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J H Miller

Publications and source records attributed to J H Miller.

At least 55 records · Page 3Linked to original sources
PubMed2001

Amplification of mutator cells in a population as a result of horizontal transfer.

Mutator cells that lack the mismatch repair system (MMR(-)) occur at rates of 10(-5) or less in laboratory populations started from wild-type cells. We show that after selection for recombinants in an interspecies mating between Salmonella enterica serovar Typhimurium and Escherichia coli, the percentage of MMR(-) cells rises to several percent of the recombinant population, and after a second successive mating and selection, greater than 95% of the recombinants are MMR(-). Coupling a single cross and selection with either mutagenesis or selection for spontaneous mutants also results in a dramatic increase in MMR(-) cells. We discuss how horizontal transfer can result in mutator strains during adaptive evolution.

Base Pair Mismatch↗
PubMed2001

Normal brain maturation during childhood: developmental trends characterized with diffusion-tensor MR imaging.

PURPOSE: To characterize the maturational changes in water diffusion within central gray matter nuclei and central white matter pathways of the human brain by using diffusion-tensor magnetic resonance (MR) imaging. MATERIALS AND METHODS: Retrospective analysis of normal MR examination findings in 153 subjects (age range, 1 day to 11 years) referred for clinical neuroimaging was performed. All studies included diffusion tensor-encoded echo-planar MR imaging. Isotropic diffusion coefficient (D) and diffusion anisotropy (A(sigma)) were measured in the corpus callosum, internal capsule, caudate nucleus, lentiform nucleus, and thalamus. RESULTS: exhibited biexponential decay with age in gray and white matter regions, except for monoexponential decay in the genu of the corpus callosum. There was a steep nonlinear increase of A(sigma) in white matter tracts that paralleled the time course of the decline in D. In basal ganglia, only a small linear increase in A(sigma) was observed in patients. A(sigma) changes in the thalamus were intermediate between basal ganglia and white matter structures. CONCLUSION: Changes in magnitude and anisotropy of water diffusion follow stereotypical time courses during brain development that can be empirically described with multiexponential regression models, which suggests that quantitative scalar parameters derived from diffusion-tensor MR imaging may provide clinically useful developmental milestones for brain maturity.

Brain↗
PubMed2001

Control of impurities in diphenhydramine hydrochloride by an ion-pairing, reverse liquid chromatography method.

A precise, sensitive, repeatable and robust reverse-phase liquid chromatographic method has been developed for the control of seven possible impurities of diphenhydramine hydrochloride. The robustness of the method was examined by varying, in turn, each of four mobile-phase parameters (acetonitrile content, buffer salt concentration, ion-pair reagent concentration and pH). The method was linear in the range 0-0.14 mg mL(-1) for diphenhydramine hydrochloride with an acceptable precision and accuracy, and a limit of detection of 0.17 microg mL(-1). Five samples of diphenhydramine hydrochloride from two sources were analysed with the developed liquid chromatographic method.

Chromatography, High Pressure Liquid↗
PubMed2001

Differentiation between bone infarction and acute osteomyelitis in children with sickle-cell disease with use of sequential radionuclide bone-marrow and bone scans.

BACKGROUND: The differentiation of bone infarction from acute osteomyelitis in patients with sickle-cell disease is challenging, as the clinical presentations of the two conditions are similar and imaging and laboratory studies are of limited value. METHODS: A combination of radionuclide bone-marrow and bone scans was performed sequentially within a twenty-four-hour period (with one exception) to aid in the differentiation between bone infarction and osteomyelitis in seventy-nine consecutive episodes of acute bone pain in children with sickle-cell disease. RESULTS: Seventy cases of bone infarction were diagnosed on the basis of decreased uptake on the bone-marrow scan and abnormal uptake on the bone scan at the site of pain. Antibiotic administration was discontinued in sixty-six of the seventy cases after the imaging results were obtained, and the bone pain resolved. In four of the seventy-nine cases, there was normal uptake on the bone-marrow scan and abnormal uptake on the bone scan at the site of pain, findings that were suggestive of acute osteomyelitis. In three of these cases, osteomyelitis was proven by culture, and the symptoms in all four resolved with antibiotic treatment. In five of the seventy-nine cases, the bone-marrow and bone scans were normal and thought to indicate neither osteomyelitis nor bone infarction; in all of these cases, the symptoms resolved without the use of antibiotics. CONCLUSIONS: These findings suggest that osteomyelitis can be differentiated from bone infarction in children with sickle-cell anemia and acute bone pain by a combination of sequential bone-marrow and bone scintigraphy.

Acute Disease↗
PubMed2000

Serum-free culture of rat post-natal and fetal brainstem neurons.

Serum-free medium is essential for cell culture studies in which complete control of the environment is required. Primary culture of post-natal brainstem neurons in defined medium has not been described in the literature, and successful culture of primary brainstem neurons is typically restricted to embryonic ages E14-E18. This study describes a method for culture of fetal and post-natal brainstem neurons using a serum-free culture medium. The culture system is based on Neurobasal medium supplemented with antioxidant-rich B27. Media and supplements are commercially available products from Life Technologies. Neuron survival was optimized by replacing glutamine with GlutaMaxI, by matching osmolality with neuronal age, and by using Hibernate medium to increase neuron survival during tissue dissociation. Fetal E14, E16, E20, and post-natal P3 and P6 cultures were examined after 4, 7, and 9 days in culture. Neuron and glial cells present in the cultures were identified using immunocytochemistry with antibodies raised against microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP), respectively. Fetal E14 cultures had more bipolar neurons than multipolar neurons compared with developmentally older P6 cultures. Early fetal cultures had a higher percentage of neurons than late fetal and early post-natal cultures. Neuron survival was similar between 4 and 9 days in culture for all age groups tested. This is the first reliable, defined culture medium that supports brainstem neurons from late fetal and early post-natal stages of the rat for up to 6 days post-partum.

Animals↗
PubMed2000

In vivo MRI measurements of tumor growth induced by dichloroacetate: implications for mode of action.

Dichloroacetate (DCA) is an important by-product of the chlorination of drinking water that produces liver cancer in rodents. Assessment of the risk that results from concentrations that occur in drinking water will be dependent upon the mode of action held responsible for these tumors. A study by Stauber and Bull [Stauber, A.J. and Bull, R. J (1997) Differences in phenotype and cell replicative behavior of hepatic tumors inducted by dichloroacetate (DCA) and trichloroacetate (TCA). Toxicol. Appl. Pharmacol. 144, 235-246] in mice treated with DCA demonstrated a lesion distribution that was skewed towards many small, altered foci of cells that are assumed to be precursor lesions [EPA, (1996). U.S. Environmental Protection Agency: Proposed Guidelines for carcinogen risk assessment; notice. Fed. Reg. 61, pp. 17960-10811]. The present study was designed to determine the extent to which the tumorigenic effects of DCA could be explained by its effect on tumor growth rates (i.e. tumor promoting activity). In vivo magnetic resonance imaging (MRI) allowed accurate determination of growth rates of individual lesions in mice that had been treated with DCA in drinking water at 2 g/l. Out of thirty treated mice, ten were found to have hepatic tumors detectable by MRI at 48 weeks of treatment. These tumor-bearing animals were assigned to two groups matched on the size of lesions observed by in vivo MR1. Treatment with DCA continued in one group of five mice and was stopped in the other. For both groups, tumor growth rates were determined by measuring changes in size of all lesions greater than 1 mm(3) in volume during a 14-day period. Removal of DCA treatment resulted in growth rates that could not be distinguished from zero across all lesion sizes represented in the sample. These data are in agreement with previous observations of DCAs effects on replication rates within tumors (Stauber and Bull, (1997)). Tumor growth rates observed in animals maintained on treatment decreased with lesion volume in a manner that is consistent with a stochastic Gompertz birth-death process proposed by Tan [Tan, W.Y. (1986) A stochastic Gompertz birth-death process. Stat. Prob. Lett. 4, 25-28]. Parameters of this model obtained by fitting measured growth rates were used to predict the lesion-size distribution expected after one year of DCA treatment. The shape of the predicted lesion-size distribution was similar to that observed by Stauber and Bull (Stauber and Bull, (1997)) in mice sacrificed after 40 weeks of DCA treatment. We conclude that the effects of DCA on the division and/or death rates of spontaneously initiated cells can account for the predominance of small lesions in DCA-treated animals.

Animals↗
PubMed2000

Cloning and characterization of a new member of the Nudix hydrolases from human and mouse.

Proteins containing the Nudix box "GX(5)EX(7)REUXEEXGU" (where U is usually Leu, Val, or Ile) are Nudix hydrolases, which catalyze the hydrolysis of a variety of nucleoside diphosphate derivatives. Here we report cloning and characterization of a human cDNA encoding a novel nudix hydrolase NUDT5 for the hydrolysis of ADP-sugars. The deduced amino acid sequence of NUDT5 contains 219 amino acids, including a conserved Nudix box sequence. The recombinant NUDT5 was expressed in Escherichia coli and purified to near homogeneity. At the optimal pH of 7, the purified recombinant NUDT5 catalyzed hydrolysis of two major substrates ADP-ribose and ADP-mannose with K(m) values of 32 and 83 microM, respectively; the V(max) for ADP-mannose was about 1.5 times that with ADP-ribose. The murine NUDT5 homolog was also cloned and characterized. mNudT5 has 81% amino acid identity to NUDT5 with catalytic activities similar to NUDT5 under the optimal pH of 9. Both NUDT5 and mNudT5 transcripts were ubiquitously expressed in tissues analyzed with preferential abundance in liver. The genomic structures of both NUDT5 and mNudT5 were determined and located on human chromosome 10 and mouse chromosome 2, respectively. The role of NUDT5 in maintaining levels of free ADP-ribose in cells is discussed.

Adenosine Diphosphate Ribose↗
PubMed2000

Selecting protein targets for structural genomics of Pyrobaculum aerophilum: validating automated fold assignment methods by using binary hypothesis testing.

Three-dimensional protein folds were assigned to all ORFs of the recently sequenced genome of the hyperthermophilic archaeon Pyrobaculum aerophilum. Binary hypothesis testing was used to estimate a confidence level for each assignment. A separate test was conducted to assign a probability for whether each sequence has a novel fold-i.e., one that is not yet represented in the experimental database of known structures. Of the 2,130 predicted nontransmembrane proteins in this organism, 916 matched a fold at a cumulative 90% confidence level, and 245 could be assigned at a 99% confidence level. Likewise, 286 proteins were predicted to have a previously unobserved fold with a 90% confidence level, and 14 at a 99% confidence level. These statistically based tools are combined with homology searches against the Online Mendelian Inheritance in Man (OMIM) human genetics database and other protein databases for the selection of attractive targets for crystallographic or NMR structure determination. Results of these studies have been collated and placed at http://www.doe-mbi.ucla.edu/people/parag/P A_HOME/, the University of California, Los Angeles-Department of Energy Pyrobaculum aerophilum web site.

Algorithms↗
PubMed2000

Monte Carlo simulation of single-cell irradiation by an electron microbeam.

A model is presented for irradiation of a cellular monolayer by an electron microbeam. Results are presented for two possible window designs, cells plated on the vacuum-isolation window and cells plated on Mylar above the vacuum-isolation window. Even for the thicker dual-membrane window that facilitates tissue culture and allows the target cell to be centered relative to the electron beam, the majority of the calculated beam spreading was contained in a volume typical of the mammalian HeLa cell line. None of the 10(4) electrons simulated at 25 keV were scattered into the spatial region occupied by neighbors of the target cell. Dose leakage was largest at 50 keV where the mean energy deposited in all neighbors was 21% of that deposited in the target cell. This ratio was reduced to 5% at 90 keV, the highest beam energy simulated. Lineal energy spectra of energy deposition events scored in the nucleus of the target cell became progressively more like the gamma-ray spectrum as the electron beam energy increased. Hence, our simulations provide strong support for the feasibility of a low-LET, single-cell irradiator.

Cell Nucleus↗
PubMed2000

Development of multi-species consortia biofilms of oral bacteria as an enamel and root caries model system.

The aim was to establish defined-species consortium plaque biofilms to investigate enamel and root caries in an artificial mouth. Strains of the putative enamel and root caries pathogens, Streptococcus mutans, Strep. sobrinus, Actinomyces naeslundii and Lactobacillus rhamnosus, were screened in batch culture for potential cariogenic properties: a low terminal pH, ability to aggregate, and catabolic diversity. The strains selected were grown as monoculture biofilms and as consortium plaque biofilms in a multiplaque artificial mouth. The biofilms were supplied with a constant flow of a simulated oral fluid and were given periodic sucrose (and in some instances glucose) to simulate meals. All the bacteria except L. rhamnosus formed large, monospecies biofilms with resting pH in the range 5.3-5.8. The consortia biofilms were larger and had a resting pH of 4.9-5.3. The consortia biofilms supplied with 8-hourly carbohydrate comprised mainly 'mutans' streptococci (58, SD 5.5%) and L. rhamnosus (42, SD 5.7%). A. naeslundii characteristically was absent or present in a low percentage (up to 4% colony-forming units). All biofilms demineralized polished bovine enamel and dentine blocks, as assessed by microradiography and enamel-surface microhardness measurement. The consortia also demineralized intact enamel and tooth roots; they were more cariogenic to enamel than any of the monoculture biofilms, as measured by enamel-surface softening, but variation in lesion depth was proportional to biofilm wet weight irrespective of acidogen composition (r = 0.93, p < 0.05). Enamel lesions had a well-mineralized intact surface and a zone of subsurface demineralization, typical of early natural lesions. Dentine and root lesions showed extensive demineralization but lacked a pronounced surface mineralized zone. Substitution of glucose for sucrose had no effect on the cariogenicity of the consortium to bovine enamel or human roots and had no major effect on the plaque composition. Continuously supplied fluoride (19 parts/10(6)) resulted in a substantially reduced enamel surface softening and subsurface demineralization of intact roots. It was concluded that consortia biofilms of selected caries pathogens generate realistic caries lesions in all tooth hard tissues under controlled growth conditions in the artificial mouth. This in vitro caries experimental model may prove useful for the study of interrelations between the plaque biofilm, tooth tissues and the oral environment, and for the development of procedures to modify the course of caries development.

Actinomyces↗
PubMed2000

Molecular mass and volume in radiation target theory.

Radiation target analysis is based on the action of ionizing radiation directly on macromolecules. Interactions of this radiation with the molecules leads to considerable structural damage and consequent loss of biological activity. The radiation sensitivity is dependent on the size of the macromolecules. There has been confusion and discrepancy as to whether the molecular mass or the molecular volume was the determinant factor in the sensitivity. Some proteins are known to change their hydrodynamic volume at low pH, and this characteristic can be utilized to compare the radiation sensitivities of these proteins in the two states. The results show that the radiation sensitivity of proteins depends on the mass of the molecule and is independent of the molecular volume/shape.

Animals↗
PubMed2000

Relationship of early inhalant use to substance use in college students.

This study examined the relationship between early inhalant use and later substance use in a sample of college students. Data were taken from four campus-wide surveys that assessed changes in rates of substance use over time at a large Southwestern university. Within these surveys, a group of students (n = 187) who reported early use of inhalants (i.e., before age 18) was identified and examined in terms of their substance use behavior while in college (i.e., during the year and month prior to the survey). These students were compared to two other groups of students: those who reported early use of marijuana but no early use of inhalants (n = 1271) and students who reported no early use of either inhalants or marijuana (n = 1479). Results show that early use of either inhalants or marijuana substantially increased risk of frequent drinking, binge drinking, smoking, illicit drug use, and substance-related consequences during the college years. However, the early use of inhalants conferred the greatest risk and was associated with twice the rate of binge and frequent drinking and significantly greater rates of tobacco and drug use than early marijuana use alone. Implications of these findings for intervention and prevention with college students are discussed.

Administration, Inhalation↗
PubMed2000

Epidermal growth factor receptor is increased in rabbit intestinal brush border membrane after small bowel resection.

A defective epidermal growth factor receptor (EGFR) abrogates adaptation, while overexpression of EGFR or exogenous epidermal growth factor (EGF) enhances adaptation following small bowel resection (SBR). EGFR is predominantly located on the enterocyte basolateral membrane, yet luminal EGF is increased in injured mucosa. We hypothesized that EGFR is both increased and redistributed to the enterocyte brush border membrane (BBM) after SBR and that parenteral EGF will reverse this redistribution. Rabbits (N = 20) were subjected to sham operation or SBR. EGF or vehicle were administrated one week postoperatively to SBR rabbits, and the gut was harvested one week later. EGFR levels in intestinal crude extracts and purified BBM were determined by Western blot analysis. No difference in EGFR level was detected in the crude extract among any of the groups. SBR more than doubled EGFR amount in BBM (P < 0.006). Parenteral EGF did not influence this redistribution. Thus, EGFR is partially redistributed to the BBM in the mucosa of SBR rabbits, and parenteral EGF does not reverse this redistribution.

Animals↗
PubMed2000

Genes involved in the determination of the rate of inversions at short inverted repeats.

BACKGROUND: Not all of the enzymatic pathways involved in genetic rearrangements have been elucidated. While some rearrangements occur by recombination at areas of high homology, others are mediated by short, often interrupted homologies. We have previously constructed an Escherichia coli strain that allows us to examine inversions at microhomologies, and have shown that inversions can occur at short inverted repeats in a recB,C-dependent fashion. RESULTS: Here, we report on the use of this strain to define genetic loci involved in limiting rearrangements on an F' plasmid carrying the lac genes. Employing mini-Tn10 derivatives to generate insertions near or into genes of interest, we detected three loci (rmuA,B,C) that, when mutated, increase inversions. We have mapped, cloned and sequenced these mutator loci. In one case, inactivation of the sbcC gene leads to an increase in rearrangements, and in another, insertions near the recE gene lead to an even larger increase. The third gene involved in limiting inversions, rmuC, has been mapped at 86 min on the E. coli chromosome and encodes a protein of unknown function with a limited homology to myosins, and some of the SMC (structural maintenance of chromosomes) proteins. CONCLUSIONS: This work presents the first example of an anti-mutator role of the sbcC,D genes, and defines a new gene (rmuC) involved in DNA recombination.

Chromosome Inversion↗
PubMed2000

The consequences of growth of a mutator strain of Escherichia coli as measured by loss of function among multiple gene targets and loss of fitness.

We have examined the composition of members of mutator populations of Escherichia coli by employing an extensive set of phenotypic screens that allow us to monitor the function of >700 genes, constituting approximately 15% of the genome. We looked at mismatch repair deficient cells after repeated cycles of single colony isolation on rich medium to generate lineages that are forced through severe bottlenecks, and compared the results to those for wild-type strains. The mutator lineages continued to accumulate mutations rapidly with each increasing cycle of colony isolation. By the end of the 40th cycle, after approximately 1000 generations, most of the lineages had reduced colony size, 4% had died out, 55% had auxotrophic requirements (increasing to 80% after 60 cycles), and 70% had defects in at least one sugar or catabolic pathway. In addition, 33% had a defect in cell motility, and 26% were either temperature-sensitive or cold-sensitive lethals. On the other hand, only 3% of the wild-type lineages had detectable mutations of any type after 40 cycles. By the 60th cycle, the typical mutator cell carried 4-5 inactive genes among the 15% of the genome being monitored, indicating that the average cell carried at least 24-30 inactivated genes distributed throughout the genome. Remarkably, 30% of the lineages had lost the ability to utilize xylose as a carbon source. DNA sequencing revealed that most of the Xyl(-) mutants had a frameshift in a run of eight G's (GGGGGGGG) in the xylB gene, either adding or deleting one -G-. Further analysis indicated that rendering E. coli deficient in mismatch repair unmasks hypermutable sites in certain genes or intergenic regions. Growth curves and competition tests on lineages that passed through 90 cycles of single colony isolation showed that all lineages suffered reduced fitness. We discuss these results in terms of the value of mutators in cellular evolution.

Adaptation, Biological↗