Biomedical subjects

J H Miller

Publications and source records attributed to J H Miller.

At least 73 records · Page 4Linked to original sources

PubMed2000

Characterization of a thermostable DNA glycosylase specific for U/G and T/G mismatches from the hyperthermophilic archaeon Pyrobaculum aerophilum.

U/G and T/G mismatches commonly occur due to spontaneous deamination of cytosine and 5-methylcytosine in double-stranded DNA. This mutagenic effect is particularly strong for extreme thermophiles, since the spontaneous deamination reaction is much enhanced at high temperature. Previously, a U/G and T/G mismatch-specific glycosylase (Mth-MIG) was found on a cryptic plasmid of the archaeon Methanobacterium thermoautotrophicum, a thermophile with an optimal growth temperature of 65 degrees C. We report characterization of a putative DNA glycosylase from the hyperthermophilic archaeon Pyrobaculum aerophilum, whose optimal growth temperature is 100 degrees C. The open reading frame was first identified through a genome sequencing project in our laboratory. The predicted product of 230 amino acids shares significant sequence homology to [4Fe-4S]-containing Nth/MutY DNA glycosylases. The histidine-tagged recombinant protein was expressed in Escherichia coli and purified. It is thermostable and displays DNA glycosylase activities specific to U/G and T/G mismatches with an uncoupled AP lyase activity. It also processes U/7,8-dihydro-oxoguanine and T/7,8-dihydro-oxoguanine mismatches. We designate it Pa-MIG. Using sequence comparisons among complete bacterial and archaeal genomes, we have uncovered a putative MIG protein from another hyperthermophilic archaeon, Aeropyrum pernix. The unique conserved amino acid motifs of MIG proteins are proposed to distinguish MIG proteins from the closely related Nth/MutY DNA glycosylases.

Amino Acid Sequence↗

PubMed2000

A theory-based motivational approach for reducing alcohol/drug problems in college.

The Campuswide Alcohol and Drug Abuse Prevention Program (CADAPP) was implemented and evaluated over a 1.5-year span at the University of New Mexico (UNM). Drawing on self-regulation theory as a basis for understanding motivation for change, the program was designed to increase risk perceptions and thereby reduce the use of alcohol and other drugs among university students. The program was evaluated from 1988 to 1989 through repeated anonymous random sample surveys of all enrolled students on the UNM campus and on a similar control campus not implementing new prevention efforts during the same period. As predicted, relative to the control campus, students on the CADAPP campus after the program showed significantly higher perceived risks from substance use and significantly reduced levels of alcohol and marijuana use. These findings provide encouraging evidence for this theory-based approach to primary and secondary prevention.

Adolescent↗

PubMed2000

Reducing alcohol use in college students: a controlled trial of two brief interventions.

This study tested two forms of alcohol reduction programming for college students. Thirty-seven moderate to heavy drinkers completed measures of quantity/frequency, drinking consequences, and attitude questionnaires. Participants were randomly assigned to one of three groups: 1) a two-hour information and motivation session plus mailed personal feedback on their drinking; 2) mailed feedback only; or 3) no treatment. At a 6-week follow-up session, the feedback-only group decreased drinks per month as compared to control. No other differences were statistically significant, though decreases favored the treatment conditions about equally over control. Implications for research and treatment are discussed.

Adult↗

PubMed1999

Cytochrome c unfolding on an anionic surface.

It is now well accepted that the adsorption of proteins to solid supports sometimes involves surface-mediated unfolding. A detailed understanding of the adsorption and surface-mediated unfolding process is lacking. We selected a well studied protein, horse heart cytochrome c, and a weakly ionic support to examine some of the characteristics of protein adsorption under near-physiological conditions. We used high-performance liquid chromatography (HPLC) to investigate the effect of temperature on surface-mediated unfolding. Samples of cytochrome c were introduced to an anionic support, and a NaCl gradient was used to desorb the protein at different times and temperatures. The profiles and retention times were monitored to examine the adhesive properties of cytochrome c to the anionic support. We found that protein retention increased with time at temperatures as low as 0 degrees C, and a significant loss of cytochrome c occurred between 55 degrees C and 70 degrees C. The loss of recovery of cytochrome c indicates irreversible surface-mediated unfolding. The changes in retention time may indicate more subtle transitions, including reversible surface-mediated unfolding of cytochrome c. These results suggest that perturbations in the structure as well as unfolding of cytochrome c can be detected at a lower temperature on an anionic surface than in solution thereby acting like a catalyst for protein unfolding.

Adsorption↗

PubMed1999

A thermostable vacuolar-type membrane pyrophosphatase from the archaeon Pyrobaculum aerophilum: implications for the origins of pyrophosphate-energized pumps.

Vacuolar-type H(+)-translocating pyrophosphatases (V-PPases) have been considered to be restricted to plants, a few species of phototrophic proteobacteria and protists. Here, we describe PVP, a thermostable, sequence-divergent V-PPase from the facultatively aerobic hyperthermophilic archaeon Pyrobaculum aerophilum. PVP shares only 38% sequence identity with both the prototypical V-PPase from Arabidopsis thaliana and the H(+)-PPi synthase from Rhodospirillum rubrum, yet possesses most of the structural features characteristic of V-PPases. Heterologous expression of PVP in Saccharomyces cerevisiae yields a M(r) 64¿ omitted¿000 membrane polypeptide that specifically catalyzes Mg(2+)-dependent PPi hydrolysis. The existence of PVP implies that PPi-energized H(+)-translocation is phylogenetically more deeply rooted than previously thought.

Amino Acid Sequence↗

PubMed1999

Chelation of iron within the erythrocytic Plasmodium falciparum parasite by iron chelators.

To examine the site of action of antimalarial iron chelators, iron ligands were added to control erythrocytes and to erythrocytes parasitized with Plasmodium falciparum, and the concentration of intracellular labile iron was monitored with the fluorescent probe, calcein. The fluorescence of calcein quenches upon binding iron and increases upon releasing iron. The chelators included desferrioxamine B, 2',2'-bipyridyl, and aminophenol II, a compound that is being newly reported as having anti-plasmodial properties. Calcein-loaded parasitized cells displayed fluorescence predominantly within the cytosol of both rings and trophozoites. The addition of chelators to both control and parasitized erythrocytes led to significant increases of fluorescence (P < 0.001). Fluorescence was observed to increase within the parasite itself after addition of iron chelators, indicating that these agents bound labile iron within the plasmodium. The relative increases of fluorescence after addition of chelators were greater in control than parasitized erythrocytes (P < 0.05) as were the estimated labile iron concentrations (P < or = 0.001). These results suggest that (i) the anti-malarial action of iron chelators might result from the ability to reach the infected cell's parasite compartment and bind iron within the parasite cytosol, and (ii) the labile iron pool of the host red cell may be either utilized or stored during plasmodial growth.

2,2'-Dipyridyl↗

PubMed1999

Small bowel adaptation is dependent on site of massive enterectomy.

BACKGROUND: Changes in amino acid transport after massive enterectomy occur in a nutrient-dependent fashion and may affect long-term outcome. Epidermal growth factor (EGF) can enhance nutrient transport and a defective epidermal growth factor receptor (EGF-R) has been noted to attenuate adaptation. Most animal studies, however, have examined only a single site of resection. This does not mimic the clinical situation where disease dictates the site of resection leading to proximal, middle, or distal enterectomies. We hypothesize that the site of massive enterectomy will alter nutrient transport and EGF-R levels in the residual gut. MATERIALS AND METHODS: New Zealand White rabbits were randomized to control, midgut division, or 70% resection (proximal, midgut, or distal). After 1 week, sodium-dependent transport of glucose, glutamine, alanine, and leucine into brush border membrane vesicles was quantitated. EGF-R protein was determined by Western blot analysis. RESULTS: At baseline, amino acid transport was greater in ileum than jejunum. Surgery alone elevated glutamine and leucine jejunal transport by 130 and 97%, respectively, over controls (P < 0.05). Midgut resection increased jejunal glutamine transport 61% over control (P < 0.05). In contrast, distal resection increased jejunal alanine transport by 150% over controls with no change in glutamine (P < 0.05). After midgut resection, EGF-R was significantly greater (124%) in ileum then in jejunum in whole mucosa homogenates. Proximal resection significantly lowered ileal EGF-R compared to that seen in midgut resected residual ileum. CONCLUSIONS: Site of massive resection is important in determining postoperative changes in nutrient transport and EGF-R.

Adaptation, Physiological↗

PubMed1999

Growth inhibitory effects of a mu opioid on cultured cholinergic neurons from fetal rat ventral forebrain, brainstem, and spinal cord.

Cholinergic pathways play a role in respiration in the mammalian brain, and agents that affect respiratory function such as opioid peptides might have positive or negative neurotrophic effects during the development of these cholinergic connections. Rat fetal nerve cell cultures from developmental stages E14-E18 were established in 96-well plates from ventral forebrain (VFB), an area rich in cholinergic neurons, and from brainstem and rostral spinal cord, areas where respiratory control systems and cholinergic neurons co-exist. High affinity 3H-choline uptake was highest in E14 VFB cultures and decreased to 20% of this value by E16 and E18. Choline uptakes in E14 brainstem and spinal cord were only 20% and 13%, respectively, of E14 VFB uptake. A mu opioid receptor agonist, d-ala2-mePhe4-gly(ol)5]-enkephalin (DAMGO), was tested for its effect on somal area and neurite outgrowth in E16 cultures. Cholinergic neurons were identified by immunostaining with choline acetyltransferase antibody. DAMGO (10(-8) M) significantly decreased somal area in VFB cultures and spinal cord, but had no effect on somal area in brainstem. Naltrexone (10(-6) M) reversed this inhibition. Spinal cord cell neurite outgrowth was inhibited by DAMGO, and this inhibition was reversed by naltrexone. DAMGO had no significant effect on neurite length in VFB. Brainstem neurite length was paradoxically increased by both DAMGO and naltrexone. It was concluded that mu-selective opioid peptides inhibit growth of cultured cholinergic neurons in VFB and spinal cord, but not in the brainstem. There was no evidence for endogenous opioid activity in either VFB or spinal cord cultures.

Analgesics, Opioid↗

PubMed1999

Congenital hemihypertrophy and epithelioid haemangioendothelioma in a 10-year-old boy: a case report.

Epithelioid haemangioendothelioma has not been previously described in a patient with congenital hemihypertrophy and diabetes mellitus. Hepatic nodules were incidentally discovered on a routine US examination searching for known associated abnormalities. Pulmonary nodules were present on chest X-ray and CT of the lungs. The diagnosis was confirmed by open biopsy of a hepatic nodule. Despite significant disease progression the patient remains symptom free.

Child↗

PubMed1999

The pattern of infertility diagnoses in women of advanced reproductive age.

OBJECTIVE: Our intention was to determine whether there is a unique pattern of infertility diagnoses in older infertile couples. STUDY DESIGN: The design of this study was a retrospective chart review study. It was performed in a tertiary referral reproductive medicine unit. There were 2 groups of patients-couples: group 1, female partner aged 20-29 (n = 105) at presentation; group 2, female partner aged 40-45 (n = 112) at presentation. All women underwent infertility evaluations between 1989 and 1994. There were no interventions. The prevalence of standard infertility diagnoses was the main outcome measure. RESULTS: The prevalence of 8 major infertility diagnoses in the younger and older groups (each couple could have >/=1 diagnosis) was as follows: (1) ovulatory factor-younger group, 56%; older group, 30%; (2) tubal factor-younger group, 34%; older group, 29%; (3) endometriosis-younger group, 13%; older group, 17%; (4) uterine factor-younger group, 1%; older group, 5%; (5) cervical factor-younger group, 4%; older group, 1%; (6) luteal deficiency-younger group, 4%; older group, 10%; (7) male factor-younger group, 32%; older group, 45%; (8) unexplained-younger group, 5%; older group, 10%. The only significant difference was an increase in ovulatory factor in the younger group. CONCLUSIONS: There is no unique pattern of infertility diagnoses in women of advanced reproductive age as seen at a tertiary referral center. We speculate that a high false-positive rate associated with standard infertility tests and a different referral pattern for older couples obscures any real differences in the etiology of infertility in older couples.

Adult↗

PubMed1999

Intracerebral haemorrhage and cerebral amyloid angiopathy: CT features with pathological correlation.

AIMS: To review the computed tomography (CT) features of intracerebral haemorrhage pathologically proven to be associated with cerebral amyloid angiopathy in order to facilitate recognition of the presence of cerebral amyloid angiopathy in life. METHODS: We prospectively collected the clinical and brain imaging records of patients dying following an intracerebral haemorrhage who underwent a post-mortem examination and were found to have cerebral amyloid angiopathy. We reviewed the brain imaging to highlight features of the haemorrhage and of the rest of the brain common to these cases. RESULTS: Seven patients aged 60-86 years were examined over a 30-month period. On CT, the notable features were that the haemorrhages appeared large, lobar, often extended through the cortex to the subarachnoid space or into the ventricles, and were multiple and recurrent in patients who survived the initial bleed. The high density (blood) within the haematoma tended to sediment posteriorly. CONCLUSIONS: There are features on CT of cerebral amyloid angiopathy associated with spontaneous intracerebral haemorrhage which should raise the possibility of this underlying diagnosis. We suspect this condition is under-recognized in life, and should perhaps be considered more widely.

Aged↗

PubMed1999

Characterization of L-leucine transport system in brush border membranes from human and rabbit small intestine.

The branched-chain amino acids (BCAAs) leucine, isoleucine, and valine are beneficial to catabolic patients by improving hepatic protein synthesis and nitrogen economy, yet their transport from the intestinal lumen is not well-defined. The leucine transport system in human and rabbit small intestine was characterized using a brush border membrane vesicle (BBMV) model. Sodium and pH dependence and transport activity along the longitudinal axis of the small bowel were determined. Transport kinetics and inhibition profiles were defined. Although previous studies in other tissues show leucine transport to be mostly a Na+-independent process, our studies show that leucine transport is a predominantly Na+-dependent process occurring mainly via a single saturable pH-independent transporter resembling system B0 in the intestine. This system B0 transporter demonstrates stereoisomeric specificity. There is also a minor Na+-independent transport component (<6% in rabbits). Leucine uptake in both rabbits and humans is significantly greater than the uptake of other clinically relevant nutrients such as glutamine. In the rabbit, ileal leucine transport is significantly greater than jejunal uptake. While the affinities of the human and rabbit transporters are similar, the rabbit transporter has greater carrier capacity (maximal transport velocity [Vmax]). These findings suggest that the transport of leucine in the gut mucosa is significantly different from the transport in other tissues.

Amino Acids↗

PubMed1999

A novel Rieske iron-sulfur protein from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum: sequencing of the gene, expression in E. coli and characterization of the protein.

The crenarchaeon Pyrobaculum aerophilum is with an optimal growth temperature of 100 degrees C one of the most thermophilic organisms known to possess an aerobic respiratory chain. The analysis of DNA sequences from the Pyrobaculum genome project lead to the identification of an open reading frame potentially coding for a Rieske iron-sulfur protein. The complete gene (named parR) was cloned and sequenced. The deduced amino acid sequence displays unusual amino acid exchanges and a so far unknown sequence insertion. The N-terminus shows similarities to bacterial signal sequences. Several forms of the gene were expressed in E. coli in order to verify the classification as a Rieske protein and to facilitate biophysical studies. Soluble, thermo-stable proteins with correctly inserted iron-sulfur clusters were expressed from two versions of the gene. The delta1-23 truncated holo-protein is redox active. It displays the typical spectroscopic properties of a Rieske protein. The redox potential was determined to be +215 mV at pH 6.5 and is pH dependent above pH 7.5 revealing the influence of two protonation equilibria with pKa values of 8.1 and 9.8. Phylogenetic analysis demonstrates that the parR protein clusters together with the two other available archaeal Rieske sequences from Sulfolobus on a separate branch of the phylogenetic tree apart from the proteins from thermophilic bacteria like Aquifex and Thermus.

Amino Acid Sequence↗

PubMed1999

Renal nerve and alpha 2-adrenergic action during acute volume expansion in the anaesthetized rat.

1. The effects of renal denervation and alpha 2-adrenoceptor blockade on diuresis and natriuresis in response to acute volume expansion (VE) were investigated in anaesthetized rats. 2. In normal rats, a 30 min intravascular VE of up to 7.5% of bodyweight caused a significant diuresis and natriuresis. 3. Denervation enhanced the diuretic effect of VE by 39%, while alpha 2 blockade by 12 mumol/kg per h rauwolscine significantly inhibited the response by 61% in innervated kidneys and 43% in denervated kidneys. 4. Similar results were found for natriuresis in both innervated and denervated kidneys. 5. The results demonstrate that renal sympathetic nerve activity inhibits part of the increase in sodium and water excretion caused by VE, since denervation increased excretion under these conditions. The contribution of renal nerves to the control of renal function, however, decreases during the response to VE. 6. alpha 2-Adrenoceptor action enhanced the diuretic and natriuretic effects of VE by mechanisms otherwise unrelated to renal nerve activity.

Anesthesia↗

PubMed1999

Drinking, binge drinking, and other drug use among southwestern undergraduates: three-year trends.

This study examined substance use patterns and consequences in college students over a three year period. Students were surveyed at a large, southwestern university, allowing for a diverse sample that included a large percentage of minority respondents. Students (total N = 2710) in 1994, 1995, and 1996 responded anonymously to the Core Survey of Alcohol and Drugs. Over 80% of students at each time point were current drinkers, and over one-third at each time period reported binge drinking. Binge drinking was associated with greater weekly drinking and with a range of negative consequences. Underage drinking was prevalent at all time points, and underage drinkers reported drinking in a range of on- and off-campus situations. Hispanic students reported higher rates of binge drinking than other ethnic groups. Nonwhite, non-Hispanic students reported greater rates of abstinence than other students. Although other drug use was much less prevalent, drug use in combination with drinking was associated with more problematic patterns of drinking and more negative consequences. Results are discussed in terms of implications for interventions with college students.

Adult↗

PubMed1999

Functional expression of hMYH, a human homolog of the Escherichia coli MutY protein.

We have previously described the hMYH cDNA and genomic clones (M. M. Slupska et al., J. Bacteriol. 178:3885-3892, 1996). Here, we report that the enzyme expressed from an hMYH cDNA clone in Escherichia coli complements the mutator phenotype in a mutY mutant and can remove A from an A. 8-hydroxydeoxyguanine mismatch and to a lesser extent can remove A from an A. G mismatch in vitro.

8-Hydroxy-2'-Deoxyguanosine↗

PubMed1999

Direct selection for mutators in Escherichia coli.

We have constructed strains that allow a direct selection for mutators of Escherichia coli on a single plate medium. The plate selection is based on using two different markers whose reversion is enhanced by a given mutator. Plates containing limiting amounts of each respective nutrient allow the growth of ghost colonies or microcolonies that give rise to full-size colonies only if a reversion event occurs. Because two successive mutational events are required, mutator cells are favored to generate full-size colonies. Reversion of a third marker allows direct visualization of the mutator phenotype by the large number of blue papillae in the full-size colonies. We also describe plate selections involving three successive nutrient markers followed by a fourth papillation step. Different frameshift or base substitution mutations are used to select for mismatch-repair-defective strains (mutHLS and uvrD). We can detect and monitor mutator cells arising spontaneously, at frequencies lower than 10(-5) in the population. Also, we can measure a mutator cascade, in which one type of mutator (mutT) generates a second mutator (mutHLS) that then allows stepwise frameshift mutations. We discuss the relevance of mutators arising on a single medium as a result of cells overcoming successive growth barriers to the development and progression of cancerous tumors, some of which are mutator cell lines.

2-Aminopurine↗

PubMed1999

Radioaerosol scintigraphy in infants and children born to mothers with HIV disease. Pediatric Pulmonary and Cardiovascular Complications (of Vertically Transmitted Human Immunodeficiency Virus) Study Group.

PURPOSE: To determine the usefulness of technetium 99m diethyltriaminepentacetic acid (DTPA) radioaerosol inhalation-clearance scintigraphy for early detection of pulmonary complications of human immunodeficiency virus (HIV) disease in children. MATERIALS AND METHODS: A total of 301 studies were performed in 132 HIV-positive children (group 1; mean age, 46.6 months). In children born to HIV-positive mothers (group 2), 273 studies were performed in 160 children who eventually were proved to be HIV negative (mean age, 10.3 months), and 80 studies were performed in 47 HIV-positive children (mean age, 15.6 months). Radioaerosol studies were performed by using commercially available radioaerosol nebulizers. Pulmonary clearance half-time was measured by using conventional gamma camera computer systems. Radioaerosol results were correlated with indexes of pulmonary health and function. RESULTS: The HIV-negative, group 2 children had a mean radioaerosol clearance half-time (58.1 minutes; 162 studies in 108 children) similar to that reported in healthy adults. Group 1 children with pulmonary involvement exhibited a faster mean clearance half-time (28.6 minutes) than did children without evidence of pulmonary involvement from either group 1 or group 2 (P < .05). A faster pulmonary clearance rate did not simply reflect the presence of chest disease that also was detectable on radiographs (P = .3). CONCLUSION: Quantitative DTPA radioaerosol clearance studies may provide useful information about pulmonary involvement in selected children with HIV disease.

Administration, Inhalation↗