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D M Sullivan

Publications and source records attributed to D M Sullivan.

At least 73 records · Page 4Linked to original sources
PubMed1987

Proliferation-dependent topoisomerase II content as a determinant of antineoplastic drug action in human, mouse, and Chinese hamster ovary cells.

We have shown previously that quiescent Chinese hamster ovary (CHO) cells are less sensitive than log phase CHO cells to the cytotoxic and DNA cleavage effects of etoposide, a drug which appears to act via DNA topoisomerase II. This loss of sensitivity was associated with a decrease in topoisomerase enzyme activity in nuclear extracts of the quiescent cells. We have now extended our observations by examining the basis for the reduction in enzyme activity during quiescence. DNA topoisomerase II content, as assayed by immunoblotting with a polyclonal rabbit anti-topoisomerase II antiserum, was virtually absent in nuclear extracts of quiescent CHO cells in contrast to logarithmically growing cells. This suggests that the previously demonstrated loss of enzyme activity in CHO cells is a function of reduction in content rather than posttranslational modifications of the enzyme. Quiescent human lymphoblastic CCRF cells also exhibited reduced topoisomerase II content compared to actively proliferating cultures, but the difference was less than that observed in CHO cells. In contrast, log and plateau phase cultures of mouse leukemia L1210 cells exhibited similar topoisomerase II content. Reduction in enzyme content correlated with the ability of these cell lines to accumulate during quiescence with a G0-G1 content of DNA. Sensitivity to the DNA cleavage effects of etoposide in dividing and nondividing cells correlated well with enzyme content. As has been observed with CHO cells, both CCRF and L1210 cells in plateau phase were more resistant to the cytotoxic effects of etoposide than those actively dividing. The result with L1210 cells was surprising, however, in light of the equivalent DNA damage observed under the two growth conditions. Our data indicate that topoisomerase II enzyme content is proliferation dependent in some but not all cells and suggest that while enzyme content may be important in drug resistance in some cell types, other factors can decrease the sensitivity of the cell to cleavable complex formation as well.

Animals↗
PubMed1987

Role of proliferation in determining sensitivity to topoisomerase II-active chemotherapy agents.

We have examined the relationship between topoisomerase II content and the DNA cleavage activity and cytotoxicity of etoposide during proliferative and quiescent culture conditions. In proliferating cultures of Chinese hamster ovary (CHO) cells, human lymphoblastic CCRF cells, and mouse leukemia L1210 cells, there was easily detectable topoisomerase II by immunoblotting. In contrast, quiescent CHO cells contained virtually no detectable topoisomerase II, while the content of L1210 cells was unchanged. Enzyme content of quiescent CCRF cells was diminished but detectable. DNA cleavage activity induced by etoposide correlated well with enzyme content in proliferating and quiescent cells. Quiescent CHO and CCRF cultures were highly resistant to the cytotoxic effects of etoposide as expected. However, despite unchanged enzyme content and DNA cleavage activity, there was also significant resistance observed in plateau L1210 cells. We have also investigated topoisomerase content and drug activity as a function of cell cycle progression. Following serum stimulation of confluent BalbC/3T3 cells, maximal etoposide-induced DNA cleavage activity is observed in G2/M and is associated with an increase in topoisomerase II content. Maximum cytotoxicity, however, occurs during mid to late S phase. Our data suggest that topoisomerase II content may be an important determinant of chemotherapeutic sensitivity during alterations in the proliferative status of the cell. However, it is clear that other factors must be involved in cell sensitivity, and elucidation of these may contribute to our understanding of the mechanism of action of these drugs.

Animals↗
PubMed1987

Mediation of multi-drug resistance in a Chinese hamster ovary cell line by a mutant type II topoisomerase.

Identification of DNA topoisomerase II as the intracellular target for the DNA cleavage activity of the epipodophyllotoxins and several intercalating agents is well established. In contrast, definite correlation of cleavable complex formation with eventual cell death has been more difficult to document. Our studies with an epipodophyllotoxin resistant cell line not only provide additional evidence that the enzyme is a multi-drug target, but also implicate drug-stimulated cleavage activity as a critical component of cytotoxic effect. When compared to WT (wild type) cells, the mutant Chinese hamster ovary cell line, VpmR-5, exhibits marked resistance to both the cytotoxic and DNA cleavage activity of etoposide as well as various intercalating agents. Steady state concentrations of these drugs in both cell lines is identical. Enzyme content as measured by immunoblot and by total decatenation activity in crude nuclear extracts is equal. Catalytic activity is also equally sensitive to inhibition by etoposide. In contrast, cleavage activity in purified enzyme from VpmR-5 cells is profoundly resistant to stimulation by drug. These data indicate that a multi-drug resistant phenotype may be acquired by a qualitative change in the enzyme that alters its interaction with drug. Further, the data strongly support a direct role for cleavable complex formation in cell death.

Amsacrine↗
PubMed1986

Proliferation dependence of topoisomerase II mediated drug action.

Topoisomerase II mediated DNA scission induced by both a nonintercalating agent [4'-demethylepipodophyllotoxin 4-(4,6-O-ethylidene-beta-D-glucopyranoside) (VP-16)] and an intercalator [4'-(9-acridinylamino) methanesulfon-m-anisidide (m-AMSA)] was studied as a function of proliferation in Chinese hamster ovary (CHO), HeLa, and mouse leukemia L1210 cell lines. Log-phase CHO cells exhibited dose-dependent drug-induced DNA breaks, while plateau cells were found to be resistant to the effects of VP-16 and m-AMSA. Neither decreased viability nor altered drug uptake accounted for the drug resistance of these confluent cells. In contrast to CHO cells, plateau-phase HeLa and L1210 cells remained sensitive to VP-16 and m-AMSA. Recovery of drug sensitivity by plateau-phase CHO cells was found to reach a maximum approximately 18 h after these cells regained exponential growth and was independent of DNA synthesis. DNA strand break frequency correlated with cytotoxicity in CHO cells; log cells demonstrated an inverse log linear relationship between drug dose (or DNA damage) and colony survival, whereas plateau-derived colony survival was virtually unaffected by increasing drug dose. Topoisomerase II activity, whether determined by decatenation of kinetoplast DNA, by cleavage of pBR322 DNA, or by precipitation of the DNA-topoisomerase II complex, was uniformly severalfold greater in log-phase CHO cells compared to plateau-phase cells.

Aminoacridines↗
PubMed1984

Intracranial vessels lack vasa vasorum.

A micro-corrosion technique was used to demonstrate an extensive vasa vasorum network in extracranial vessels but did not reveal this system in intracranial vessels of comparable size in three species of animals. The absence of a vasa vasorum network in cerebral vessels may result in a higher level of susceptibility to periarterial abnormalities, such as cerebral vasospasm secondary to subarachnoid hemorrhage.

Animals↗
PubMed1983

Fractionation and kinetic properties of rat liver and kidney methionine adenosyltransferase isozymes.

Three isozymes of methionine adenosyltransferase (EC 2.5.1.6; MAT-I, -II, and -III) exist in normal rat liver and are conveniently purified (MAT-III to homogeneity) by a three-step column chromatography procedure. MAT-I shows Michaelis-Menten kinetics with a Km (L-methionine) of 41 microM and a molecular weight of 208 000 and is slightly inhibited by S-adenosyl-L-methionine (Adomet). MAT-II, which is also the only isozyme found in normal rat kidney, shows negative cooperativity with a Hill coefficient of 0.7. It has a L-methionine concentration required for half-maximal velocity [S0.5(Met)] of 8 microM and a molecular weight of 120 000 and is strongly inhibited by Adomet. MAT-I and -II comprise 15% and 5%, respectively, of total MAT activity in rat liver. The predominant isozyme in rat liver, MAT-III, demonstrates positive cooperativity with a Hill coefficient of 1.8. It has a molecular weight of 97 000 and apparently consists of two subunits of identical molecular weight (47 000). This liver-specific isozyme is strongly activated by both dimethyl sulfoxide and Adomet and has a S0.5(Met) of 215 microM.

Animals↗
PubMed1982

Structural and functional properties of a Ca2+-ATPase from human platelets.

An antibody prepared against highly purified rabbit muscle Ca2+-ATPase from sarcoplasmic reticulum has been observed to cross-react with proteins in human platelet membrane vesicles. The antibody specifically precipitated Ca2+-ATPase activity from solubilized human platelet membranes and recognized two platelet polypeptides denatured in sodium dodecyl sulfate with Mr = 107,000 and 101,000. Ca2+-ATPase activity from Brij 78-solubilized platelet membranes was purified up to 10-fold. The purified preparation consisted mainly of two polypeptides with Mr approximately 100,000, and 40,000. The lower molecular weight protein appeared unrelated to Ca2+-ATPase activity. The Ca2+-ATPase in human platelet membrane vesicles exhibited "negative cooperativity" with respect to the kinetics of ATP hydrolysis. The apparent Km for Ca2+ activation of ATPase activity was 0.1 microM. Ca2+-dependent phosphorylation of platelet vesicles by [gamma-32P]ATP at 0 degrees C yielded a maximum of 0.2-0.4 nmol of PO4/mg of protein that was labile at pH 7.0 and 20 degrees C. This result suggests that only about 2-4% of the total protein in platelet membrane vesicles is the Ca2+-ATPase, which agrees with an estimate based on the specific activity of the Ca2+-ATPase in platelet membranes (20-50 nmol of ATP hydrolyzed/min/mg of protein at 30 degrees C). Calmodulin resulted in only a 1.6-fold stimulation of Ca2+-ATPase activity even after extensive washing of membranes with a calcium chelator or chlorpromazine. It is concluded that human platelets contain a Ca2+-ATPase immunochemically related to the Ca2+ pump from rabbit sarcoplasmic reticulum and that the enzymatic characteristics and molecular weight of the platelet ATPase are quite similar to those of the muscle ATPase.

Adenosine Triphosphate↗
PubMedDate not supplied

Seven years of biosolids versus inorganic nitrogen applications to tall fescue.

Repeated applications of municipal wastewater biosolids is cost effective for biosolids managers, but may lead to undesirable accumulations of nutrients or contaminants. We evaluated the effects of seven years of biosolids applications on tall fescue (Festuca arundinacea Schreb.) production and nutrient availability. We compared two types of Class A biosolids applied to tall fescue on a sandy loam in western Washington. Mean annual biosolids rates of 290, 580, and 870 kg total N ha(-1) yr(-1) were compared with inorganic N and zero-N controls using a randomized complete block design. We measured yield and N uptake for each forage harvest, plant tissue metals at selected harvests, soil nitrate each fall, diethylenetriaminepentaacetic acid (DTPA)-extractable metals after five years of applications, and soil pH, available P, and organic C after seven years. Forage yields increased with biosolids rate. Apparent nitrogen recovery (ANR) for biosolids averaged 18% in 1993 (Year 1), 35% in 1994, and 46% in 1999. The ANR for inorganic N averaged 62% from 1994-1999. Residual soil nitrate was less than 25 kg ha(-1) for all treatments through 1995, but increased beginning in 1996 for the high biosolids rate. Biosolids increased soil organic C levels by 2 to 5 g kg(-1) and Bray-1 P levels by 300 to 600 mg kg(-1) (0-15 cm depth). Plant tissue Zn increased from 24 to 66 mg kg(-1) at the highest application rate. Nearly all of the DTPA-extractable metals remained in the 0- to 8-cm soil depth.

Chelating Agents↗
PubMedDate not supplied

Potential hazards of radiative electromagnetic hyperthermia in the presence of multiple metallic surgical clips.

Three-dimensional computer simulations of a patient under consideration for heating by the BSD Sigma 60 phased array applicator showed marked specific absorption rate (SAR) enhancement in the region where there was a concentration of metallic surgical clips. Subsequent tests in tissue-equivalent phantom material confirmed that metallic objects much shorter than the wavelength in tissue of the radiative electromagnetic field can cause significant changes in the power deposition near the implanted metal. The placement of extensive numbers of surgical clips may therefore hinder the ability to subsequently utilize radiative electromagnetic heating modalities.

Adult↗
PubMedDate not supplied

Peripheral neuropathy and myonecrosis following hyperthermia and radiation therapy for recurrent prostatic cancer: correlation of damage with predicted SAR pattern.

During the past 10 years, numerous phase I-II studies were conducted and provided clinical experience with combined radiation therapy and hyperthermia treatments. Among the rare complications reported in these combined radiation therapy-hyperthermia trials were myonecrosis and peripheral neuropathy which were felt, mainly on a clinical basis, to be caused by local heat damage. Recently, such complications were noted in two patients with recurrent prostatic cancer treated in our department with radiation therapy combined with deep regional hyperthermia delivered by the Sigma-60 applicator of the BSD 2000 hyperthermia system (Salt Lake City, UT, USA). Analysis of the results of three-dimensional modelling of the SAR (specific absorption rate, W/kg) pattern in these patients indicated high SAR at the sites of the complications. Pretreatment three-dimensional modelling or other methods of predicting potential areas of high power deposition may have a role for future hyperthermia treatment planning aiding in the prevention of possible local heat damage and providing improved delivery of heat to the target volume.

Aged↗
PubMedDate not supplied

Stanford 3D hyperthermia treatment planning system. Technical review and clinical summary.

In the field of deep regional hyperthermia cancer therapy the Sigma 60 applicator of the BSD-2000 Hyperthermia System is one of the most widely used devices. This device employs four independent sources of radiofrequency electromagnetic energy to heat tumour sites deep within the body. The difficulty in determining the input parameters for the four sources has motivated the development of a computer-based three-dimensional (3D) treatment planning system. The Stanford 3D Hyperthermia Treatment Planning System has been in clinical use at Stanford Medical Center for the past 2 years. It utilizes a patient-specific, three-dimensional computer simulation to determine safe and effective power deposition plans. An optimization programme for the selection of the amplitudes, phases and frequency for the sources has been developed and used in the clinic. Examples of the application of the treatment planning for hyperthermia treatment of pulmonary, pelvic, and mediastinal tumours are presented. Methods for quantifying the relative effectiveness of various treatment plans are reviewed.

Adult↗
PubMedDate not supplied

Determination of gold in animal tissue by graphite furnace atomic absorption spectrophotometry.

Historically, compounds that contain gold have been used to treat conditions such as rheumatoid arthritis in humans. However, understanding of the metabolic fate of gold in biological tissues has been limited by lack of sensitive quantitative methods of analysis. We addressed this problem by developing a graphite furnace atomic absorption (GFAA) spectrophotometric method to measure trace amounts of gold. This method was validated on small samples of beef liver, kidney, and bone. The samples were digested in micro-Kjeldahl flasks with a mixture of sulfuric, perchloric, and nitric acids; the residue was treated with aqua regia and extracted into methylisobutyl ketone (MIBK); levels of gold were then measured by GFAA. All the reagents were of an ultra-pure grade and were monitored for gold content. We established that the linear range of quantitation was from 1 to 2500 ppb. Multiple extractions with MIBK were not necessary to recover all the gold, and, in most cases, use of ultra-pure acids was not necessary. A scan of the extracts by inductively coupled argon plasma atomic emission spectrophotometry revealed no appreciable concentration of elements that would be most likely to interfere with the determination of gold. Average recoveries of gold ranged from 102 to 111%, and the overall coefficient of variation was 5.5%.

Animals↗