Biomedical subjects
Publications and source records attributed to M E Jones.
Screening patients or healthcare workers for Human Immunodeficiency Virus is unlikely to be cost-effective if the benefit is seen only as the prevention of infection of one group by the other within the healthcare institution. Overwhelmingly, the benefit of screening comes from interrupting the chains of infection within the ongoing epidemic in the population at large.
Human uridine monophosphate (UMP) synthase, a bifunctional protein containing orotate phsophoribosyltransferase (OPRTase, EC 2.4.2.10) and orotidine 5'-monophosphate decarboxylase (ODCase, EC 4.1.1.23) activities, has been overproduced by construction and use of a recombinant baculovirus containing the cDNA for this protein. Expression of the virus in cabbage looper larvae produces a crude larval homogenate having UMP synthase enriched about 180-fold over human placental homogenates and allows larger quantities of this human protein as well as analog proteins to be prepared for structure/function studies. A vastly improved purification procedure using a monoclonal immunoaffinity column was developed. Human UMP synthase purified from larval extracts yielded a product which comigrates in SDS gel electrophoresis with UMP synthase purified from human placenta; pure proteins prepared from these two tissue sources have the same specific activities. We found that OPRTase requires Pi ions in the assay buffers for optimal OPRTase activity; BSA in the assay vessel increases to a lesser degree both OPRTase and ODCase activities. These changes in the assay are essential to observe a parallel enrichment of the two enzyme activities. The baculovirus system was used to express human UMP synthase because it usually yields a product with posttranslational modifications that reflect those of the organism that provided the cDNA. We report data to show that human UMP synthase derived from either human placenta or larval extracts both have a sequence in which the N-terminal methionine has been removed and the formerly penultimate alanine has been acetylated.
The proteasome plays a central role in ubiquitin-dependent and -independent proteolysis in eukaryotic cells. The hawkmoth proteasome was purified from larval body wall and characterized with respect to substrate specificity, sensitivity to protease inhibitors, and cross-reactivity with monoclonal antibodies (mAbs) raised against human placenta proteasome. Leupeptin selectively inhibited the trypsin-like activity (T-L) and N-ethylmaleimide inhibited both T-L and chymotrypsin-like activities, whereas 0.02% sodium dodecyl sulfate stimulated the peptidylglutamyl peptide hydrolase, branched-chain amino acid preferring, and caseinolytic activities 20-, 18-, and 3.8-fold, respectively. All four peptidase activities were inhibited by 3,4-dichloroisocoumarin. One-dimensional immunoblot analysis showed that the level and subunit composition of the proteasome varied between tissues. The relative levels of proteasome were high in intersegmental muscle and ovary, lower in Malpighian tubule, male accessory gland, and ventral nerve cord, and lowest in flight muscle and fat body. The tissues differed in the relative amount of a 41-kDa doublet; a 22-kDa subunit was present only in the male accessory gland. Two-dimensional polyacrylamide gel electrophoresis showed that the hawkmoth proteasome contained at least 26 subunits, compared with 28 subunits in lobster. Immunological analysis using four subunit-specific mAbs identified the putative homologs of the human zeta, C2, C3, and C8 alpha-type subunits in the hawkmoth and lobster enzymes. Two of the four mAbs reacted with three or more of the hawkmoth subunits and three of the mAbs reacted with two or more of the lobster subunits. In addition, two other mAbs that recognize epitopes shared by a number of alpha-type subunits indicated that at least 15 (lobster) or 16 (hawkmoth) subunits were alpha-type. These results suggest that much of the subunit complexity of the arthropod proteasomes is a consequence of extensive post-translational modifications.
The intersegmental muscles (ISMs) of the tobacco hawkmoth Manduca sexta are a well-characterized model system for examining the biochemical changes that accompany programmed cell death during development. These giant muscles die during a 30-hr period in response to a decline in the circulating titer of the insect molting hormone 20-hydroxyecdysone. When the ISMs become committed to die, there are dramatic increases in both ubiquitin expression and ubiquitin-dependent proteolysis. Since the multicatalytic proteinase (MCP) is responsible for ATP/ubiquitin-dependent proteolysis in cells, we examined its composition and properties. The purified enzyme from whole larval integumentary tissues resembles MCPs isolated from other species with respect to subunit composition and general catalytic properties. However, when MCP was isolated from condemned ISMs, we observed an approximately ninefold increase in proteinase activity compared to MCP from precommitment muscles. This increase in proteolytic activity was correlated with an approximately eightfold increase in the absolute amounts of MCP protein as determined by Western blotting and densitometry. When purified MCP from condemned muscles was examined by two-dimensional polyacrylamide gel electrophoresis, four new subunits that were not detected in the precommitment muscles were present. Correlated with the addition of these new subunits was a dramatic increase in the levels of immunodetectable MCP throughout the cytoplasm and within the nuclei of dying muscles. These changes in MCP were regulated by the same hormonal signals that mediate cell death. These data are consistent with the hypothesis that when the ISMs become committed to die, more MCP accumulates in cells and new subunits are synthesized that change both the enzymatic properties and the conformation of MCP, which in turn participates in the dramatic proteolysis that accompanies cell death.
From 1990 to 1991 in the Hobart region there was a marked fall in both hotel patronage and the proportion of patrons subsequently driving with their blood alcohol concentration above the legal limit. This was associated with smaller falls in the number of drink drivers charged and alcohol-related road accidents, which continued in the following year. It appears that the pattern of drinking and driving is changing, presumably in response to random breath testing and tougher penalties for offences.
This study aimed to characterize the molecular basis of beta-lactamase induction in Citrobacter diversus. The chromosomal beta-lactamase encoding region from C. diversus, strain NF85, was cloned and expressed in Escherichia coli. The cloned region was sequenced and open-reading frames encoding a class A beta-lactamase, designated cdiA, and a putative LysR-type transcriptional regulator protein, divergently transcribed from the beta-lactamase gene and designated cdiR, were identified. The nucleotide sequence of the NF85 cdiA was identical to that of the published C. diversus ULA27 ampC sequence. A putative helix-turn-helix DNA-binding motif was located at the N-terminus of CdiR, and homology with enterobacterial AmpR proteins was noted. CdiR was demonstrated to bind to the C. diversus cdiAR intergenic region but not to the C. freundii ampCR intergenic region. A putative CdiR binding motif was identified in the cdiAR intergenic region. The cloned cdiAR region was inducible in E. coli strains SNO3 and HfrH. The inducible phenotype was dependent on the E. coli ampD and ampG gene products. We conclude that the molecular basis of inducible cdiA expression in C. diversus is similar to that of C. freundii ampC.
The development of leadership potential is recognized to be an essential component of nurse practitioner education to prepare graduates for a dynamic health care system. Leadership potential refers to those skills that enable a nurse practitioner to function as a change agent and to effectively resolve conflict. The purpose of the study was to examine personal variables (personality type, self-esteem, and learning style) with respect to the leadership potential of pediatric nurse practitioner graduates over a 3-year period. The research study was descriptive, and data were analyzed using descriptive statistics. The subjects consisted of 21 female students who completed five instruments used to measure the relationships between variables and leadership potential. Significant results were found between those students who learned abstractly and the ability to function effectively as a change agent post-program.
This study examines the importance of left ventricular (LV) dilatation, and evidence of multivessel coronary artery disease identified on thallium-201 (TI-201) single-photon emission computed tomographic (SPECT) scintigrams, for predicting long-term outcome in patients with an extensive left anterior descending (LAD) perfusion deficit. Impaired contractility of the left ventricle determined by low ejection fraction, elevated LV end-systolic volume, and dilatation of the left ventricle are known as major predictors of mortality after myocardial infarction. TI-201 single-photon emission computed tomography primarily reveals status of perfusion/redistribution, but also contains indirect information on LV function. To date, there are no TI-201 SPECT data on impaired function of the left ventricle (LV dilatation) and extent of perfusion deficits, discussed together as correlates of survival. Patient data were prospectively collected in the computer data base at Emory University. A large perfusion defect involving more than one third of the LAD territory was identified in 291 of 2,652 consecutive patients examined with TI-201 SPECT initial and 3-hour redistribution studies. Follow-up data were obtained for 284 patients (98%) at 38 +/- 14 months. Of the 291 patients, 58 died. The most powerful multivariate correlates of death were LV dilatation, multivessel disease, and the ratio of the LAD severity stress score to total severity of SDs stress score. Cox model analysis was used to determine correlates of survival. Three-year survival for patients with LV dilatation was 73% versus 89% without LV dilatation (p < 0.001). Three-year survival in patients with 1-vessel disease ("LAD only") was 94% versus 78% for multivessel disease (p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)
Cells growing in culture are subject to mutation, and as mutation is a random event, the number of mutants in a culture will be a random variable. The size of the clone of mutants arising from a single mutational event depends on the timing of the mutation; the earlier the mutation the larger the corresponding clone of mutants. The frequency with which new mutations arise may be estimated from examining the number of mutants found in a number of parallel cultures, each culture arising from a single cell. An efficient estimator of mutation rate in such an experiment is a maximum likelihood estimator. The use of such an estimator presupposes knowledge of the probability distribution for the number of mutants to be detected in a culture, given the mutation rate. In turn this depends on the probability distribution for the size of the clone of detected mutants arising from any single mutation. This latter distribution depends on the relative cell cycle time, tau, of the mutants, and on the probability, s, that a mutant which exists will be detected. This paper develops the required probability distribution.
This study reports on an intervention programme to promote responsible drinking in hotels. The licensees of eight hotels agreed to participate in a trial of measures designed to assist patrons to avoid drink-driving, and seven other hotels were used as controls. The interventions acceptable to licensees comprised commercial-quality promotional material with the theme "0.05 Know Your Limits", and a breath analysis machine and poster on its use. Patrons leaving the hotels on Thursday, Friday and Saturday nights were interviewed and breath-tested. Although the intervention material had been seen by one-third of patrons in the intervention hotels, there was no significant difference between them and control hotel patrons in either median BAC or the proportion who were going to drive with BAC over the legal limit. There was poor compliance by hotels with the intervention procedures, indicating that a major impediment to the implementation and evaluation of programmes to promote responsible drinking is a lack of motivation by many licensees, despite support by some licensees and the Australian Hotels Association.
Luria and Delbrück, in a seminal paper, introduced fluctuation analysis primarily as a means to elucidate the timing of mutation in relation to the imposition of selective conditions. Their work, and subsequently that of LEA and COULSON, established also a basis for measuring the frequency of mutational events. The several estimators proposed by these authors differ both in complexity and in efficiency, and the published literature relies mainly on the less efficient but computationally trivial estimators. The estimators as originally proposed assume that all mutants occurring in culture will be counted in the subsequent assay, but a relaxation of this assumption suggests an alternative experimental design and alternative estimators which offer advantages over those currently in common use.
We examined the negative relation between temperature and the sudden infant death syndrome (SIDS) in 22 communities in seven countries. We estimated the percentage increase in SIDS rate for a 1 degree C drop in climatic temperature. The relation differed substantially among communities. In New Zealand and Australia (10 communities), the association was consistently strong; in Europe (seven communities), it varied from strong to weak; and in the USA (five communities), it was moderate or weak. We postulate that low climatic temperature indirectly increases the incidence of SIDS, particularly in countries where outdoor climatic temperature modifies the indoor temperature and clothing habits.
Considerable biochemical diversity and polynucleotide sequence variation have been reported amongst strains of Citrobacter spp. However, sequence heterogeneity has not been investigated at gene loci of clinical relevance. In this study, sequence heterogeneity in the beta-lactamase structural gene, ampC, amongst 91 clinical isolates of Citrobacter spp. that showed resistance to various third-generation cephalosporins was investigated. Variation was examined by high-stringency polymerase chain reactions (PCR) with primers homologous to the known ampC sequences of C. freundii strains OS60 and I113, and C. diversus NF85. If an isolate contained an ampC gene homologous to one of these three characterised ampC genes, a single PCR band of a predictable size was generated with the appropriate primer set; 50 (60%) of isolates gave a PCR product of the expected size with the OS60 primer set and nine (10%) gave a product with the I113 primer set. All these 59 isolates were identified as C. freundii by API-20E strips. Six isolates (7%) gave a product with the C. diversus NF85 primer set but only four of these were identified as C. diversus in API-20E tests; the other two isolates were identified as C. freundii. Of the 91 isolates, 28 (31%), were identified as either C. freundii or C. diversus, but gave no PCR product with any primer set tested. Five of these showed no homology to any of the reference strain ampC PCR products in hybridisation tests. Nevertheless, all showed beta-lactamase activity. Overall, this method allowed the identification of novel ampC gene loci, which may serve as a basis for the identification of Citrobacter spp. rapidly at a molecular level.
We have tested the hypothesis that the composition of alveolar surfactant varies with pattern of breathing and level of fitness. We examined three major components of surfactant, surfactant protein A (SP-A), disaturated phospholipids (DSP), and cholesterol (CHOL) in bronchoalveolar lavage (BAL) fluid from 12 healthy men before and after exercise. Fitness was assessed as work load/heart rate ([kpm.min-1]/[HR.HRmax-1]) achieved during cycling for 30 min at 90% theoretical maximal heart rate. Using a bronchoscope, four 20-ml vols of 0.15 M NaCl at 37 degrees C were instilled and then recovered from first a right upper and then a right lower lobe segmental bronchus. As we found no differences in the BAL from upper and lower lobes, the fluid was combined. We found a direct relationship between CHOL and DSP (rs = 0.84, p < 0.001), SP-A and CHOL (rs = 0.40, p < 0.025), and between SP-A and DSP (rs = 0.44, p < 0.025). The change in the ratios CHOL/DSP, SP-A/CHOL, and SP-A/DSP immediately after exercise was correlated with fitness (rs = -0.56, p < 0.025; rs = 0.75, p < 0.005; rs = 0.62, p < 0.025, respectively). We conclude that the composition of surfactant can change rapidly with exercise in a manner related to fitness, and we suggest that this is consistent with the existence of at least two pools of tissue surfactant of different composition supplying the alveolar compartment.
As one ages, sensory discrimination in the oral cavity progressively diminishes, and dysphagia and aspiration are more likely to occur. Whether similar age-related laryngeal and pharyngeal sensory abnormalities exist and contribute to dysphagia and aspiration is unknown. The purpose of this study was to determine if sensory discrimination in the area innervated by the superior laryngeal nerve diminishes with increasing age. By applying a previously described new device and technique that utilizes brief air pulse stimulation of the anterior wall of the pyriform sinus, sensory discrimination can be reliably determined. We carried out 672 trials in 56 healthy adults divided into three age groups: 20 to 40, 41 to 60, and 61 to 90 years of age. Overall, the average sensory discrimination was 2.30 +/- 0.50 mm Hg. In subjects 20 to 40 years of age, sensory discrimination was 2.07 +/- 0.20 mm Hg, while in subjects 61 to 90 years of age, sensory discrimination was 2.68 +/- 0.63 mm Hg (p < .05). There also was a statistically significant difference between the 41- to 60-year and 61- to 90-year age groups (p < .05). Progressive diminution in pharyngeal and supraglottic sensitivity with increasing age might be a contributing factor in the development of dysphagia and aspiration in the elderly.
There is a popular misconception that the meniscus is avascular and that tears of the meniscus do not heal. Over the years this notion has gradually been dispelled as being incorrect and the fact that the periphery of the meniscus is in fact vascular has been accepted. Attempts to resuture this particular structure in the knee have, in some cases, proved to be very successful.
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During development, large numbers of cells die by a nonpathological process referred to as programmed cell death. In many tissues, dying cells display similar changes in morphology and chromosomal DNA organization, which has been termed apoptosis. Apoptosis is such a widely documented phenomenon that many authors have assumed all programmed cell deaths occur by this process. Two well-characterized model systems for programmed cell death are (i) the death of T cells during negative selection in the mouse thymus and (ii) the loss of intersegmental muscles of the moth Manduca sexta at the end of metamorphosis. In this report we compare the patterns of cell death displayed by T cells and the intersegmental muscles and find that they differ in terms of cell-surface morphology, nuclear ultrastructure, DNA fragmentation, and polyubiquitin gene expression. Unlike the T cells, which are known to die via apoptosis, we find that the intersegmental muscles display few of the features that characterize apoptosis. These data suggest that more than one cell death mechanism is used during development.