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Biomedical subjects

R A Floyd

Publications and source records attributed to R A Floyd.

At least 127 records · Page 7Linked to original sources
PubMed1986

Effect of aging on pancreatic lipolytic enzymes.

Pancreatic lipolytic enzyme activities and plasma lipids were measured in three age groups of female Fisher 344 rats (3 months [young], 12 months [adult], and 27 months [old]) in order to evaluate age-related changes and a possible correlation of these parameters. Cholesterol esterase activity was measured in pancreas homogenate, while the lipase activity was further fractionated into heparin-Sepharose unretained (lipase I) and retained (lipase II) fractions. In analogy to the lipolytic enzymes of the human pancreas, lipase I corresponds to pancreatic lipase and lipase II corresponds to pancreatic carboxylesterase. Plasma triglyceride and cholesterol levels were significantly higher in old as compared with adult and young rats. There was no significant correlation between plasma lipid and pancreatic enzyme activity levels. Cholesterol esterase and pancreatic lipase (lipase I) activity did not show any consistent change with age. Pancreatic carboxylesterase (lipase II), on the other hand, was consistently lower in adult and old animals. Although the importance of pancreatic carboxylesterase for triglyceride hydrolysis remains to be established, our results suggest that this enzyme is under long-term metabolic control. We suggest that early in life, high levels of this enzyme may be needed to supplement pancreatic lipase in order to optimize digestion of dietary fat required for optimal growth.

Age Factors↗
PubMed1986

Hydroxyl free radical adduct of deoxyguanosine: sensitive detection and mechanisms of formation.

DNA or 2-deoxyguanosine reacts with hydroxyl free radical to form 8-hydroxy-deoxyguanosine (8-OH-dG). We found that 8-OH-dG can be effectively separated from deoxyguanosine by high pressure liquid chromatography and very sensitively detected using electrochemical detection. The sensitivity of electrochemical detection is about one-thousand fold enhanced over optical detection. Utilizing deoxyguanosine in bicarbonate buffer it was found that ferrous ion, but not ferric ion, was effective in forming 8-OH-dG. The hydroxyl free radical scavenging agents, thiourea and ethanol, were very effective in quenching Fe(11) mediated 8-OH-dG formation, but superoxide dismutase had very little effect.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1985

Studies on the effect of iron overload on rat cortex synaptosomal membranes.

Iron as ferrous ammonium sulfate was injected into the cerebral spinal fluid of rats. After three consecutive days of injection of 4 mumol of iron, the total iron content of brain cortex synaptosomes from the iron-treated animals was 2-fold higher than that from control animals receiving the saline vehicle only. Spin label studies of the synaptosomal membranes demonstrated that the lipid region of the membranes became more rigid and, in addition, the mobility of labeled SH groups of membrane proteins decreased after the iron treatment. The cholesterol content was significantly higher in iron-treated animals as compared to controls. Centrophenoxine pretreatment (100 mg/kg body weight daily for 6 weeks) diminished the iron effects. Synaptosomal membrane alterations observed after iron treatment were similar to changes observed previously during aging. This lends support to the notion that free-radical induced damage occurs in brain membranes with increasing age.

Animals↗
PubMed1985

Regional lipid peroxidation in rat brain in vitro: possible role of endogenous iron.

Lipoperoxidative capacity of various brain areas of aging rats was examined in vitro using the thiobarbituric acid test. Significant regional differences in the generation of lipid peroxides were found in freshly prepared homogenates from different areas of brain incubated under air. Incubation under oxygen resulted in marked stimulation of lipid peroxidation, with highest increases in hypothalamus (144%). Addition of exogenous Fe2+ and ascorbic acid resulted in stimulation of lipid peroxidation ranging from 10-fold in cortex to 20-fold in hypothalamus homogenates during incubation in air. A linear relationship was found between endogenous iron content in brain regions and their ability to produce lipid peroxides in vitro under oxygen for all areas except striatum. Several iron chelating agents effectively inhibited lipid peroxidation under hyperbaric oxygen whereas oxygen-free radical scavengers, as well as catalase and superoxide dismutase were not effective. It is concluded that regional differences in lipoperoxidative capacity of brain areas in vitro are in part governed by local endogenous iron content and may indicate regional susceptibility to oxidative damage.

Animals↗
PubMed1985

Phase I and pharmacologic study of oral ftorafur and X ray therapy in advanced gastrointestinal cancer.

We have treated 15 patients with advanced gastrointestinal carcinoma with a cyclical regimen of combined Ftorafur (N1-((2-furanidyl-))-5-Fluorouracil, a 5-FU pro-drug) and external beam radiation. The Ftorafur (FT) was administered orally in daily doses of between 1.0 and 2.5 g/m2/day in 3 divided doses in a Phase I format. The drug was given daily for 5 days along with conventional X ray treatment portals and daily radiation doses of 250 rad on each of the first 4 days of each treatment cycle. The patients were then rested for a minimum of 10 days or until all significant side effects had passed. The total number of 1,000 rad cycles and radiation dose were dictated by tolerance and by normal organ dose limitations. The most common toxicity in general, and the most common limiting toxicity was nausea and vomiting, in contrast to oral FT alone where diarrhea is more prominent. Stomatitis was seen only once and no other form of serious toxicity was encountered. Two-thirds of the patients responded in subjective terms (pain relief). There was 1 partial response to FT alone (pulmonary metastases outside the treatment field). The sole patient whose treatment field was outside the abdomen (chest portals for esophageal carcinoma) developed pneumonitis which contributed to his death. No other delayed effects were noted. Serum FT levels were related to the ingested dose and in the microgram range while serum 5-FU levels were in the nanogram range indicating slow decomposition of FT into 5-FU. The therapy was reasonably well tolerated at doses of 2.0 g/m2/day or lower with abdominal radiation. FT offers the potential for replacing intra-venous infused 5-FU as a clinical radiosensitizer.

Administration, Oral↗
PubMed1985

Phase I and pharmacologic study of 72-hour infused 5-fluorouracil in man.

The relationships between the administered dose, clearance, and the toxicity spectrum of 5-fluorouracil (5-FU) administered as 72-hour constant infusion have been studied in 21 patients with advanced cancer. This was done as a pilot study for possible future combination using 5-FU as a radiosensitizer. Individual patients tolerated up to 65 mg/kg/24 hours, but serious toxicity appeared once as low as 35 mg/kg. Limiting toxicity proved to be "mixed" with upper intestinal symptoms (nausea and vomiting), stomatitis, and central nervous system signs all occurring in various patients. Central nervous system effects (both cerebellar and vomiting) proved as troublesome as stomatitis. There was only a general link between the administered dose and the subsequent toxicity grade, but a reasonably quantitative relationship emerged when the serum 5-Fu levels obtained and the degree of patient toxicity were compared. The clearance of 5-FU was confirmed to be nonlinear over the entire dose range studied (25-65 mg/kg/24 hours), consistent with a two-compartment model of drug metabolism. One compartment appears to be systemic (extra-hepatic) metabolism (probably anabolic removal) which is saturated at just below 15 mg/kg/day. Doses above that level lead to drug accumulation. No steady state was reached, contrary to previous reports. At the higher infusion rates, clearance progressively approaches that predicted by the assumption that the second compartment is splanchnic blood flow and catabolism. While 5-FU can be administered as a 72-hour infusion as one possible schedule for use as a single agent or for combined modality studies, CNS effects are quite troublesome in comparison to longer infusions to toxicity.

Drug Evaluation↗
PubMed1984

Hydroxyl free radical reactions with amino acids and proteins studied by electron spin resonance spectroscopy and spin-trapping.

It has recently been shown that Fe(I) complexes of ADP or ATP generate OH radicals with H2O2 in a Fenton-type reaction. The OH radicals can be detected by using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap in electron spin resonance spectroscopy. All the biologically occurring amino acids, some related compounds and several proteins (histone, bovine serum albumin, collagen) were tested as OH radical scavengers against DMPO. The tested compounds competed with DMPO in trapping OH radicals to various extents as shown by the decrease of signal intensity of DMPO-OH spin-adduct. The tested compounds did not oxidize Fe(II) itself, with the only exception being tyrosine, as revealed by properly designed ferrozine reaction. Some of the amino acids reacted also with the DMPO-OH spin-adduct to a certain extent, whereas others did not. The formation of carbon centered organic radicals of the amino acids could be detected under the influence of OH radicals by using the spin traps phenyl-tert-butylnitrone (PBN) and alpha-pyridyl-1-oxide-N-tert-butylnitrone (4-POBN). The proteins, however, did not react with these spin traps. One can conclude that the amino acids and proteins can be targets of OH radical damage even in vivo, and such phenomena may be of importance in the deterioration of the conformation of proteins, e.g., during aging or in some pathological processes.

Amino Acids↗
PubMed1984

Formation of long-lived hydroxyl free radical adducts of proline and hydroxyproline in a Fenton reaction.

Proline and hydroxyproline when exposed to the hydroxyl free radical generating system of ADP-Fe(II)-H2O2 yielded long-lived free radicals. An analysis of the electron paramagnetic resonance spectra of the long-lived hydroxyl free radical adducts of proline and hydroxyproline is consistent with a free electron on a nitroxyl group interacting with the nitrogen atom as well as with three separate protons. In the case of proline, nitroxide formation was observed under the influence of tert-butyl-hydroperoxide, giving a similar EPR spectrum (Lin, J.S., Tom, T.C. and Olcott, H.S. (1974) J. Agr. Food Chem. 22, 526-528); however, the hydroxyl free radical adduct of hydroxyproline has not been described yet. In the case of the proline nitroxide radical, two of the three protons involved interact with the free electron equivalently. The coupling constants for the hydroxyl free radical adduct of proline are AN = 1.58 mT, AH1 beta = AH2 beta = 2.13 mT, AH3 beta = 1.77 mT and for hydroxyproline are AN = 1.54 mT, AH1 beta = 2.56 mT, AH2 beta = 2.03 and AH3 beta = 1.51. The data are consistent with the amine nitrogen of proline and hydroxyproline being oxidized to a nitroxyl group and the free electron of the nitroxyl interacting with the beta-protons of these amino acid hydroxyl free radical adducts.

Adenosine Diphosphate↗
PubMed1984

Sensitive assay of hydroxyl free radical formation utilizing high pressure liquid chromatography with electrochemical detection of phenol and salicylate hydroxylation products.

Evidence is presented for a sensitive method useful for the detection of hydroxyl free radical generation in various systems. The methodology employs high pressure liquid chromatography with electrochemical detection (LCED) for the quantification and identification of the hydroxylation products from the reaction of OH with both phenol and salicylate. A detection limit of less than 1 pmol for the hydroxylation products has been achieved with electrochemical detector responses linear over at least three orders of magnitude. Detection and quantitation of the hydroxylation products obtained and formed during OH generation from biologically meaningful systems have been demonstrated. The three systems utilized were ADP/FE(II)/H2O/, hypoxanthine/xanthine oxidase plus chelated iron, and UV photolysis of H2O2.

Chromatography, High Pressure Liquid↗
PubMed1984

Electron spin resonance spectroscopic demonstration of the hydroxyl free radical scavenger properties of dimethylaminoethanol in spin trapping experiments confirming the molecular basis for the biological effects of centrophenoxine.

The ADP-Fe(II)-H2O2 system generates OH free radicals which can be trapped by 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) thus yielding a measurable signal by electron spin resonance spectroscopy. The amount of DMPO-OH spin adduct formed under certain conditions decreased considerably, if dimethylaminoethanol (DMAE), p-chlorophenoxyacetic acid (PCPA) or centrophenoxine (CPH) were present in comparable concentrations to that of DMPO. It has been demonstrated that such an effect cannot be attributed to any interference of the tested compounds with the Fe(II) and its oxidability by H2O2. The reaction of DMAE with OH free radicals was demonstrated also by using other spin traps. These spin traps reacted with OH free radicals either not at all (phenyl-tert-butylnitrone, PBN) or only to a slight extent (alfa-pyridyl-l-oxide-N-tert-butylnitrone, 4-POBN). DMAE was also a competitive OH free radical scavenger with proline and hydroxyproline, both of which have recently been shown to react with OH free radicals to form nitroxyl free radicals. On the basis of the experimental results, the OH free radical scavenger property of DMAE can be regarded as firmly established. This result supports the molecular mechanism proposed for the explanation of the anti-aging effects of CPH in terms of the membrane hypothesis of aging.

2,4-Dichlorophenoxyacetic Acid↗
PubMed1984

EEG spectra following phenytoin infusion in seizure patients.

Diphenylhydantoin (DPH) kinetics were studied in five male patients with poorly controlled seizures. EEG recordings were taken on four of the patients immediately prior to and 30 minutes following a 60 minute i.v. infusion of phenytoin sodium. Mean phenytoin levels following post-infusion EEG recordings were 17.1 mg/L. Spectral analysis revealed that phenytoin produced a decrease in the variance of EEG spectral power as a function of time in frequencies between 8-18 Hz. This decrease in the variance of the time series of EEG spectra suggests increased EEG stability and may represent an electrographic marker of the anticonvulsant action of phenytoin.

Adult↗
PubMed1984

Protease inhibition of natural killer (NK) and antibody-dependent cell-mediated cytoxicity (ADCC) activities.

The ability of protease inhibitors to modulate NK and ADCC cell activities was studied. NK and ADCC cell activities were suppressed by 1,10 phenanthroline (PHE), N-alpha-p-tosyl-L-lysine chloromethyl ketone (TLCK), L-1-tosylamide-2-phenylethylchoromethyl ketone (TPCK), N-benzoyl-DL-tyrosine ethyl ester (BTEE), and phenylethylchoromethyl fluoride (PMSF). The effect of these protease inhibitors on ADCC was not mediated through the early phase of cytotoxicity involving Fc-receptor binding, since they failed to alter EoxA rosetting of effector cells. The results suggest that an activated esterase system might play a role in the later phase of the cytotoxic mechanism by ADCC cells and perhaps NK cells.

Animals↗
PubMed1983

Hydroxyl free-radical spin-adduct in rat brain synaptosomes. Observations on the reduction of the nitroxide.

Understanding the prevalence and action of hydroxyl free-radicals, damaging species in biological systems, is improved by spin-trapping techniques. The hydroxyl free-radical rapidly adds to the spin-trap 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) to form a relatively stable nitroxyl free-radical spin-adduct (DMPO-OH), but it was found that DMPO-OH becomes diamagnetic, presumably by chemical reduction, when it is added to rat brain synaptosomes. DMPO-OH reduction by synaptosomes was a time-dependent process, starting immediately and continuing up to 20 min or more, when almost all the spin-adduct is reduced. During synaptosomal reduction, the electron paramagnetic resonance (EPR) high field line of DMPO-OH is broadened indicating restrictions on the motion of the nitroxide. This was not due to simple absorption of DMPO-OH to proteins. Experiments with the spin-adduct in glycerol at low temperatures showed that DMPO-OH experienced, during synaptosomal reduction, restricted motion having a correlation time of about 3.3 X 10(-9)s. This value would be expected if DMPO-OH was near the polar region of the hydrocarbon layer of a membrane bilayer. Synaptosomal membrane transport of DMPO-OH was tested by competition studies, since it was presumed that this nitroxide was transported and then reduced inside the synaptosomes. Several compounds tested, including amino acids, neurotransmitters, food dyes, and many others, that may have acted competitively in transport were not effective in slowing down reduction of DMPO-OH. However, it was found that azide, cyanide and heating of the synaptosomes markedly decreased the rate of synaptosomal reduction of DMPO-OH. Azide at 50 microM manifested one-half of its maximal inhibitory effect. Experiments with 1-octanol to obtain a measure of the membrane partition of DMPO-OH demonstrated that the octanol/water distribution was 1.83. Experiments with glass microfibre filtration to determine synaptosomal binding revealed that DMPO, as well as apparently DMPO-OH, were not specifically retained by synaptosomes. All of these results combined with past work on nitroxide reduction by biological systems, suggest that DMPO-OH reduction by synaptosomes is due to its interaction with and reduction by electron transport carriers of mitochondria within the synaptosomes.

Animals↗
PubMed1983

Direct demonstration that ferrous ion complexes of di- and triphosphate nucleotides catalyze hydroxyl free radical formation from hydrogen peroxide.

Utilizing an electron paramagnetic resonance (EPR) spin-trapping technique it was demonstrated that the di- and triphosphate nucleotides of adenosine, cytidine, thymidine, and guanosine in the presence of Fe(II) catalyze hydroxyl free radical formation from H2O2. The triphosphate nucleotides in general were about 20% more effective than the diphosphate nucleotides. The amount of OH produced from H2O2 as a function of nucleotide level tended to increase in a sigmoidal fashion beginning at a nucleotide/Fe(II) ratio of 2 but then rose rapidly up to a ratio of 5 at which point the increase became more gradual. The monophosphate nucleotides did not cause an increase in the amount of hydroxyl free radical produced from H2O2 over the low level obtained in the buffer system only. The cations, Mg2+ and Ca2+, even at much higher than physiological levels and much higher than the level of added Fe(II), did not cause a substantial diminution of the Fe(II)-nucleotide-catalyzed breakdown of H2O2 to yield OH. A study of the time course of the effectiveness of Fe(II)-nucleotide-mediated OH formation from H2O2 demonstrated that Fe(II) in the presence of nucleotides remained in an effective catalytic state with a halftime of about 160 s whereas in the absence of the nucleotides the halftime was 7.5 s. All observations indicate that Fe(II) ligates with di- and triphosphate nucleotides and remains in the ferrous state which is then capable of catalyzing OH formation from H2O2; but with time, oxidation of the metal ion to the ferric state occurs, which either ligated to the nucleotide or to buffer ions, is ineffective in H2O2 catalysis to yield OH. Iron-nucleotide complexes may be of importance in mediating oxygen free radical damage to biological systems. The observations presented here indicate that hydroxyl free radicals will be produced when H2O2 is present with ferrous-nucleotide complexes.

Adenosine Diphosphate↗
PubMed1983

Circulating immune complexes in rats with metastasizing or nonmetastasizing mammary tumor.

The dual rat metastatic and nonmetastatic mammary tumor model was characterized by the level of immune complexes in the sera of rats bearing these tumors. Circulating immune complexes (CIC) were measured at various intervals posttumor injection in Wistar-Furth (W/Fu) rats inoculated at 2 months of age with either 1 X 10(6) viable metastasizing (TMT-081) or nonmetastasizing (MT-100) tumor cells into the mammary fat pads. The Raji cell assay was used to measure CIC. No correlation between tumor size and CIC levels in the tumors were observed. While none of the sera from the rats bearing the nonmetastasizing tumors had CIC levels higher than 30 micrograms/ml, a small percentage of the animals bearing the metastasizing tumors had serum CIC levels higher than 30 micrograms/ml. This study suggests no clear difference between the amount of CIC in the sera of animals bearing either metastasizing or nonmetastasizing tumors.

Adenocarcinoma↗