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Biomedical subjects

D Smith

Publications and source records attributed to D Smith.

At least 721 records · Page 40Linked to original sources
PubMed1992

Influence of metal ions on the ribonuclease P reaction. Distinguishing substrate binding from catalysis.

A high yield, photoactivated cross-linking reaction between a modified tRNA and RNase P RNA was used as a quantitative assay of substrate binding affinity. The cross-linking assay allows the effects of metal ions on substrate binding to be measured independently and in the absence of the pre-tRNA cleavage reaction. The results of this assay, in conjunction with the conventional cleavage assay, support the following conclusions about the nature of the RNase P RNA-tRNA binding interaction. (i) Monovalent cations act primarily to enhance enzyme-substrate binding, presumably by functioning as counterions. This enhancement can be attributed to a reduction in the tRNA off-rate. (ii) Although divalent cation is required for cleavage, the enzyme-substrate complex can form in the absence of divalent cation; the essential role of divalent cation in the reaction is thus catalytic. (iii) Ca2+ is as efficient as Mg2+ in promoting binding but supports catalysis only at a low rate.

Autoradiography↗
PubMed1992

Contributions of phylogenetically variable structural elements to the function of the ribozyme ribonuclease P.

Ribonuclease P (RNase P) is a ribonucleoprotein enzyme which participates in processing precursor tRNAs. The RNA subunit contains the catalytic site and is capable of catalysis in the absence of the protein subunit. RNase P RNAs from various eubacteria consist of a core of conserved sequence and secondary structure which is evolutionarily modified in different organisms by the presence of discrete helical elements at various sites in the RNAs. The variable occurrence of these helical elements suggests that they have no important functional role in the enzyme. The Escherichia coli RNase P RNA contains four such elements. It has been shown that simultaneous deletion of all four of them produces an RNA that is functional but has several significant defects which could arise from general disruption of the RNA or from the loss of element-specific functions. This paper describes a more detailed analysis of the role of the variable elements in E. coli RNase P RNA. Removal of one of the elements had no apparent effect on RNase P activity in vitro. Two other elements are required for correct folding of the RNA: their absence confers a requirement for extremely high monovalent salt concentrations, apparently to reduce intramolecular electrostatic repulsion. The fourth element that was tested participates in a long-range structural interaction (pseudoknot) which contributes to the structural stability of the enzyme and affects substrate binding affinity. In the absence of this helix, the RNA becomes temperature-sensitive, and the KM increases 100-fold.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1992

Urban black women's perceptions of breast cancer and mammography.

The purpose of this study was to examine differences in perceptions of breast cancer and mammography between black women who wanted a mammogram and those who did not. The subjects were 186 low socioeconomic black women who attended an inner city community health clinic (83% response rate). There were no significant differences on the demographic and background variables between women who did (N = 139) and did not (N = 47) want a mammogram. The knowledge level of both groups regarding breast cancer was poor. Those who desired a mammogram perceived themselves as more susceptible to breast cancer, and considered breast cancer more severe than those who did not want a mammogram. Neither group identified many barriers to obtaining a mammogram. The majority (at least 88 percent of those who wanted a mammogram and at least 55 percent of those who did not) agreed with each of the five benefit items. Eighty-five percent of both groups agreed they would receive a mammogram if their physician told them to do so. The two Health Belief Model components which accounted for the largest percentage of the variance between women who wanted a mammogram and those who did not were perceived benefits (13 percent) and perceived susceptibility (3 percent).

Adult↗
PubMed1992

Ventilatory mechanisms of exercise intolerance in chronic heart failure.

Mechanisms that have been suggested to underlie the abnormal ventilatory response to exercise in patients with chronic congestive heart failure (CHF) include high pulmonary pressures, ventilation-perfusion mismatching, early metabolic acidosis, and abnormal respiratory control. To evaluate the role that ventilation and gas exchange play in limiting exercise capacity in patients with CHF, data from 33 patients with CHF and 34 normal subjects of similar age who underwent maximal exercise testing were analyzed. Maximal oxygen uptake was higher among normal subjects (31.7 +/- 6 ml/kg/min) than among patients with CHF (17.7 +/- 4 ml/kg/min; p less than 0.001). The ventilatory equivalent for oxygen, expressed as a percentage of maximal oxygen uptake, was 25% to 35% higher among patients with CHF compared with normal subjects throughout exercise (p less than 0.01). A steeper component effect of ventilation on maximal oxygen uptake was observed among normal subjects compared with patients with CHF, which suggests that a significant portion of ventilation in CHF is wasted. Maximal oxygen uptake was inversely related to the ratio of maximal estimated ventilatory dead space to maximal tidal volume (VD/VT) in both groups (r = -0.73, p less than 0.001). Any given oxygen uptake at high levels of exercise among patients with CHF was accompanied by a higher VD/VT, lower tidal volume, and higher respiratory rate compared with normal subjects (p less than 0.01). Relative hyperventilation in patients with CHF started at the beginning of exercise and was observed both below and above the ventilatory threshold, which suggests that the excess ventilation was not directly related to earlier than normal metabolic acidosis. Thus abnormal ventilatory mechanisms contribute to exercise intolerance in CHF, and excess ventilation is associated with both a higher physiologic dead space and an abnormal breathing pattern. The high dead space is most likely due to ventilation-perfusion mismatching in the lungs, which is related to poor cardiac output, and the abnormal breathing pattern appears to be an effort to reduce the elevated work of breathing that is caused by high pulmonary pressures and poor lung compliance.

Adult↗
PubMed1992

Recombinant pseudorabies virus carrying a plasmodium gene: herpesvirus as a new live viral vector for inducing T- and B-cell immunity.

In Balb/c mice, the sterile protective immunity induced by immunization with radiation-attenuated Plasmodium yoelii sporozoites is eliminated by in vivo depletion of CD8+ T lymphocytes, suggesting that cytotoxic T lymphocytes (CTL) against malaria antigens expressed on infected hepatocytes are required for mediating this protective immunity. To produce a vaccine that would induce CTL against the P. yoelii circumsporozoite protein (CS), we constructed an attenuated pseudorabies virus (PRV) containing a gene encoding this protein. Balb/c mice that received three doses of 10(7) plaque-forming units (p.f.u.) of this vaccine intravenously at 3 week intervals developed high levels of antibodies to sporozoites (indirect fluorescent antibody titre = 4096) and CTL against a 16 amino acid epitope (SYVPSAEQILEFVKQI, amino acids 281-296) from the P. yoelii CS protein designated PYCTL1. The cytotoxic activity of the CTL was antigen-specific, MHC-restricted, and dependent on CD8+ T cells. Furthermore, these CTL eliminated P. yoelii-infected hepatocytes from in vitro culture, indicating that they recognize this peptide on the surface of infected hepatocytes. However, all nine mice that were challenged with 200 sporozoites developed a blood-stage malaria infection. We attribute this lack of protection to the great difficulty of inducing sterile immunity against this highly infectious parasite P. yoelii. We conclude that recombinant pseudorabies virus (PRV) worked successfully as a live vaccine vector to induce both antibodies and CTL, albeit non-protective in vivo, and the herpesviruses should be considered as subunit vaccines where T- and B-cell immunity is required.

Amino Acid Sequence↗
PubMed1992

Virulence of Mycobacterium tuberculosis for guinea pigs: a quantitative modification of the assay developed by Mitchison.

One of the more accepted methods of assay of virulence of tubercle bacilli is one developed by Mitchison in which guinea pigs were infected by the intramuscular route with 1.0 mg of tubercle bacilli freshly harvested from Lowenstein Jensen medium and in which virulence was based on a subjective score of the extent of gross disease in the animal 6 weeks after infection. Due to the practical difficulties involved in such an assay when routinely performed, the following modifications were made: frozen stocks of the culture were prepared prior to the date of infection, the inoculum was quantitated by means of colony forming units, and virulence was based on the number of tubercle bacilli recovered from the spleen of infected animals 6 weeks after infection. A significant correlation was obtained between the findings from the Mitchison assay of virulence and the modification. The latter assay is recommended for its quantitative and objective features.

Animals↗
PubMed1992

Multiple phantom limbs in a child.

This case report describes multiple phantom feet in a child after amputation of a leg. The subject is a 16-year-old girl who was born with a right leg 10 cm shorter than the left and who at the age of 6 was amputated below the right knee so that she could wear a prosthesis that would give her normal mobility. The girl reports that she subsequently experienced 2 phantom feet and 3 sets of phantom toes which have persisted to the present time. Each phantom has a distinct size, length and position in relation to the others and each is also the site of vivid sensations such as heat, tickle, and fatigue as well as voluntary and involuntary movement. She also describes sensations that resemble sensations experienced before the amputation: one of her phantom feet feels flat and locked into a forward position which corresponds with the actual shape and position of her congenitally deformed amputated foot. The implications of multiple phantoms are discussed with reference to recent concepts of phantom limbs.

Adolescent↗
PubMed1992

A comparison of two supplemental procedures for confirmation of antibody to hepatitis C virus c100-3 antigen in Louisiana blood donors.

In a pilot study designed to evaluate the performance of supplemental hepatitis C virus (HCV) tests, 146 consecutive HCV enzyme immunoassay (EIA)-reactive samples (0.98% of 14,949 donors) were comparatively evaluated with two sets of supplemental tests: HCV antibody neutralization/c100-3 peptide EIA and the first-generation HCV recombinant immunoblot assay (RIBA). Of these samples, 68.5 percent were positive and 17.8 percent were negative on both supplemental assays. Nineteen samples were discordant. Eleven samples were positive on one assay (9 on neutralization/peptide, 2 on RIBA) and negative or indeterminate on the alternate supplemental test, but reacted with two additional HCV antigens outside the c100-3 region in a second-generation dot immunoblot assay. The dot immunoblot assay was used as a reference and reactive samples were considered confirmed. The remaining eight discordant samples were indeterminate or negative on either assay and did not react on the dot immunoblot assay. These data indicate a 0.74-percent prevalence of HCV exposure detected by reactivity with the c100-3 antigen in blood donors in southern Louisiana.

Adolescent↗
PubMed1992

Decreased alpha 1-adrenergic receptors after experimental brain injury.

The magnitude of neuronal damage in central nervous system (CNS) injury may be related, in part, to alterations in the balance between excitatory and inhibitory neurotransmitters. Previous studies have implicated a role of central inhibitory noradrenergic mechanisms in the pathophysiologic sequelae of traumatic brain injury. In the present study, we examined alpha 1-adrenergic receptor binding after parasaggital lateral fluid percussion (FP) brain injury of moderate severity (2.3 atm) in the rat. At 30 min following injury, the specific binding of [3H]prazosin to membranes isolated from left cortex (injury site) was reduced by 37% in brain-injured animals when compared to sham-operated noninjured animals (p < 0.05). However, there were no significant differences in [3H]prazosin binding to membranes of either contralateral (right) cortex or left and right hippocampi between brain-injured and sham-operated animals. Conversely, at 24 h posttrauma, specific binding to membranes of left cortex, cortex adjacent to injury site, contralateral (right) cortex, and left hippocampus was reduced by 25%, 16%, 27%, and 24%, respectively (all p < 0.05). Scatchard analysis revealed that a reduction of [3H]prazosin binding to membranes of injured animals resulted from a decrease in alpha 1-receptor binding density (B-max) but not from changes in ligand affinity. Histopathologic assessment of neuronal damage at 24 h postinjury revealed neuronal loss within injury site cortex and left hippocampus but no clearly discernible cell loss in contralateral right cortex, suggesting that the decrease in B-max might be a consequence of early pathophysiology of trauma rather than of neuronal cell loss. We suggest that alterations in alpha 1-adrenergic receptors after brain injury may result in decreased inhibitory neurotransmitter action of norepinephrine and may thus contribute to the pathophysiology of traumatic brain injury.

Animals↗
PubMed1992

The novel 21-aminosteroid U74006F attenuates cerebral edema and improves survival after brain injury in the rat.

The present study evaluated the effect of the nonglucocorticoid 21-aminosteroid U74006F, an inhibitor of iron-dependent lipid peroxidation, on the development of regional cerebral edema after lateral fluid-percussion (FP) brain injury. Male Sprague-Dawley rats (n = 40) were anesthetized and subjected to FP brain injury of moderate severity centered over the left parietal cortex (2.5-2.6 atms). Fifteen minutes after brain injury, animals randomly received an i.v. bolus of either U74006F (3 mg/kg, n = 21) followed by a second bolus (3 mg/kg) at 3 hr or buffered sodium citrate vehicle (equal volume, n = 15). An additional group of 12 surgically prepared but uninjured animals served as preinjury controls. At 48 hr after injury, animals were sacrificed and brain tissue assayed for water content and regional cation concentrations. With the use of specific gravimetric techniques, no significant differences were observed in posttraumatic cerebral edema between drug- and control-treated animals. However, using wet weight/dry weight methodology, we found that administration of U74006F significantly reduced water content in the right hippocampus (contralateral to the site of injury) compared to saline-treated animals (p less than 0.05). U74006F also significantly prevented the postinjury increase in sodium concentrations in the ipsilateral hippocampus (p less than 0.05) and thalamus (p less than 0.03). Regional concentrations of potassium were unaltered after drug treatment. Administration of U74006F significantly reduced postinjury mortality, from 28% in control animals to zero in treated animals (p = 0.01). These results suggest that lipid peroxidation may be involved in the pathophysiological sequelae of brain injury and that 21-aminosteroids may be beneficial in the treatment of brain injury.

Animals↗
PubMed1992

Routine pelvic x-ray studies in awake blunt trauma patients: a sensible policy?

To evaluate the usefulness of routine pelvic x-ray films in the resuscitation of blunt trauma victims, 1395 patients were prospectively evaluated over a 13-month period. Of these, 810 (58%) were awake with Glasgow Coma Scale scores greater than or equal to 13 and were enrolled into the study. A history, with directed questions regarding pelvic pain, a clinical examination of the pelvis, and an anterior-posterior pelvic x-ray film (APPX) were obtained for each patient. Thirty-nine patients (5%) had fractures identified on the x-ray films. Of these patients with radiographically identified fractures, 34 (87%) complained of pain and had positive results on clinical examination, two (5%) either complained of pain or had positive results on examination and three (8%) had neither complaint of pain nor positive examination results. Of the 771 patients without fractures 743 (96%) lacked pain complaints or positive examination results. The likelihood of fracture was greatest in patients with complaints of pain and positive examination results (65%) followed by patients with either complaint of pain or positive examination results (16%). Only three (0.4%) of the 743 patients having no complaints of pain and a negative clinical examination had fractures diagnosed roentgenographically. These were minor fractures that did not affect the clinical course. Total charges incurred to diagnose pelvic fractures in this low-yield patient group were $88,028. We conclude that the practice of obtaining a screening APPX is not necessary or cost-effective in the management of awake blunt trauma patients who do not complain of pain and who have normal pelvic physical examination results.

Adolescent↗
PubMed1992

Comparison of four immunochemical methods for the measurement of oestrogen receptor levels in breast cancer.

Four methods of assessing oestrogen receptor (ER) status were compared in 33 patients with operable primary breast cancer. The methods used to assess the ER status were immunocytochemical assay (ER-ICA) of frozen sections, fine needle aspirates and imprint material and enzyme immunoassay (ER-EIA) of tumour tissue. A mean overall ER positivity of 45% (15 out of 33), 41% (13 out of 32) and 21% (six out of 29) was obtained by immunocytochemical (ER-ICA) staining of frozen sections, fine needle aspirates and tumour imprints, respectively, and a mean overall ER positivity of 42% (14 out of 33) was obtained by ER-EIA. The concordance of ER positivity in pairs of data obtained from different method combinations was found to range between 72 and 91%. However, statistically there was no significant difference between the four methods on the basis of the data obtained. Good comparability has been shown between the three tissue analyses and therefore the method of choice is technically not immediately apparent.

Biopsy, Needle↗
PubMed1992

Occurrence of a Photosystem II polypeptide in non-photosynthetic membranes of cyanobacteria.

Cytoplasmic and thylakoid membranes have been purified from the cyanobacteria Anacystis nidulans R2 and Phormidium laminosum by sucrose density gradient centrifugation. Probing of Western blots of proteins from these purified membrane fractions with antibodies directed against the 33 kDa polypeptide of Photosystem II from pea indicates that this protein is present in both the thylakoid and cytoplasmic membranes, rather than just the thylakoid membranes. This has been confirmed by immunogold labelling of cells. Oxygen evolution assays have been used to show that the 33 kDa polypeptide is not assembled into a functional Photosystem II complex in the cytoplasmic membranes. This may be due to the absence of other Photosystem II components.

Bacterial Proteins↗
PubMed1992

Maternal serum screening for Down's syndrome: the effect of routine ultrasound scan determination of gestational age and adjustment for maternal weight.

OBJECTIVE: To investigate the effect of using a routine ultrasound estimate of gestational age and maternal weight adjustment on maternal serum alpha-fetoprotein (AFP), unconjugated oestriol (uE3) and human chorionic gonadotrophin (hCG) levels in antenatal screening for Down's syndrome. DESIGN: Women with a singleton pregnancy without Down's syndrome were screened using the three serum markers and an estimate of gestational age based on 'dates' (time since first day of the last menstrual period) and one based on an ultrasound scan examination was recorded together with maternal weight. SETTING: Women attending the Homerton Hospital, Hackney, for their antenatal care between February 1989 and January 1990. SUBJECTS: 2113 women with a singleton pregnancy without Down's syndrome. RESULTS: The use of ultrasound to estimate gestational age (usually based on the biparietal diameter of the fetal skull) led to a significant reduction in the variance of each marker at a given week of pregnancy. The level of each marker was negatively associated with maternal weight, so that adjustment for weight also led to a reduction in variance. These data on gestational age and maternal weight, taken together with published data on pregnancies associated with Down's syndrome, indicate that the routine use of ultrasound to estimate gestational age will increase the detection rate from 58% to 67% while maintaining the false-positive rate at 5%, or reduce the false-positive rate from 5.7% to 3.1% while maintaining the detection rate at 60%. Routine maternal weight adjustment for the serum marker levels was much less useful, increasing the detection rate by about 0.5% for a given false-positive rate, or reducing the false-positive rate about 0.1% for a given detection rate. CONCLUSION: An ultrasound gestational age estimate available at the time of Down's syndrome screening confers a substantial advantage to screening performance with a further small benefit resulting from maternal weight adjustment, which is worth adopting if it can be done without difficulty or extra cost.

Biomarkers↗
PubMed1992

The pharmacokinetics of oral itraconazole in AIDS patients.

To test the hypothesis that absorption of orally administered itraconazole may be reduced in patients with the Acquired Immunodeficiency Syndrome (AIDS), 8 patients with AIDS were given two 100 mg capsules of itraconazole for 15 days. Plasma levels were measured by HPLC hourly for the first 8 h following ingestion, then at 24, 48, 72 and 96 h on days 1, 8 and 15. Peak plasma levels occurred after approximately 4 h. Mean peak plasma concentrations were 446 +/- 196 and 530 +/- 214 ng mL-1 at days 8 and 15. The area under the curve over 24 h (AUC0-24) was 2105 +/- 1241, 7679 +/- 3838 and 8748 +/- 4385 ng mL-1 h for days 1, 8 and 15, respectively. Steady-state was achieved after one week of dosing. There was no evidence of hepatotoxicity and one patient stopped itraconazole due to a rash. It may be concluded that the absorption of oral itraconazole capsules is reduced in AIDS patients, by a factor of approximately 50%, when compared with normal volunteers. AIDS patients may require higher doses of itraconazole than used in non-AIDS patients to achieve comparable plasma levels.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Comparison of nebuliser efficiency for aerosolizing pentamidine.

Inhaled pentamidine has become an important method of treatment and prophylaxis for Pneumocystis carinii pneumonia and we have compared nebuliser efficiency in terms of drug output and droplet sizes in four brands of jet nebuliser (Acorn-22, Inspiron, Cirrus, Respirgard II) and one brand of ultrasonic nebuliser (Fisoneb), at 2 pentamidine concentrations and 3 flow rates, using a laser particle sizer. Droplet size (which varied from 1.2 to 4.7 microns mass median diameter) was dependent in all cases, except with the Respirgard II system, on the flow rate of the gas driving the equipment and the concentration of pentamidine used. Drug output varied significantly between nebuliser brands and for a 300 mg dose of pentamidine was: 61% for the Acorn-22, 62% for the Inspiron, 49% for the Fisoneb and 43% for the Respirgard II. Both droplet size and drug output are important in determining nebuliser efficiency.

Aerosols↗
PubMed1992

Complex recognition site for the group I intron-encoded endonuclease I-SceII.

We have characterized features of the site recognized by a double-stranded DNA endonuclease, I-SceII, encoded by intron 4 alpha of the yeast mitochondrial COX1 gene. We determined the effects of 36 point mutations on the cleavage efficiency of natural and synthetic substrates containing the Saccharomyces capensis I-SceII site. Most mutations of the 18-bp I-SceII recognition site are tolerated by the enzyme, and those mutant sites are cleaved between 42 and 100% as well as the wild-type substrate is. Nine mutants blocked cleavage to less than or equal to 33% of the wild-type, whereas only three point mutations, G-4----C, G-12----T, and G-15----C, block cleavage completely. Competition experiments indicate that these three substrates are not cleaved, at least in part because of a marked reduction in the affinity of the enzyme for those mutant DNAs. About 90% of the DNAs derived from randomization of the nucleotide sequence of the 4-bp staggered I-SceII cleavage site are not cleaved by the enzyme. I-SceII cleaves cloned DNA derived from human chromosome 3 about once every 110 kbp. The I-SceII recognition sites in four randomly chosen human DNA clones have 56 to 78% identity with the 18-bp site in yeast mitochondrial DNA; they are cleaved at least 50% as well as the wild-type mitochondrial substrate despite the presence of some substitutions that individually compromise cleavage of the mitochondrial substrate. Analysis of these data suggests that the effect of a given base substitution in I-SceII cleavage may depend on the sequence at other positions.

Base Sequence↗