antibody response to sRBC were reversed by treatment with AAN. Conversely, induction of cytochrome P450, by pretreatment of mice with ethanol prior to treatment with CCl4, resulted in the potentiation of the immunosuppressive effects of CCl4. AAN and ethanol administered alone had no effect on antibody responses. In order to assess the effect of CCl4 treatment on cytochrome P450 activity at doses which cause immunosuppression, measurements of total microsomal protein and specific substrate activities were determined. Significant decreases were observed in both total hepatic microsomal protein as well as in aminopyrine N-demethylase activity, aniline hydroxylase activity, and aryl hydrocarbon hydroxylase activity following treatment with CCl4 for 7 days at doses ranging from 5 to 1000 mg/kg. All of the cytochrome P450 parameters that were measured, following CCl4 treatment, demonstrated very flat dose-response curves which appeared to parallel the effects of CCl4 on antibody responses.

Aminoacetonitrile↗
PubMed1990

Mitogenic activity from trout embryos.

An extract of 21-day rainbow trout embryos stimulated growth of several piscine cell lines in the absence of added serum. Established lines from trout (RTG-2 and STE-137), salmon (CHSE-214), carp (EPC), and goldfish (CAR) and early-passage cells initiated from trout embryos grew in serum-free medium containing the embryo extract. In addition the extract was sufficient for maintaining long-term cultures of CHSE-214 cells for several months through a minimum of 20 passages (approximately 50 population doublings) in the absence of serum. Optimal response was achieved with 100 micrograms of extract protein per ml, but a significant growth-promoting effect was observed with as little as 2.5 micrograms/ml. The activity was nondialyzable, protease-sensitive, and stable in 200 mM acetic acid. The level of mitogenic response induced by the extract could not be duplicated with purified mammalian growth factors added individually or in combination, and the extract did not stimulate DNA synthesis in quiescent mouse fibroblasts. These results suggest that trout embryo extract may contain a novel growth-promoting activity for fish cells.

Animals↗
PubMed1990

The relative biological effectiveness of fractionated doses of fast neutrons (42 MeVd----Be) for normal tissues in the pig. II. Late effects on cutaneous and subcutaneous tissues.

The late effects of irradiation with single and fractionated doses of X rays (250 kV) and fast neutrons (42 MeVd----Bc), on the cutaneous and subcutaneous tissues of the pig, have been evaluated from measurements of changes in relative field length. These were determined at intervals of 26-104 weeks after irradiation. For fractionated irradiation with X rays the average fractions exponent, N, obtained from a log-log plot of iso-effect dose (ED50) against fraction number was 0.41. This was independent of the period of assessment, with no significant indication of a time factor. However, the exponent N did vary with the level of effect and was in the range 0.33-0.51. It was greatest for a greater than or equal to 10% reduction in relative field length. Assuming the validity of the linear quadratic model of cell survival, the alpha/beta ratio was 1.95 Gy. However, this model fitted the data less well for the least severe levels of damage, and for these the alpha/beta ratios were not significantly different from zero. Irradiation with fast neutrons showed a small effect of fractionation for doses given in greater than or equal to 6 fractions compared with a single dose. There was no significant increase in iso-effect dose when the dose was given in 30 fractions compared with 6 fractions. The relative biological effectiveness for late cutaneous and subcutaneous damage for the energy of fast neutrons used did not vary with the period of assessment, i.e. 26-52 weeks compared with 65-104 weeks, and was not significantly different from that previously obtained for ischaemic dermal necrosis, seen after higher doses, at 12-20 weeks after irradiation.

Animals↗
PubMed1990

The relative biological effectiveness of fractionated doses of fast neutrons (42 MeVd----Be) for normal tissues. III. Effects on lung function.

The effect of single and fractionated doses of fast neutrons (42 MeVd----Be) on the early and late radiation responses of the pig lung have been assessed by the measurement of changes in lung function using a 133Xe washout technique. The results obtained for irradiation schedules with fast neutrons have been compared with those after photon irradiation. There was no statistically significant difference between the values for the relative biological effectiveness (RBE) for the early and late radiation response of the lung. The RBE of the neutron beam increased with decreasing size of dose/fraction with an upper limit value of 4.39 +/- 0.94 for infinitely small X-ray doses per fraction.

Animals↗
PubMed1989

Meeting report: human fetal tissue transplantation research panel.

On September 14 through 16, 1988, a meeting on the use of human fetal tissue in transplantation was held at the National Institutes of Health, Bethesda Maryland, USA. The meeting sponsored by NIH for the Human Fetal Tissue Transplantation Research Panel, a consultant group to the Advisory Committee to the Director. The consultant group was convened to deal with the scientific, judicial and moral questions associated with research involving transplantation of human fetal tissue obtained after induced abortions. The first day of the meeting was devoted to presentations addressing scientific issues. Included among the speakers was Dr. Lars Olson, Professor of Neurobiology, Karolinska Institute, Stockholm, who described the use of transplanted human fetal tissue in the treatment of patients with Parkinson's disease and Dr. Eugene Redmond, Professor of Psychiatry, Yale University School of Medicine, who showed results of work with transplantation of tissue to correct induced Parkinson-like disease in monkeys. Other speakers addressed the present, past or potential use of fetal tissue in the treatment of diabetes, immune disorders, and other diseases, as well as the use of fetal cells in the production of biologicals. At the conclusion of the meeting the panel did not recommend that research be halted on fetal tissue within the context discussed, although the recommendation of the committee is not binding, and an additional assembly of the panel will probably occur before the final recommendation to an NIH advisory committee is made in November. Other meetings on this subject include a meeting on the use of fetal tissue sponsored by the American Association of Tissue Banks, March 6-7, 1989, in Washington D. C. (Crystal City) and a meeting June 10, 1989, the day before the annual meeting of the Tissue Culture Association, USA, in Orlando, Florida, on fetal cells and ownership of cultured cells and products derived from clinical specimens. Following are statements to the Human Fetal Tissue Transplantation Research Panel presented September 14, 1988, by Dr. David Barnes, Associate Professor of Biochemistry and Biophysics in the Environmental Health Sciences Center at Oregon State University, USA, who was asked to address for the panel recent advances in cell culture related to fetal tissue, and Dr. Robert E. Stevenson, Director of the American Type Culture Collection, President of the Tissue Culture Association, USA, and Chairman of the Committee on Cells and Tumors of the American Association of Tissue Banks.

Cells, Cultured↗
PubMed1989

Modification of the radiation response of pig skin by manipulation of tissue oxygen tension using anesthetics and administration of BW12C.

The importance of tissue oxygen tension on radiosensitivity was studied by examining modifications in the incidence of moist desquamation in pig skin after irradiation with strontium-90 plaques. The effects were analyzed using quantal dose-response data and comparisons were made using ED50 values for moist desquamation. Under standard anesthetic conditions of 2% halothane, approximately 70% oxygen, and approximately 30% nitrous oxide, the ED50 value (+/- SE) for moist desquamation was 27.32 +/- 0.52 Gy with no significant variation in radiosensitivity between dorsal, lateral, and ventral skin sites on the flank. Irradiation with 2% halothane and air increased the ED50 to 31.25 +/- 0.94 Gy, primarily due to an increased radioresistance of the dorsal sites. When combined with BW12C, a drug which binds oxygen selectively to hemoglobin and hence reduced the oxygen availability to tissues, a further increase in the ED50 values was observed. This was approximately 39 Gy with BW12C concentrations of 30 mg/kg and 50 mg/kg b.w. of BW12C, indicating a dose modification factor (DMF) of approximately 1.26. However, when animals were breathing the standard gas mixture, this DMF was reduced to 1.15 for 30 mg/kg of BW12C, indicating that a higher level of oxygen partly counteracted the effects of the drug in these studies with BW12C. The greatest variability in radiosensitivity was seen in the dorsal fields. This suggested complex physiological adaptation, a phenomenon that might also explain the absence of any modification of the radiation response when 100 mg/kg of BW12C was used.

Aldehydes↗
PubMed1988

Attachment and multiplication, morphology and protein production of human fetal primary liver cells cultured in hormonally defined media.

We established for human fetal liver cells (cultured for 2 wk) in a hormonally defined medium, optimal conditions for attachment, multiplication, and preservation of epithelial morphology as well as production and secretion of serum proteins characteristic of fetal (alpha l-fetoprotein, AFP) and adult (albumin and hemopexin) life. Conditions were considered optimal when cell number, albumin, and hemopexin levels were maintained throughout the 2-wk culture period. However, the decrease in AFP concentration, which occurred after a few days of culture, could not be reversed. The culture system developed is a suitable model for studying regulatory mechanisms governing structure and function during differentiation and may prove useful for testing the effect of toxic agents during fetal development of the human liver.

Cell Adhesion↗
PubMed1988

An immunotoxicological evaluation of 4,4'-thiobis-(6-t-butyl-m-cresol) in female B6C3F1 mice. 1. Body and organ weights, hematology, serum chemistries, bone marrow cellularity, and hepatic microsomal parameters.

Adult female B6C3F1 mice were gavaged with 4,4'-thiobis-(6-t-butyl-m-cresol) (TBBC) in corn oil at doses of 10, 100, or 200 mg/kg daily for 14 consecutive days. There was no overt toxicity, as manifested by grossly observable behavioral changes, decreased growth rate over the exposure period, or mortality. There were also no marked effects on serum chemistries or hematology, with the exception of a significant increase (41%) in the number of leukocytes at the highest dose. Absolute differential counts indicated that significant increases occurred in the number of lymphocytes (31%) and neutrophils (177%). Studies with bone marrow indicated a significant 30% increase in the number of cells/femur from animals treated with the highest dose of TBBC. The number of macrophage progenitors (CFU-M)/femur was significantly increased by 28%, while the number of granulocyte-monocyte progenitors (CFU-GM)/femur was nonsignificantly increased by 20% in the high dose animals. The weight of both the spleen and liver was increased in a dose-related fashion, although the histopathology of the spleen of TBBC-treated mice was not different from control. The livers of mice receiving the high dose showed mild focal hydropic degeneration, mild hepatitis, and a slight increase in the number of Kupffer cells. No other organs were affected. Liver microsomal protein and cytochrome P-450 levels were increased in a dose-related fashion. Enzyme activities of aminopyrine demethylase and aniline hydroxylase, but not arylhydrocarbon hydroxylase, were also increased in a dose-related fashion.

Animals↗
PubMed1988

The relative biological effectiveness of fractionated doses of fast neutrons (42 MeVd----Be) for normal tissues in the pig. I. Effects on the epidermis and dermal vascular/connective tissues.

The effects of fractionated doses of fast neutrons (42 MeVd----Be) on the early epithelial and later dermal response of pig skin have been assessed and compared with those after X irradiation. For the early epithelial reaction, i.e. moist desquamation, the relative biological effectiveness (RBE) of the neutron beam increased with the decreasing size of the X-ray dose/fraction. There was an experimentally observed upper RBE value of approximately 2.75 for X-ray doses/fraction of between 2 and 5 Gy. For the late reaction of ischaemic dermal necrosis the RBE was greater than 3.0 for X-ray doses/fraction of less than 3 Gy and, based on the assumptions made in the linearquadratic model of cell survival, an upper limiting RBE of 4.32 +/- 0.39 was calculated for infinitely small doses/fraction. These findings were compared with other radiobiological data and the conclusions drawn from the results of clinical trials. It was concluded that for the sparing of late effects in skin and subcutaneous tissues, relative to acute reactions, a relatively small number of fractions in a short overall treatment time may be optimal for fast neutron therapy.

Animals↗