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Biomedical subjects

W J Sullivan

Publications and source records attributed to W J Sullivan.

At least 19 recordsLinked to original sources
PubMed2002

The importance of patient autonomy at birth.

It has always been recognised that the practice of medicine requires an ethical base. This ethical base also provides a backbone for a legal framework. Until recently paternalism was the accepted norm in the physician-patient relationship. Based on knowledge and experience the physician decided what treatment was in the best interests of the patient. However, in recent years medicine has changed from a predominantly paternalistic profession to one that is more patient centred. The physician informs and advises the patient but it is the patient who makes the decision. This is reflected in the change to principle-based medical ethics. The central concept in principle-based medical ethics is patient autonomy. This paper explores the application of autonomy and the legal requirement of informed consent to the clinical practice of obstetric anaesthesia. Autonomy requires that a patient has capacity to make a decision. Whether labouring women have capacity is discussed. There are times when patients are not able to act autonomously and the roles of advance directives (which can include birth plans) and substitute decision makers become important. The application of ethical practice to research in obstetric anaesthesia is considered.

Journal Article↗
PubMed2000

Incidence of intravascular uptake in lumbar spinal injection procedures.

STUDY DESIGN: Multicenter, prospective, observational study. OBJECTIVES: To document the incidence of and factors associated with intravascular uptake during lumbar spinal injection procedures. SUMMARY OF BACKGROUND DATA: In prior reports, the incidence of inadvertent intravascular needle placement during contrast-enhanced, fluoroscopically guided lumbar spinal injection procedures has been incidentally noted to range from 6.4% to 9.2%. We present the first systematic prospective documentation of intravascular uptake of contrast dye during different types of lumbar injection procedures. METHODS: Fifteen interventional spine physicians in seven centers recorded data regarding intravascular uptake during 1219 contrast-enhanced, fluoroscopically guided lumbar spinal injection procedures. RESULTS: The overall incidence of intravascular uptake during lumbar spinal injection procedures as determined by contrast enhanced fluoroscopic observation is 8.5%. Caudal and transforaminal routes have the highest rates at 10.9% and 10.8%, respectively, followed by zygapophyseal joint (6.1%), sacroiliac joint (5.3%), and translaminar (1.9%) injections. Intravascular uptake is twice as likely to occur in those patients over rather than under 50 years of age. Preinjection aspiration failed to produce a flashback of blood in 74% of cases that proved to be intravascular upon injection of contrast dye. CONCLUSION: The incidence of intravascular uptake during lumbar spinal injection procedures is approximately 8.5%. The route of injection and the age of the patient greatly affect this rate. Absence of flashback of blood upon preinjection aspiration does not predict extravascular needle placement. Contrast-enhanced, fluoroscopic guidance is recommended when doing lumbar spinal injection procedures to prevent inadvertent intravascular uptake of injectate.

Adolescent↗
PubMed2000

Cloning and characterization of a novel histone acetyltransferase homologue from the protozoan parasite Toxoplasma gondii reveals a distinct GCN5 family member.

In an effort to identify gene products involved in transcriptional regulation in apicomplexan parasites, the Toxoplasma gondii expressed sequence tag (EST) database was examined for sequences containing similarity to known transcriptional components. One EST (dbEST ID #466792) exhibited strong similarity to yeast GCN5 and other histone acetyltransferases (HATs). Primers were designed based on the EST sequence and used to amplify an 850 bp fragment (containing an intron) from T. gondii genomic DNA which was used to identify four cDNA clones from a tachyzoite cDNA library. The complete open reading frame (ORF) of 3.5 kb was elucidated using 5' RACE and genomic sequence. The deduced amino acid sequence of the coding region shows that the C-terminal domain possesses unequivocal similarity to GCN5 family members. However, unlike other lower eukaryotes, T. gondii GCN5 has an extended N-terminal domain similar in length, but not in composition, to metazoan HAT proteins. These features distinguish T. gondii GCN5 as a novel member of the GCN5 family. A portion of the cDNA sequence was used as a probe to isolate three overlapping clones from a T. gondii genomic library, generating a approximately 7.5 kb map of the GCN5 locus which contains seven exons separated by six introns. Southern analysis verifies the predicted map and suggests that a similar locus may be present elsewhere in the genome.

Acetyltransferases↗
PubMed1999

The adenosine transporter of Toxoplasma gondii. Identification by insertional mutagenesis, cloning, and recombinant expression.

Purine transport into the protozoan parasite Toxoplasma gondii plays an indispensable nutritional function for this pathogen. To facilitate genetic and biochemical characterization of the adenosine transporter of the parasite, T. gondii tachyzoites were transfected with an insertional mutagenesis vector, and clonal mutants were selected for resistance to the cytotoxic adenosine analog adenine arabinoside (Ara-A). Whereas some Ara-A-resistant clones exhibited disruption of the adenosine kinase (AK) locus, others displayed normal AK activity, suggesting that a second locus had been tagged by the insertional mutagenesis plasmid. These Ara-A(r) AK+ mutants displayed reduced adenosine uptake capability, implying a defect in adenosine transport. Sequences flanking the transgene integration point in one mutant were rescued from a genomic library of Ara-A(r) AK+ DNA, and Southern blot analysis revealed that all Ara-A(r) AK+ mutants were disrupted at the same locus. Probes derived from this locus, designated TgAT, were employed to isolate genomic and cDNA clones from wild-type libraries. Conceptual translation of the TgAT cDNA open reading frame predicts a 462 amino acid protein containing 11 transmembrane domains, a primary structure and membrane topology similar to members of the mammalian equilibrative nucleoside transporter family. Expression of TgAT cRNA in Xenopus laevis oocytes increased adenosine uptake capacity in a saturable manner, with an apparent K(m) value of 114 microM. Uptake was inhibited by various nucleosides, nucleoside analogs, hypoxanthine, guanine, and dipyridamole. The combination of genetic and biochemical studies demonstrates that TgAT is the sole functional adenosine transporter in T. gondii and a rational target for therapeutic intervention.

Adenosine↗
PubMed1999

Recombinant expression, purification, and characterization of Toxoplasma gondii adenosine kinase.

Toxoplasma gondii lacks the capacity to synthesize purines de novo, and adenosine kinase (AK)-mediated phosphorylation of salvaged adenosine provides the major route of purine acquisition by this parasite. T. gondii AK thus represents a promising target for rational design of antiparasitic compounds. In order to further our understanding of this therapeutically relevant enzyme, an AK cDNA from T. gondii was overexpressed in E. coli using the pBAce expression system, and the recombinant protein was purified to apparent homogeneity using conventional protein purification techniques. Kinetic analysis of TgAK revealed Km values of 1.9 microM for adenosine and 54.4 microM for ATP, with a k(cat) of 26.1 min(-1). Other naturally occurring purine nucleosides, nucleobases, and ribose did not significantly inhibit adenosine phosphorylation, but inhibition was observed using certain purine nucleoside analogs. Adenine arabinoside (AraA), 4-nitrobenzylthioinosine (NBMPR), and 7-deazaadenosine (tubercidin) were all shown to be substrates of T. gondii AK. Transgenic AK knock-out parasites were resistant to these compounds in cell culture assays, consistent with their proposed action as subversive substrates in vivo.

Adenosine Kinase↗
PubMed1999

Insertional tagging of at least two loci associated with resistance to adenine arabinoside in Toxoplasma gondii, and cloning of the adenosine kinase locus.

A genetic approach has been exploited to investigate adenylate salvage pathways in the protozoan parasite Toxoplasma gondii, a purine auxotroph. Using a new insertional mutagenesis vector designed to facilitate the rescue of tagged loci even when multiple plasmids integrate as a tandem array, 15 independent clonal lines resistant to the toxic nucleoside analog adenine arabinoside (AraA) were generated. Approximately two-thirds of these clones lack adenosine kinase (AK) activity. Parallel studies identified an expressed sequence tag (EST) exhibiting a small region of weak similarity to human AK, and this locus was tagged in several AK-deficient insertional mutants. Library screening yielded full-length cDNA and genomic clones. The T. gondii AK gene contains five exons spanning a approximately 3 kb locus, and the predicted coding sequence was employed to identify additional AK genes and cDNAs in the GenBank and dbEST databases. A genomic construct lacking essential coding sequence was used to create defined genetic knock-outs at the T. gondii AK locus, and AK activity was restored using a cDNA-derived minigene. Hybridization analysis of DNA from 13 AraA-resistant insertional mutants reveals three distinct classes: (i) AK-mutants tagged at the AK locus; (ii) AK- mutants not tagged at the AK locus, suggesting the possibility that another locus may be involved in regulating AK expression; and (iii) mutants with normal AK activity (potential transport mutants).

Adenosine Kinase↗
PubMed1999

Transport and trafficking: Toxoplasma as a model for Plasmodium.

Like Plasmodium, the protozoan parasite Toxoplasma gondii is a member of the phylum Apicomplexa, and an obligate intracellular pathogen. Unlike Plasmodium, however, Toxoplasma is highly amenable to experimental manipulation in the laboratory. The development of molecular transformation protocols for T. gondii has provided both scientific precedent and practical selectable markers for Plasmodium. Beyond the feasibility of molecular biological experimentation now possible in both systems, the high frequency of stable transformation in Toxoplasma allows this parasite to be used for molecular genetic analysis. The ability to control homologous vs. non-homologous recombination in T. gondii permits gene knockouts/allelic replacements at previously cloned loci, and saturation insertional mutagenesis of the entire parasite genome (and cloning of the tagged loci). T. gondii also exhibits unusual ultrastructural clarity, facilitating cell biological analysis. The accessibility of Toxoplasma as an experimental system allows this parasite to be used as a surrogate for asking many questions that cannot easily be addressed in Plasmodium itself. T. gondii also serves as a model system for genetic exploration of parasite biology and host-parasite interactions. Success stories include: biochemical analysis of antifolate resistance mechanisms; pharmacological studies on the mechanisms of macrolide activity; genetic identification of nucleobase/nucleoside transporters and metabolic pathways; and cell biological characterization of the apicomplexan plastid. As with any model system, not all questions of interest to malariologists can be addressed in Toxoplasma; differentiating between sensible and foolish questions requires familiarity with the biological similarities and differences of these systems.

Animals↗
PubMed1999

Advance directives: a third option.

As autonomy can only be exercised when a person is competent, advance directives give the potential to extend autonomy during incapacity. Most advance directives specify when intervention measures are not to be used. The elderly, or those with a fatal or chronic illness commonly give these directions. This type of advance directive does not satisfy he concerns of the younger and healthier segment of the population, and may be unsuitable for the older person. Healthy people are less concerned with procedures; they are concerned with outcome. All groups are concerned with certainty. These concerns may be better addressed by combining an instructional and a proxy directive, and for the healthy, by emphasizing ends rather than means. An example of such a directive is proposed.

Advance Directives↗
PubMed1998

Cardiovascular disease in patients with dysvascular amputation.

Cardiovascular diseases are the most common causes of morbidity and mortality in individuals with peripheral vascular disease (PVD). Among patients who have undergone lower extremity amputation as a result of PVD, the prevalence of concomitant cardiovascular disease may be as high as 75%. Comorbid heart disease may complicate the postamputation course of recovery, delay initiation of rehabilitation training, and inhibit the achievement of maximal functional independence. A variety of methods have been used to assess cardiac status and risk in amputation patients undergoing physical training; these have included clinical evaluation, resting electrocardiography, and continuous dynamic electrocardiography during either standard physical therapy exercise or adapted ergometry. Several conditioning training programs have been developed to improve the cardiovascular fitness of patients with dysvascular amputation, the results of which have been favorable. These assessment and intervention strategies have extensive applicability in the clinical management of patients with dysvascular amputation.

Amputation, Traumatic↗
PubMed1997

Tagging genes and trapping promoters in Toxoplasma gondii by insertional mutagenesis.

Plasmid vectors that incorporate sequence elements from the dehydrofolate reductase-thymidylate synthase (DHFR-TS) locus of Toxoplasma gondii integrate into the parasite genome with remarkably high frequency (>1% of transfected parasites). These vectors may-but need not-include mutant DHFR-TS alleles that confer pyrimethamine resistance to transgenic parasites. Large genomic constructs integrate at the endogenous locus by homologous recombination, but cDNA-derived sequences lacking long stretches of contiguous genomic DNA (due to intron excision) typically integrate into chromosomal DNA by nonhomologous recombination. Nonhomologous integration occurs effectively at random; and coupled with the high frequency of transformation, this allows a large fraction of the parasite genome to be tagged in a single electroporation cuvette. Genomic tagging permits insertional mutagenesis studies conceptually analogous to transposon mutagenesis in bacteria, yeast, Drosophila, etc. In theory (and, thus far, in practice), this allows identification of any gene whose inactivation is not lethal to the haploid tachyzoite form of T. gondii and for which a suitable selection or screen is available. Transformation vectors can be engineered to facilitate rescue of the tagged locus and to include a variety of reporters or selectable markers. Genetic strategies are also possible, using reporters whose function can be assayed by metabolic, visual, or immunological screens to "trap" genes that are activated (or inactivated) under various conditions of interest.

Animals↗
PubMed1971

Potassium and sodium transport across single distal tubules of Amphiuma.

The transport properties of potassium (K) and sodium (Na) were studied in single distal tubules of Amphiuma using free-flow micropuncture techniques and stationary microperfusion methods. The transepithelial movement of labeled potassium was measured utilizing a three-compartment system in series in which the time course of tracer disappearance from the lumen was followed. Under control conditions, in blood- and doubly-perfused kidneys, extensive active net reabsorption of sodium and potassium obtains along single distal tubules. Tubular potassium reabsorption is abolished by ouabain at a concentration of 5 x 10(-6)M. Significant net secretion of K can be induced by exposing Amphiuma to a high K environment (100 mM KCl) or by adding acetazoleamide (1 x 10(-4)M) to the perfusion fluid. Transepithelial movement of potassium involves mixing with only a small fraction of total distal tubular cell potassium. This transport pool of potassium increases significantly with the transition from tubular net reabsorption to net secretion. Indirect evidence is presented which indicates that increased active K uptake across the peritubular cell boundary may be of prime importance during states of net K secretion.

Acetazolamide↗