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M J Miller

Publications and source records attributed to M J Miller.

At least 361 records · Page 20Linked to original sources
PubMed1988

Inter- and intraclonal variability of polypeptides synthesized in a rat hepatoma cell line. Quantitative two-dimensional gel analysis.

To examine the degree of clonal heterogeneity in the synthesis of polypeptides in neoplastic cells, single-cell subclones from the rat hepatoma cell line H4-II-E were isolated. Polypeptides from the clones were resolved on high resolution two-dimensional polyacrylamide gels (PAGE), and quantitatively analyzed with a computerized two-dimensional PAGE analysis system developed in this laboratory. Only four qualitatively different spots were found which were synthesized in one of the subclones in four out of five experiments. In contrast, 5-20% of the spots showed statistically significant quantitative differences when any one subclone was compared to any other. These differences were generally quite small, averaging about 1.5-fold in intensity, although variations of fourfold or more were observed. Different cultures of the same subclone showed quantitative differences of the same order as seen in different subclones, indicating that this variability was primarily intraclonal in nature, i.e. associated with the cultures rather than the subclones. The distribution of quantitatively variable spots indicates that 50% or more of the polypeptides in these cells may display intraclonal variability. Similar results were obtained with a second set of subclones derived from these primary ones. Time course studies were conducted where cells were maintained continuously for 12 weeks, with samples taken for two-dimensional PAGE analysis once a week. The fraction of polypeptides that vary significantly generally increased with time between sampling points. Experiments with independent cultures grown in parallel indicate that about 4% of this variability can be correlated to the age of the culture media, although the majority appears due to uncontrolled and/or random differences that arise between cultures. These results indicate that independent cultures quickly develop detectable quantitative differences in the expression of a large fraction of their polypeptides. These differences cannot, at present, be associated with the observable biology of the cells and probably reflect time-associated variations in the balance of cellular macromolecular synthesis which arise in tissue culture cells.

Animals↗
PubMed1988

Comparison of gene expression in preneoplastic and neoplastic rat liver to adult, fetal, regenerating, and tumor-promoted liver.

Computer-assisted analysis was performed on the in vitro translation products of polyadenylated RNA samples isolated from normal adult Fischer rat liver and from preneoplastic and neoplastic rat liver samples which were generated by the Solt Farber technique (Solt, D. and Farber, E., Nature 263:701-703, 1976). The vast majority of the differences in translation products observed throughout the progressive development of hepatocellular carcinoma was quantitative in nature. Importantly, this quantitative heterogeneity first became prevalent at the very early preneoplastic stage of hepatoma formation. Only 3 consistent qualitative alterations in translation products were observed to be associated with the hepatocarcinogenesis process. The appearance of two new polypeptides of molecular weight and isoelectric point of 32/5.2 and 43/5.1 appeared to be related to an early preneoplastic event in hepatoma development and the transition from a preneoplastic to a neoplastic state, respectively. Importantly, these new polypeptides were not observed in the in vitro translation products generated from fetal or regenerating liver samples or from liver samples which were chronically treated with phenobarbital or terachlorodibenzo-p-dioxin. One translation product (located at 35/6.6) of normal adult, fetal, and regenerating liver RNA samples was undetected in all preneoplastic, neoplastic, phenobarbital-, and terachlorodibenzo-p-dioxin-treated liver RNA translation products. The possibility exists that the specific loss of this gene product may promote the development of the transformed phenotype.

Animals↗
PubMed1988

The active form of the cytochrome d terminal oxidase complex of Escherichia coli is a heterodimer containing one copy of each of the two subunits.

The cytochrome d complex is a component of the aerobic respiratory system of Escherichia coli. The enzyme functions as a terminal oxidase, oxidizing ubiquinol-8 within the cytoplasmic membrane and reducing oxygen to water. The enzyme is of particular interest because it is a coupling site in the electron transfer chain. The electron transfer reaction catalyzed by this enzyme is coupled to the translocations of protons across the membrane (H+/e-approximately equal to 1). The oxidase contains two subunits by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, with molecular weights of 58,000 and 43,000. In this paper, the question of the quaternary structure is addressed. Quantitative N-terminal analysis of the isolated enzyme and relative mass quantitation following sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicate the subunits are present in equimolar amounts. Sedimentation velocity and sedimentation equilibrium studies were used to characterize the hydrodynamic properties of the purified enzyme solubilized in Triton X-100, under conditions where the enzyme is active. It is concluded that the active enzyme in Triton X-100 is a heterodimer, containing one copy of each subunit. This is likely the structure of the enzyme in the E. coli membrane.

Algorithms↗
PubMed1988

Elsie 4: quantitative computer analysis of sets of two-dimensional gel electrophoretograms.

We have developed and refined a system for quantitative computer analysis of two-dimensional polyacrylamide gel electrophoretograms. The system, named Elsie 4, is based on one described by Vo et al. (Anal. Biochem. 112, 258 (1981]. It is highly automated. Elsie 4 can find, and measure the intensity of, almost any spot resolvable on two-dimensional gels, including spots visible only as shoulders off larger spots and spots so close together that there is no "valley" between them. It can automatically match the spot patterns of different gels, potentially without the need for a user to provide landmark matches. The matches between paired gels let us follow the synthesis of any spot through a set of gels. Information about a group of matched spots can be obtained by referring to any spot in the group. There is generally no need for a standard or reference gel. Data for two experiments can be combined and compared by matching any gel in one experiment with any gel in the other. There are ways to automatically find possible mismatches in sets of gels. Scans and the results of the analysis can be shown on an image displayer. The programs use function libraries; this helps ensure consistency and increase portability. The programs and functions can be linked together in many ways; this lets users build custom programs for analysis of specific experiments.

Computer Systems↗
PubMed1988

Electroconvulsive treatment compared with lithium in the management of manic states.

Thirty-four hospitalized manic patients were randomized to treatment with either lithium carbonate or an average series of nine bilateral electroconvulsive treatments (ECTs), followed by maintenance with lithium carbonate. Weekly ratings of manic, depressive, and psychotic symptoms were obtained for eight weeks, and patients were followed up monthly for up to two years. Ratings by nonblind and blind observers indicated that the patients who underwent ECT improved more during the first eight weeks than did patients who were treated with lithium carbonate. This was especially true of patients with mixed symptoms of mania and depression and/or extreme manic behavior. Clinical ratings after eight weeks showed no significant differences between the lithium carbonate- and ECT-treated patients. Likewise, the two groups had comparable rates of relapse, recurrence, and rehospitalization during the follow-up period.

Adult↗
PubMed1988

Assessment of aortic pressure-volume relationships with an impedance catheter.

Volume measurements obtained with an impedance catheter (impedance volume measurements) have been used previously in determining pressure-volume relationships in the right and left ventricles and in the right atrium. The purpose of the present study was to determine the utility of impedance volume techniques in the assessment of aortic pressure-volume relationships in animals and humans. Experiments to develop this methodology were first performed in ten anesthetized dogs, with simultaneous measurements of aortic pressure (micromanometer), diameter (ultrasonic crystals), and impedance volume. Transient (20 sec) inferior vena cava (IVC) occlusion and nitroprusside infusion resulted in a reproducible series of aortic pressure-volume and pressure-diameter points. In each case changes in impedance volume closely paralleled changes in diameter. From the wide range of pressure-volume points, it was possible to construct pressure-volume plots for a single cardiac cycle, as well as longer loops over a 20-40 mm Hg pressure range. End-systolic and end-diastolic pressure-volume points could subsequently be identified, which outlined a more precise pressure-volume relationship that was linear in six cases and curvilinear in four cases, depending on the level of aortic pressure. We extended these techniques to human subjects undergoing diagnostic cardiac catheterization. Studies using the impedance catheter in the aorta were performed in 11 patients. Aortic pressure-volume curves were elicited during Valsalva maneuver to obtain data over a range wider than that of a single cardiac cycle. Measurements of relative volume were calibrated with simultaneous M-mode echocardiography in three patients. We conclude that pressure-volume relationships in the aorta can be assessed with impedance volume techniques, making possible in vivo measurements of arterial compliance. Potential nonlinearities in aortic P-V relationships require that the assessment of aortic compliance encompass as much as possible of the physiologic range of pressure and volume.

Animals↗
PubMed1988

Propranolol and inspiratory muscle performance in normal subjects.

We studied the effects of propranolol on respiratory muscle performance during inspiratory threshold-loaded endurance breathing in eight normal subjects. Propranolol (mean daily dose = 160 + 17 mg, SEM) reduced loaded 15-second MVV (92 versus 81 L/min;P = .01) and maximal sustainable ventilatory capacity (52.3 versus 44.5 L/min, P = .02) but did not affect the fraction of MVV, which was sustainable. Maximal static inspiratory pressures were reduced at two of three lung volumes, whereas maximal static expiratory pressures were unaffected by propranolol. The reduction in inspiratory muscle performance in the whole population could be accounted for almost entirely by four subjects who developed symptoms of "tiredness" and easy fatigability while receiving propranolol. There was no significant difference in propranolol dose, in degree of beta-receptor blockade, or in physical fitness in symptomatic and asymptomatic groups, and in neither group did propranolol alter pulmonary function test results. Propranolol directly depresses inspiratory muscle strength in subjects who develop drug-induced symptoms of fatigue by a mechanism probably unrelated to beta-adrenergic-receptor blockade.

Adult↗
PubMed1988

Methods for increasing the resolution of two-dimensional protein electrophoresis.

A two-dimensional gel elctrophoresis protocol has been developed which provides for a 1.5-to 3-fold increase in the resolution of proteins compared to other frequently used methods. The major variations from previous protocols include increased pore size in the isoelectric focusing gels; cholamidopropyldimethylhydroxypropanesulfonate, a zwitterionic detergent, replaces most of the Nonidet P-40, a nonionic detergent, in the isoelectric focusing gels; no equilibration step is employed between the first and second dimensional separation. The use of a stacking gel in the second dimension has been eliminated; a more efficient and evenly distributed cooling system has been designed for the molecular mass separation, allowing faster migration with higher current. Finally, the crosslinker diacrylylpiperazine is employed which improves protein separation and detection with ammoniacal silver staining. Silver-stained two-dimensional gel electrophoretograms of human plasma and hamster brain tissues and autoradiographs of rat liver cells are compared to the results obtained from previous methods.

Animals↗
PubMed1988

Extracellular metabolites of 2-aminofluorene in cultures of rapid and slow acetylator rabbit hepatocytes as a model for urinary and biliary metabolism.

N-acetylation is involved in determining species susceptibility to carcinogenicity by certain aromatic amines. In order to further investigate this relationship, the biotransformation of 2-aminofluorene (2-AF) by monolayer cultures of hepatocytes isolated from rapid and slow acetylator rabbits was studied. Analysis of biotransformation products liberated by cells was used as an indication of metabolites that would be excreted in the urine and bile. Hepatocytes from both acetylator phenotypes were found to extensively biotransform 2-AF. The overall rates of metabolism and the types of products formed were similar in the two phenotypes, although the quantity of several products differed. Hepatocyte cultures from rapid acetylators released a greater proportion of acetylated metabolites. Rapid acetylator hepatocytes released predominantly ring-hydroxylated-2-acetylaminofluorene (2-AAF) while the major product from the slow acetylator cultures was conjugated 2-AF. The amounts of extracellular N-hydroxy-2-acetylaminofluorene were similar in both phenotypes. No phenotype-dependent differences in extracellular metabolites were noted when hepatocytes were incubated with 2-AAF. These results indicate that hepatocytes from both phenotypes have similar capacities to excrete N-hydroxy-2-AAF and to detoxify the parent aromatic amine. These findings can be related to the carcinogenicity of 2-AF in either phenotype.

2-Acetylaminofluorene↗
PubMed1988

Effect of head position on distribution of nasal airflow in preterm infants.

Supine preterm infants characteristically adopt a lateral head position; however, it is not known whether this influences the distribution of nasal airflow. Ventilation was measured in 12 healthy preterm infants (postconceptional age 34 +/- 2 weeks) by employing a nasal mask pneumotachygraph that separated airflow between the left and right nasal passages. In the midline supine position, the percent of total tidal volume (%VT) through the right nasal passage ranged from 31% to 64% and varied by less than 5% between active and quiet sleep in any infant. Lateral positioning of the head caused %VT to increase on the dependent side and decrease through the upper nasal passage. When the right side was dependent, mean %VT on that side increased from 52 +/- 9% to 67 +/- 14% (P less than 0.01) and decreased to 43 +/- 10% (P less than 0.05) when the right side was up. In the midline position, the presence of a nasogastric tube caused %VT through the nasal passage with the tube to fall from 54 +/- 8% to 39 +/- 8% (P less than 0.01). The %VT fell farther, to 25 +/- 10% (P less than 0.01), when the nasal passage with the nasogastric tube was up. Despite these changes in VT distribution, total VT remained constant during these maneuvers. We speculate that when supine preterm infants adopt a lateral head position, the decrease in airflow through the upper nasal passage results from partial obstruction of the oropharyngeal or nasopharyngeal airway on that side.

Head↗
PubMed1988

Effects of protein, mucin, and human tears on adherence of Pseudomonas aeruginosa to hydrophilic contact lenses.

Pseudomonas aeruginosa isolated from a corneal ulcer was examined for its ability to adhere to hydrophilic contact lenses with adsorbed organic films. Generally, adherence was enhanced by mucin, lactoferrin, lysozyme, immunoglobulin A, bovine serum albumin, and a mixture of these macromolecules. The water content of the lenses also influenced bacterial adherence and protein adsorption. Adherence to lenses coated with human tear films was more complex; both enhanced and inhibited adherence were observed on lenses obtained from different individuals.

Bacterial Adhesion↗
PubMed1988

SOD prevents damage and attenuates eicosanoid release in a rabbit model of necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) was produced in anesthetized rabbits by transmural injection of intestinal loops with an acidified solution of casein and calcium gluconate, mimicking the luminal milieu of afflicted neonates. Intravenous infusion of superoxide dismutase (SOD) 15 min after NEC induction prevented intestinal damage. In ex vivo perfused intestinal loops, we determined the sites of eicosanoid release and their contribution to the vascular effects of N-formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF) in damaged and SOD-salvaged intestine. The vascular effluent was the primary site of stimulated eicosanoid release. The vascular responses to fMLP (vasoconstriction) and PAF (vasodilation) were not altered by SOD, although vascular resistance was higher in the SOD group. SOD treatment attenuated 1) transmural fluid shifts in ex vivo perfused intestinal preparations, an index of vascular permeability, 2) fMLP-induced prostaglandin E2, 6-ketoprostaglandin F1 alpha (6-keto-PGF1 alpha), and leukotriene B4 (LTB4) release, and 3) PAF-induced release of 6-keto-PGF1 alpha and LTB4. Stimulated thromboxane B2 release was not altered by SOD. Thus NEC can be established by a luminal insult that causes local generation of free radicals and exaggerated release of prostaglandins and leukotrienes.

6-Ketoprostaglandin F1 alpha↗
PubMed1988

Renal cytochrome P-450-related arachidonate metabolism in rabbit aortic coarctation.

Cells of the medullary segment of the thick ascending limb of Henle's loop (TALH) convert arachidonic acid (AA) via the cytochrome P-450 monooxygenase pathway to biologically active metabolites: P1, a vasorelaxant, and P2, an inhibitor of Na+-K+-ATPase activity. These AA metabolites may contribute to the renal vascular and metabolic adjustments in response to renal hypoperfusion and the attendant elevation of blood pressure produced by suprarenal aortic coarctation. On the eighth postoperative day, the blood pressures of hypertensive and sham-operated control rabbits were 105 (90-115) and 63 (60-64) mmHg (medians with semiquartile values), respectively (P less than 0.01). Formation of P1 and P2 was increased twofold in TALH cells obtained from hypertensive rabbits: 2.35 (1.79-4.83) and 1.28 (1.56-4.56) micrograms AA converted.mg protein-1.30 min-1 compared with sham-operated rabbits: 1.27 (1.03-1.53) and 0.64 (0.58-1.10) micrograms AA converted.mg protein-1.30 min-1 (P less than 0.05). The profile of biological activity of AA metabolites contained within P1 and P2 was unaffected by aortic coarctation. The cytochrome P-450 monooxygenase-derived AA metabolites may exert a defensive function to limit the degree of TALH cell injury in response to renal hypoperfusion and associated zonal anoxia by reducing energy-dependent Na+-K+-ATPase activity and affecting local vasodilatation.

Animals↗
PubMed1988

Airway obstruction during periodic breathing in premature infants.

To characterize changes in pulmonary resistance, timing, and respiratory drive during periodic breathing, we studied 10 healthy preterm infants (body wt 1,340 +/- 240 g, postconceptional age 35 +/- 2 wk). Periodic breathing in these infants was defined by characteristic cycles of ventilation with intervening respiratory pauses greater than or equal to 2 s. Nasal airflow was recorded with a pneumotachometer, and esophageal or pharyngeal pressure was recorded with a fluid-filled catheter. Pulmonary resistance at half-maximal tidal volume, inspiratory time (TI), expiratory time (TE), and mean inspiratory flow (VT/TI) were derived from computer analysis of five cycles of periodic breathing per infant. In 80% of infants periodic breathing was accompanied by completely obstructed breaths at the onset of ventilatory cycles; the site of airway obstruction occurred within the pharynx. The first one-third of the ventilatory phase of each cycle was accompanied by the highest airway resistance of the entire cycle (168 +/- 98 cmH2O.l-1.s). In all infants TI was greatest at the onset of the ventilatory cycle, VT/TI was maximal at the midpoint of the cycle, and TE was longest in the latter two-thirds of each cycle. A characteristic increase and subsequent decrease of 4.5 +/- 1.9 ml in end-expiratory volume also occurred within each cycle. These results demonstrate that partial or complete airway obstruction occurs during periodic breathing. Both apnea and periodic breathing share the element of upper airway instability common to premature infants.

Airway Obstruction↗
PubMed1988

Increasing arterial carbon dioxide tension: influence on transcutaneous carbon dioxide tension measurements.

Despite widespread use of transcutaneous PCO2 (TcPCO2) monitoring, the precise relationship between TcPCO2 and PaCO2 remains unclear. It has been widely assumed that theoretical correction of TcPCO2 (combining temperature correction with a constant metabolic factor of 4 mm Hg) accounts for the elevation of TcPCO2 over PaCO2. To test this assumption, TcPCO2 was measured with a 44 degrees C electrode and compared to PaCO2 in 60 normotensive infants with cardiorespiratory disease during the first four +/- six days of life (mean +/- SD) (range one to 36 days). During hypocapnea, from PaCO2. In contrast, during normocapnea, theoretically corrected TcPCO2 exceeded PaCO2 by 5 +/- 4 mm Hg (P less than .001), and similarly during hypercapnea, theoretically corrected TcPCO2 exceeded PaCO2 by 9 +/- 6 mm Hg (P less than .001). These data suggest that, as PaCO2 increases, there may be an imbalance between tissue CO2 production and removal, resulting in a progressively increasing gradient between TcPCO2 and PaCO2. Clarification of the relationship between TcPCO2 and PaCO2 should enhance the interpretation of TcPCO2 measurements in infants.

Blood Gas Monitoring, Transcutaneous↗
PubMed1988

Contribution of oxygen-derived free radicals to experimental necrotizing enterocolitis.

Oxygen-derived free radicals, particularly superoxide anion, are considered important mediators of intestinal injury induced by ischemia/reperfusion based on the protective effects of superoxide dismutase and allopurinol. A role for free radicals was investigated in a model of necrotizing enterocolitis (NEC) which was initiated by a luminal, as opposed to a vascular, insult. Intestinal loops of weanling rabbits received either saline (control loops) or a solution of 10 mg/ml casein and 50 mg/ml calcium gluconate acidified to pH 4 with proprionic acid (treated loops). When the animals were sacrificed 3 hours later, severe damage was noted in the treated loops, which included blunting of villi and edema, with all animals surviving. At 16 hours only 5 of 8 rabbits survived, and 3 had hemorrhagic necrosis. Control loops were normal in each case. Intravenous infusion of superoxide dismutase (4 mg/kg/hr), commencing 15 minutes after NEC induction, totally prevented intestinal injury. On the other hand, pretreatment with allopurinol, an inhibitor of xanthine oxidase, for 2 days (30 and 60 mg/kg by mouth) was not protective against intestinal damage. A cellular infiltration in treated loops was not histologically evident in the majority of animals at 3 hours after treatment, a finding confirmed by the minimal accumulation of 111In-labeled leukocytes in damaged and intact intestinal tissue. These results suggest that superoxide generated locally from sources other than xanthine oxidase play a critical and early role in experimental NEC and that superoxide dismutase may prove to be an effective therapy in this devastating neonatal disease.

Allopurinol↗
PubMed1987

Sensitivity of RNA synthesis to actinomycin D inhibition is dependent on the frequency of transcription: a mathematical model.

The synthesis of ribosomal RNA is known to be up to 100-fold more sensitive to inhibition by actinomycin D than is messenger RNA. A model is presented here to explain the dose-response kinetics of this inhibition. The basic concept is that on very actively transcribed genes, such as ribosomal DNA, the first bound actinomycin D will sterically affect those densely packed polymerase molecules between it and the promoter, causing them to stack up into the promoter and interfere with the initiation of RNA synthesis. However, on less active genes, where the polymerases are widely spaced, the drug will inhibit individual polymerasas independently and only at the actinomycin blockade. Counteracting these inhibitory effects will be the tendency of genes bound with actinomycin to accumulate additional polymerases. A mathematical model is described which successfully explains previously reported dose-response kinetics of actinomycin inhibition in both frequently and infrequently transcribed genes. The analysis indicates that actinomycin inhibition is dependent on both polymerase packing and on gene size. The dose-response kinetics can be used to estimate both the size and transcriptional efficiency of individual genes. The model is also able to explain several other independent observations regarding the kinetics of inhibition of RNA synthesis by actinomycin.

Dactinomycin↗