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Biomedical subjects

L Stewart

Publications and source records attributed to L Stewart.

At least 73 records · Page 4Linked to original sources
PubMed1998

Crystal structures of human topoisomerase I in covalent and noncovalent complexes with DNA.

Topoisomerases I promote the relaxation of DNA superhelical tension by introducing a transient single-stranded break in duplex DNA and are vital for the processes of replication, transcription, and recombination. The crystal structures at 2.1 and 2.5 angstrom resolution of reconstituted human topoisomerase I comprising the core and carboxyl-terminal domains in covalent and noncovalent complexes with 22-base pair DNA duplexes reveal an enzyme that "clamps" around essentially B-form DNA. The core domain and the first eight residues of the carboxyl-terminal domain of the enzyme, including the active-site nucleophile tyrosine-723, share significant structural similarity with the bacteriophage family of DNA integrases. A binding mode for the anticancer drug camptothecin is proposed on the basis of chemical and biochemical information combined with these three-dimensional structures of topoisomerase I-DNA complexes.

Antineoplastic Agents, Phytogenic↗
PubMed1998

A model for the mechanism of human topoisomerase I.

The three-dimensional structure of a 70-kilodalton amino terminally truncated form of human topoisomerase I in complex with a 22-base pair duplex oligonucleotide, determined to a resolution of 2.8 angstroms, reveals all of the structural elements of the enzyme that contact DNA. The linker region that connects the central core of the enzyme to the carboxyl-terminal domain assumes a coiled-coil configuration and protrudes away from the remainder of the enzyme. The positively charged DNA-proximal surface of the linker makes only a few contacts with the DNA downstream of the cleavage site. In combination with the crystal structures of the reconstituted human topoisomerase I before and after DNA cleavage, this information suggests which amino acid residues are involved in catalyzing phosphodiester bond breakage and religation. The structures also lead to the proposal that the topoisomerization step occurs by a mechanism termed "controlled rotation."

Amino Acid Sequence↗
PubMed1998

Laparoscopic repair of paraesophageal hiatal hernias.

BACKGROUND: Regardless of symptoms, paraesophageal hiatal hernias should be repaired in order to prevent complications. This study reports the University of California San Francisco experience with laparoscopic repair of paraesophageal hiatal hernias, emphasizing the technical steps essential for good results. PATIENTS AND METHODS: From May 1993 to September 1997, 55 patients, 27 women and 28 men, with a mean age of 67 years (range, 35-102 years) underwent laparoscopic repair of paraesophageal hernias at the University of California San Francisco. Symptoms, which had been present an average of 85 months before surgery, consisted mainly of pain (55%), heartburn (52%), dysphagia (45%), and regurgitation (41%). Of the four patients who presented with acute illness, two had gastric obstruction, one had severe dyspnea, and one had gastric bleeding. Endoscopy demonstrated esophagitis in 25 (69%) of 36 patients, and 24-hour pH-monitoring demonstrated acid reflux in 22 (67%) of 33 patients. Manometry detected severely impaired distal esophageal peristalsis in 17 (52%) of 33 patients. The preferred operation consisted of reduction of the hernia, excision of the sack and the gastric fat pad, closure of the enlarged hiatus without mesh, and construction of a fundoplication anchored by sutures within the abdomen. RESULTS: Of the 55 patients, the operations of 49 were completed laparoscopically using the following reconstructions: Guarner (270-degree) fundoplication (30 patients); Nissen fundoplication (10 patients); and gastropexy (9 patients). Five (9%) operations were converted to laparotomies. The average operating time was 219 minutes; the average blood loss was less than 25 mL; resumption of an unrestricted diet, 27 hours; and mean hospital stay, 58 hours. Intraoperative technical complications occurred in five (9%) patients. One patient died during surgery from a sudden pulmonary embolus. Two (4%) patients required a second operation for recurrent paraesophageal hernias. CONCLUSIONS: Laparoscopic repair of paraesophageal hiatal hernias is safe and effective, but the operation is difficult and good results hinge on details of the operative technique and the surgeon's experience. In this series, the crus could always be closed securely without using mesh. We realized early that a fundoplication should be a routine step, because it corrects reflux and is the best method to secure the gastroesophageal junction in the abdomen.

Adult↗
PubMed1998

Individual patient data meta-analysis in cancer.

As in many areas of health care, treatments for cancer may differ only moderately in their effects on major end points, such as death. But, such differences are worth knowing about, particularly in common diseases in which they could represent a substantial benefit to public health. Large-scale randomized evidence allows moderate differences to be investigated reliably, and one way to achieve this is by meta-analyses of updated and centrally collected individual patient data from all relevant trials. This paper illustrates why this form of research can often be important in cancer. It also offers the first list of such projects, as a source of information on current and past research in this area.

Humans↗
PubMed1998

beta-Galactosidase staining following intracoronary infusion of cationic liposomes in the in vivo rabbit heart is produced by microinfarction rather than effective gene transfer: a cautionary tale.

The myocardium is a potential target for the expression of exogenous genes to treat inherited and acquired diseases. Although adenovirus-mediated gene transfer has resulted in high-level gene transfer in vivo via direct intramyocardial injection and via a percutaneous intra-arterial route, the time-course of gene expression is limited by host immune responses. It was the aim of this study to test whether cationic liposome-mediated gene transfer, which does not suffer from the aforementioned problems, was feasible in the adult rabbit myocardium via a percutaneous transluminal approach. Doses of plasmid DNA encoding lacZ from 200-800 micrograms complexed to cationic liposomes resulted in X-gal conversion at day 3 with associated myocardial damage. We hypothesised that the damage was associated with macro-aggregates of cationic liposomes-DNA occluding the microcirculation. When such aggregates were excluded no X-gal conversion was seen in vivo. In order to show that X-gal conversion occurs in areas of infarction in the myocardium we caused closed chest infarction by deploying a platinum micro-embolisation coil in the circumflex coronary artery. At day 3 X-gal conversion was observed in the territory supplied by the occluded artery. Thus, microinfarction causes the false positive appearance of gene transfer when using a lacZ reporter gene.

Animals↗
PubMed1998

Endothelial cell transfection with cationic liposomes and herpes simplex-thymidine kinase mediated killing.

Endothelial cells are a promising target for cancer gene therapy because neoangiogenesis is vital for the supply of oxygen and nutrients to solid tumours. However, endothelial cells have been reported to be difficult to transfect. We demonstrate high rates of transfection with the reporter gene pSV40 beta gal using DC-Chol/DOPE cationic liposomes and lower rates with the novel polyamine cationic liposomes ACHx/DC-Chol/DOPE and ACO/DC-Chol/DOPE. Endothelial cells transfected with HSV-thymidine kinase using DC-Chol/DOPE demonstrated 3 log10 increased cytotoxicity compared with controls when exposed to the prodrug ganciclovir, thereby demonstrating significant biological effect.

Cations↗
PubMed1998

Culture shock and travelers.

As travel has become easier and more affordable, the number of people traveling has risen sharply. People travel for many and varied reasons, from the business person on an overseas assignment to backpackers seeking new and exotic destinations. Others may take up residence in different regions, states or countries for family, business or political reasons. Other people are fleeing religious or political persecution. Wherever they go and for whatever reason they go, people take their culture with them. Culture, like language, is acquired innately in early childhood and is then reinforced through formal and complex informal social education into adulthood. Culture provides a framework for interpersonal and social interactions. Therefore, the contact with a new culture is often not the exciting or pleasurable experience anticipated. When immersed in a different culture, people no longer know how to act when faced with disparate value systems. Contact with the unfamiliar culture can lead to anxiety, stress, mental illness and, in extreme cases, physical illness and suicide. "Culture shock" is a term coined by the anthropologist Oberg. It is the shock of the new. It implies that the experience of the new culture is an unpleasant surprise or shock, partly because it is unexpected and partly because it can lead to a negative evaluation of one's own culture. It is also known as cross-cultural adjustment, being that period of anxiety and confusion experienced when entering a new culture. It affects people intellectually, emotionally, behaviorally and physically and is characterized by symptoms of psychological distress. Culture shock affects both adults and children. In travelers or workers who have prolonged sojourns in foreign countries, culture shock may occur not only as they enter the new culture, but also may occur on their return to their original culture. Children may also experience readjustment problems after returning from leading sheltered lives in expatriate compounds. This readjustment back to their own culture after a period of time abroad has been termed "reverse culture shock, a condition which has been studied in both corporate managers and Peace Corps volunteers. With culture shock and many other processes of psychological adjustment, people tend to suffer alone, thinking that they are the only ones not coping well with their new circumstance. The objective of this paper was to bring the phenomenon of culture shock to the awareness of travel health advisors, who can in turn advise travelers, especially longer term travelers, about having realistic expectations of their travel and life in new cultures.

Acculturation↗
PubMed1998

Developing and implementing an intervention. Evaluation of an emergency department pilot on partner abuse.

This article discusses the role of formative and process evaluation in the development and implementation of a pilot intervention to improve the identification, treatment, and referral of women abused by their partners who present to an emergency department (ED). These evaluations were undertaken in conjunction with an outcome evaluation of training in and use of a five-step protocol of care piloted in a New Zealand public hospital. The outcome evaluation showed there was an improvement in identification and acute care of abused women following the intervention. The article highlights key factors that were relevant to the intervention's development and implementation, including social context, development processes, appropriateness for the setting, and level of support from key stakeholders. Factors identified as key to intervention effectiveness included its appropriateness for abused women and responsiveness to specific hospital, department, and staff needs. The key role of formative and process evaluation in the development and implementation of pilot interventions is highlighted, and the particular lessons gained from this study have relevance and application to other interventions.

Adolescent↗
PubMed1998

Role of surgical resection in the treatment of hypothalamic hamartomas causing precocious puberty. Report of six cases.

Hypothalamic hamartomas (HHs) are benign tumors that are often associated with central precocious puberty. Resection of HHs has been recommended as a treatment option for selected cases of pedunculated lesions, especially in young children. The role of surgery has to be evaluated in the light of the availability of effective medical treatment with gonadotropin-releasing hormone analogs (GnRHas). The authors report the long-term results of total resection of HHs in two children with central precocious puberty and compare it with medical management in four children. The two surgically treated patients underwent total resection of pedunculated HHs at 1.75 (Case 1) and 3.25 years (Case 2) and have received follow-up care for 9 and 8 years, respectively. There were no postoperative complications and computerized tomography scanning confirmed complete tumor removal in both patients. Both patients subsequently experienced some regression of secondary sexual characteristics. The response of luteinizing hormone to GnRH became prepubertal in one patient and was diminished in the other. However, the growth velocity remained elevated (> 7 cm/year), bone age remained advanced (> +2 standard deviations) 5 years after surgery, decreasing adult height prediction. In one child, GnRHa therapy was initiated 7 years postsurgery. Four children were treated solely with GnRH agonists and have received follow-up care for 2.3, 6, 9, and 9 years, respectively. These patients have had a complete regression of endocrinological abnormalities, including a normalization of growth velocity and reduction in the rate of skeletal maturation. No side effects were noted with decapeptyl treatment, and one child developed sterile abscesses while receiving Lupron-Depot. The proven efficacy of GnRHa in suppressing puberty and reducing bone age advancement leads the authors to advise against surgery as the initial management of central precocious puberty caused by HHs.

Child↗
PubMed1997

Reconstitution of human topoisomerase I by fragment complementation.

Human topoisomerase I (topo I, 91 kDa) is composed of four major domains; the unconserved and highly charged "N-terminal" domain (24 kDa), the conserved "core" domain (54 kDa), a poorly conserved and positively charged "linker" region (5 kDa), and the highly conserved "C-terminal" domain (8 kDa) which contains the active site tyrosine at position 723. Here we demonstrate that human topo I activity can be reconstituted by mixing a 58 kDa recombinant core domain (residues Lys175 to Ala659) with any one of a series of recombinant C-terminal fragments that range in size from 12 kDa (linker and C-terminal domains, residues Leu658 to Phe765) to 6.3 kDa (C-terminal domain residues Gln713 to Phe765). The C-terminal fragments bind tightly to the core domain, forming a 1:1 complex that is stable irrespective of ionic strength (0.01 to 1 M). The reconstituted enzymes are active only over a relatively narrow range of salt concentrations (25 to 200 mM KCl) as compared to the intact topo70 enzyme (missing the N-terminal domain). Under physiological conditions (150 mM KCl and 10 mM Mg2+) they are much more distributive in their mode of action than topo70. The reconstituted enzyme binds DNA with an affinity that is approximately 20-fold lower than that of the intact topo70. In addition, the cleavage/religation equilibrium of the reconstituted enzyme appears to be biased towards religation relative to that of the intact enzyme. Despite differences in the cleavage/religation equilibrium and affinity for DNA, the reconstituted and intact enzymes have identical sequence specificities for the cleavage of duplex DNA or suicide cleavage of oligonucleotide substrates.

Animals↗
PubMed1997

Improved parameters for the prediction of RNA hairpin stability.

Thermodynamic parameters are reported for hairpin formation in 1 M NaCl by RNA sequences of the type GGXANmAYCC, where XY is the set of four Watson-Crick base pairs and the underlined loop sequences are three to nine nucleotides. A nearest neighbor analysis of the data indicates the free energy of loop formation at 37 degrees C is dependent upon loop size and closing base pair. The model previously developed to predict the stability for RNA hairpin loops (n > 3) includes contributions from the size of the loop, the identity of the closing base pair, the free energy increment (deltaGo(37mm)) for the interaction of the closing base pair with the first mismatch and an additional stabilization term for GA and UU first mismatches [Serra, M. J., Axenson, T. J., & Turner, D. H. (1994) Biochemistry 33, 14289]. The results presented here allow improvements in the parameters used to predict RNA hairpin stability. For hairpin loops of n = 4-9, deltaGo(37iL)(n) is 4.9, 5.0, 5.0, 5.0, 4.9, and 5.5 kcal/mol, respectively, and the penalty for hairpin closure by AU or UA is +0.6 kcal/mol. deltaGo(37iL)(n) is the free energy for initiating a loop of n nucleotides. The model for predicting hairpin loop stability for loops larger than three becomes deltaGo(37L)(n) = deltaGo(37iL)(n) + deltaGo(37mm) + 0.6(if closed by AU or UA) - 0.7(if first mismatch is GA or UU). Hairpin loops of three are modeled as independent of loop sequence with deltaGo(37iL)(3) = 4.8 and the penalty for AU closure of +0.6 kcal/mol. Thermodynamic parameters for hairpin formation in 1 M NaCl for 11 naturally occurring RNA hairpin sequences are reported. The model provides good agreement with the measured values for both T(M) (within 10 degrees C of the measured value) and deltaGo(37) (within 0.8 kcal/mol of the measured value) for hairpin formation. In general, the nearest neighbor model allows prediction of RNA hairpin stability to within 5-10% of the experimentally measured values.

Models, Chemical↗
PubMed1997

Optimization of liposome mediated transfection of a neuronal cell line.

A cell line derived from sensory neurons was transfected with high efficiency using cationic liposomes, formulated from 3 beta [N-(N',N'-dimethylaminoethane)carbamoyl]-cholesterol (DC-Chol) and dioleoyl L-alpha-phosphatidylethanolamine (DOPE). This is the first time that cationic liposomes of this type have been reported to transfect a neuronal cell line. We used a reporter gene construct expressing beta-galactosidase under the control of the cytomegalovirus immediate early promoter and routinely observed transfection efficiencies > 40%. Parameters affecting transfection efficiency were examined and the ratio of DNA to liposome proved to be crucial. Liposome formulation procedures and cell transfection protocols devised here will be used as a basis for further in vivo and in vitro work.

Animals↗
PubMed1997

Topoisomerase I enhances TFIID-TFIIA complex assembly during activation of transcription.

The mechanism of coactivation by DNA topoisomerase I (topo I) was examined in a highly defined in vitro transcription system containing Pol II and purified factors. Both stimulation of the basal reaction and coactivation occurred dependent on TAF(II)s. Activation was first observed at the TFIID-TFIIA stage of initiation and maximal activation required the concomitant presence of TFIID, TFIIA, topo I, and activator. Electrophoretic mobility shift assay demonstrated a dramatic enhancement in the formation of the TFIID-TFIIA complex by topo I and activator, dependent on the TAF(II)s. DNase I footprinting confirmed this recruitment. A catalytically inactive topo I, which coactivated transcription, similarly stimulated the rapid formation of the TFIID-TFIIA complex in the presence of activator. A camptothecin-mediated DNA cleavage assay demonstrated the recruitment of topo I to the template by TFIID. Topo I likely functions during activation by enhancing the formation of an active TFIID-TFIIA complex on the promoter.

Camptothecin↗
PubMed1997

Approaches to preventing alcohol-related problems: the experience of New Zealand and Australia.

New Zealand and Australia have experienced similarities in their drinking cultures and patterns since colonization. This paper briefly reviews those experiences and looks at the approaches both countries have taken in dealing with alcohol-related problems. Over 10 years ago Australia developed a national strategy embracing a harm reduction approach and more recently New Zealand embarked upon a similar process. As well as examining the development of these, other initiatives are examined. These include approaches to licensing of alcohol sales, taxation, alcohol promotion, control of drinking on licensed premises, drinking and driving intervention, community action, initiatives by Maori and Aboriginal people and efforts to educate the individual drinker.

Journal Article↗
PubMed1997

The efficacy and effectiveness of process consultation in improving staff morale and absenteeism.

OBJECTIVES: The purpose of this randomized controlled trial was to test the efficacy and effectiveness of process consultation consisting of a series of nurse manager-consultant problem-solving meetings for leadership development that would lead to their staff's improved morale, quality of care, and reduced absenteeism. METHODS: Thirteen consenting clinical inpatient units were stratified for four variables known to affect outcome and were then randomly assigned to treatment and control conditions. The nurse managers from the seven experimental units were paired with outside nurse consultants from the McMaster University School of Nursing (Hamilton, Ontario, Canada) in a cooperative form of retraining in problem-solving through process consultation. Morale was determined through measures of perceptions of the work environment, an attitude scale concerning work (alienation), a personality measure (hardiness), and one of each scale for work satisfaction and for sources of satisfaction and dissatisfaction. Demographic data and information regarding family life responsibilities were collected as well. Assessment of quality of care was determined by the frequency and type of incident reports and by patients' perception of their satisfaction. A ratio of absence hours to total paid hours of work was used to compare experimental and control units' absenteeism rates. RESULTS: Experimental subjects reported a statistically significant improvement in the characteristics of their work setting and in the quality of working relationships. The consultation process facilitated a perceived change in the organizational context of the experimental hospital units with less centralization of authority and more clarity about expectation. These organizational changes were accompanied by improved working relationships and less alienation from work. There was no statistically significant difference in absenteeism. However, a subanalysis of the units by "dose" of the intervention identified those who benefited from the effects of the consultation. Subjects whose nurse managers participated more actively in the consultation process accounted for the changes in working relationships and perceived organizational changes. The "low-dose" experimental subjects were more like the control subjects, with the exception of absenteeism, for which they had higher rates. CONCLUSIONS: The findings of this study suggest that process consultation can alter the staff's perceptions of the characteristics of the work setting, can reverse negative attitudes (alienation), and can create a more positive and supportive working environment (improved working relationship). However, the results suggest the need to target this type of intervention to managers who have the personal resources to engage in self-evaluation and personal development and thus to participate in the consultative process.

Absenteeism↗