Biomedical subjects

S N Smith

Publications and source records attributed to S N Smith.

At least 19 recordsLinked to original sources

PubMed2004

Status epilepticus after the initial diagnosis of epilepsy in children.

OBJECTIVES: To determine the risk and predictors of status epilepticus in children after they have been diagnosed with epilepsy. METHODS: In a prospective community-based cohort study of 613 children, the occurrence of status epilepticus after the initial diagnosis of epilepsy was ascertained. Parents were called every 3 months, and interval medical records were reviewed every 6 months. Predictors of primary interest included a history of status before the diagnosis of epilepsy, age at onset, underlying etiology, and epilepsy syndrome. Data were analyzed with chi2 tests, Kaplan-Meier analyses, and Cox proportional hazards models. RESULTS: Of 613 children followed a median of 8.0 years, 58 (9.5%) had > or =1 episode of status epilepticus during follow-up evaluation. The first episode occurred a median of 2.5 years after initial diagnosis (range, <1 month to 8.8 years). A history of previous status epilepticus was strongly associated with subsequent status epilepticus (18/56 [32.1%] vs 40/557 [7.2%]; p < 0.0001). Younger age at onset and symptomatic etiology contributed independently to the risk of status epilepticus. Mortality was higher in children with status epilepticus before diagnosis, largely secondary to underlying cause. CONCLUSIONS: Status epilepticus occurs in approximately 10% of children after initial diagnosis of epilepsy. Status epilepticus before initial diagnosis, young age at onset, and symptomatic etiology independently influence the risk of status epilepticus. In those without status epilepticus before diagnosis, the risk is modest and is realized over a prolonged period. For children at highest risk, maintaining abortive therapy in the home may be a reasonable precaution.

Adolescent↗

PubMed2004

Analysis of Acanthamoeba polyphaga surface carbohydrate exposure by FITC-lectin binding and fluorescence evaluation.

AIMS: Characterization of the representative protozoan Acanthamoeba polyphaga surface carbohydrate exposure by a novel combination of flow cytometry and ligand-receptor analysis. METHODS AND RESULTS: Trophozoite and cyst morphological forms were exposed to a panel of FITC-lectins. Population fluorescence associated with FITC-lectin binding to acanthamoebal surface moieties was ascertained by flow cytometry. Increasing concentrations of representative FITC-lectins, saturation binding and determination of K(d) and relative B(max) values were employed to characterize carbohydrate residue exposure. FITC-lectins specific for N-acetylglucosamine, N-acetylgalactosamine and mannose/glucose were readily bound by trophozoite and cyst surfaces. Minor incremental increases in FITC-lectin concentration resulted in significant differences in surface fluorescence intensity and supported the calculation of ligand-binding determinants, K(d) and relative B(max), which gave a trophozoite and cyst rank order of lectin affinity and surface receptor presence. CONCLUSIONS: Trophozoites and cysts expose similar surface carbohydrate residues, foremost amongst which is N-acetylglucosamine, in varying orientation and availability. SIGNIFICANCE AND IMPACT OF THE STUDY: The outlined versatile combination of flow cytometry and ligand-receptor analysis allowed the characterization of surface carbohydrate exposure by protozoan morphological forms and in turn will support a valid comparison of carbohydrate exposure by other single-cell protozoa and eucaryotic microbes analysed in the same manner.

Acanthamoeba↗

PubMed2001

Approaches to study of three Chinese national groups.

BACKGROUND: Although numerous studies have examined the learning approaches of Chinese students, very few comparative studies have been carried out with Chinese students from different nations. AIMS: The present research was designed to identify differences in study approach between Chinese university students drawn from Malaysia, Singapore, and Hong Kong. SAMPLES: The sample consisted of 192 Chinese students with 89 students from Malaysia (43 males and 46 females), 65 students from Hong Kong (41 males and 24 females), and 38 students from Singapore (12 males and 26 females). METHOD: All students completed Entwistle and Ramsden's (1983) Approaches to Studying Inventory by rating on a 4-point Likert scale how well each of the 64 items described their own learning behaviour. RESULTS: The hypotheses that, in comparison with their respective counterparts, Malaysian Chinese students would identify themselves as being more dependent in their learning, Singaporean students as being more adept in presenting ideas/concepts in a clear and systematic fashion, and Hong Kong students as being more anxious in their learning approach, were all supported. However, the hypothesis that Hong Kong students would be more strategic in their learning approach than their national counterparts was not supported. CONCLUSIONS: Based on the significant differences in learning approaches noted among the different Chinese subgroups, caution must therefore be taken against forming fixed conceptualisations of cultural characteristics and considerable care be given in sample definition and selection in cross-cultural research.

Adolescent↗

PubMed2000

Indoloquinoxaline compounds that selectively antagonize P-glycoprotein.

Tumor cells often develop drug resistance through overexpression of membrane transport proteins that effectively efflux anticancer agents. The pharmacologies of the two best-studied transporters, P-glycoprotein (Pgp) and MRP1, are partially overlapping but distinct. To improve the therapeutic potential of drug resistance reversing agents, we have developed a program to identify compounds with selectivity for Pgp or MRP1. Screening of a commercial library of compounds identified indoloquinoxaline compounds with transporter selectivity, and certain examples were synthesized and further evaluated. 1,4-Dibutoxy-6H-indolo[2,3-b]quinoxaline and 4,7-dibutoxy-2,3-dihydrobenzimidazole-2-spiro-3-indolin-2-one were synthesized by condensation of 3,6-dibutoxy-1,4-diaminobenzene and isatin. Neither compound was cytotoxic to MCF-7 cells, nor did either one affect the sensitivity of MCF-7/VP or HL-60/ADR cells at doses up to at least 20 microM, indicating that they do not antagonize MRP1. In contrast, each compound, at doses as low as 0.25 microM, sensitized NCI/ADR cells to vinblastine, actinomycin D, Taxol, and doxorubicin, indicating that they effectively reverse Pgp-mediated multidrug resistance (MDR). Furthermore, the compounds sensitized two additional cell lines that overexpress Pgp to this panel of anticancer drugs. However, these compounds did not affect the sensitivities of MCF-7 or T24 cells to these cytotoxic drugs, and did not alter the sensitivities of any of the tested cell lines to cisplatin or 5-fluorouracil. Both compounds enhanced the intracellular accumulation of [3H]vinblastine by NCI/ADR cells, but did not inhibit photoaffinity labeling of Pgp by [3H]azidopine at concentrations up to at least 100 microM. Therefore, these novel nontoxic indoloquinoxalines selectively sensitize Pgp-overexpressing cells to drugs that are subject to transport by this protein, without modulating the sensitivities of MRP1-overexpressing or non-Pgp cells to cytotoxic drugs. Because of this transporter selectivity, we predict that these compounds will be effective MDR modulators in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1999

Cationic lipid-mediated CFTR gene transfer to the lungs and nose of patients with cystic fibrosis: a double-blind placebo-controlled trial.

BACKGROUND: We and others have previously reported significant changes in chloride transport after cationic-lipid-mediated transfer of the cystic fibrosis transmembrane conductance regulator (CFTR) gene to the nasal epithelium of patients with cystic fibrosis. We studied the safety and efficacy of this gene transfer to the lungs and nose of patients with cystic fibrosis in a double-blind placebo-controlled trial. METHODS: Eight patients with cystic fibrosis were randomly assigned DNA-lipid complex (active) by nebulisation into the lungs followed 1 week later by administration to the nose. Eight control patients followed the same protocol but with the lipid alone (placebo). Safety was assessed clinically, by radiography, by pulmonary function, by induced sputum, and by histological analysis. Efficacy was assessed by analysis of vector-specific CFTR DNA and mRNA, in-vivo potential difference, epifluorescence assay of chloride efflux, and bacterial adherence. FINDINGS: Seven of the eight patients receiving the active complex reported mild influenza-like symptoms that resolved within 36 h. Six of eight patients in both the active and placebo groups reported mild airway symptoms over a period of 12 h following pulmonary administration. No specific treatment was required for either event. Pulmonary administration resulted in a significant (p<0.05) degree of correction of the chloride abnormality in the patients receiving active treatment but not in those on placebo when assessed by in-vivo potential difference and chloride efflux. Bacterial adherence was also reduced. We detected no alterations in the sodium transport abnormality. A similar pattern occurred following nasal administration. INTERPRETATION: Cationic-lipid-mediated CFTR gene transfer can significantly influence the underlying chloride defect in the lungs of patients with cystic fibrosis.

Adult↗

PubMed1999

The in vivo effects of milrinone on the airways of cystic fibrosis mice and human subjects.

Previous studies have indicated that milrinone, a specific type III phosphodiesterase inhibitor, may be able to induce chloride secretion in cystic fibrosis (CF) tissues. We have now assessed the effect of this agent in vivo on the nasal epithelium of CF mutant mice and also in the nose and lungs of human subjects with CF. Wild-type mice showed a small hyperpolarization of the nasal potential difference (PD) in response to milrinone (100 microM, 1.6 +/- 0.6 mV, n = 8, P < 0.05). In contrast, CF mice carrying either the most common human mutation of the gene for the CF transmembrane regulator (CFTR), DeltaF508 (protein mislocalized), or the G551D mutation (protein normally localized) failed to demonstrate this response. Milrinone perfused alone had no significant effect on the baseline nasal PD of human subjects without CF (14.7 +/- 4.0 mV preperfusion; 15.3 +/- 4.6 mV postperfusion), but significantly (P < 0.05) augmented the hyperpolarization induced by a subsequently perfused low-chloride solution (with milrinone, 36.8 +/- 3.0 mV, n = 6; without milrinone, 18.1 +/- 2.2 mV, n = 19). In contrast, in human subjects with CF (n = 6), milrinone alone significantly (P < 0. 05) altered the nasal baseline PD (52.2 +/- 3.3 mV preperfusion; 57. 4 +/- 4.2 mV, postperfusion) but not the subsequent responses to the low-chloride solution (with milrinone, 1.1 +/- 2.2 mV, n = 4; without milrinone, 0.6 +/- 0.5 mV, n = 28) or to isoproterenol (100 microM). In a separate study in subjects (n = 6) with the DeltaF508 mutation, nasal coadministration of milrinone with isoproterenol produced no effect in the presence of amiloride and a low-chloride solution (-0.8 +/- 0.5 mV). This was also the case in the nasal epithelium of CF subjects (n = 4) carrying at least one G551D allele (-0.3 +/- 0.8 mV). Similarly, milrinone did not hyperpolarize the PD of either the tracheal (n = 6) or segmental (n = 6) airways of CF subjects (DeltaF508) when applied topically in vivo in the presence of amiloride, isoproterenol, or adenosine triphosphate (all 100 microM) in a low-chloride solution. These data do not support the use of milrinone to induce chloride secretion in CF airways in vivo.

Amiloride↗

PubMed1998

Effect of IBMX and alkaline phosphatase inhibitors on Cl- secretion in G551D cystic fibrosis mutant mice.

Some cystic fibrosis transmembrane conductance regulator (CFTR) mutations, such as G551D, result in a correctly localized Cl- channel at the cell apical membrane, albeit with markedly reduced function. Patch-clamp studies have indicated that both phosphatase inhibitors and 3-isobutyl-1-methylxanthine (IBMX) can induce Cl- secretion through the G551D mutant protein. We have now assessed whether these agents can induce Cl- secretion in cftrG551D mutant mice. No induction of Cl- secretion was seen with the alkaline phosphatase inhibitors bromotetramisole or levamisole in either the respiratory or intestinal tracts of wild-type or cftrG551D mice. In contrast, in G551D intestinal tissues, IBMX was able to produce a small CFTR-related secretory response [means +/- SE: jejunum, 1.8 +/- 0.9 microA/cm2, n = 7; cecum, 3.7 +/- 0.8 microA/cm2, n = 7; rectum (in vivo), 1.9 +/- 0.9 mV, n = 5]. This was approximately one order of magnitude less than the wild-type response to this agent and, in the cecum, was significantly greater than that seen in null mice (cftrUNC). In the trachea, IBMX produced a transient Cl- secretory response (37.3 +/- 14.7 microA/cm2, n = 6) of a magnitude similar to that seen in wild-type mice (33.7 +/- 4.7 microA/cm2, n = 9). This response was also present in null mice and therefore is likely to be independent of CFTR. No effect of IBMX on Cl- secretion was seen in the nasal epithelium of cftrG551D mice. We conclude that IBMX is able to induce detectable levels of CFTR-related Cl- secretion in the intestinal tract but not the respiratory tract through the G551D mutant protein.

1-Methyl-3-isobutylxanthine↗

PubMed1996

Cystic fibrosis mice carrying the missense mutation G551D replicate human genotype-phenotype correlations.

We have generated a mouse carrying the human G551D mutation in the cystic fibrosis transmembrane conductance regulator gene (CFTR) by a one-step gene targeting procedure. These mutant mice show cystic fibrosis pathology but have a reduced risk of fatal intestinal blockage compared with 'null' mutants, in keeping with the reduced incidence of meconium ileus in G551D patients. The G551D mutant mice show greatly reduced CFTR-related chloride transport, displaying activity intermediate between that of cftr(mlUNC) replacement ('null') and cftr(mlHGU) insertional (residual activity) mutants and equivalent to approximately 4% of wild-type CFTR activity. The long-term survival of these animals should provide an excellent model with which to study cystic fibrosis, and they illustrate the value of mouse models carrying relevant mutations for examining genotype-phenotype correlations.

Animals↗

PubMed1996

Local anaesthesia to the airway reduces sedation requirements in patients undergoing artificial ventilation.

Patients in the intensive care unit require large doses of sedative/analgesic drugs to tolerate the presence of a tracheal tube and other unpleasant stimuli. The ideal regimen for sedatives and analgesics has not yet been found. We have investigated the effects of topical local anaesthesia to the pharynx and airway on sedative/analgesic requirements in 30 ICU patients (25-75 yr old) with no obvious brain injury, undergoing mechanical ventilation. Oral tracheal tubes were changed to a modified tube which allowed instillation of local anaesthetic solutions onto the pharyngeal, laryngeal and tracheal mucosa. Lignocaine 1% (5 ml) or 5 ml of 0.9% saline were instilled hourly for 12 h each for a total of 24 h, in a double-blind, randomized crossover design. Baseline sedation was maintained with propofol or alfentanil infusions, or both, which were titrated to patient comfort and to maintain an optimum sedation score throughout. Twenty-five patients completed the study. Mean total propofol and alfentanil requirements were 766 (SD 524) mg and 17 (7.6) mg, respectively, during 12 h of lignocaine instillation, and 1321 (862) mg and 25 (11.4) mg, respectively, during 12 h of saline instillation. There was a significant reduction (P < 0.05) in the requirements for both agents during the period of lignocaine instillation.

Adult↗

PubMed1996

Studies on the toxicity of analogues of dapsone in-vitro using rat, human and heterologously expressed metabolizing systems.

Three metabolizing systems (rat, heterologously expressed CYP3A4 and human liver) were used to evaluate 12 analogues of dapsone (4,4'diaminodiphenylsulphone) in-vitro. Methaemoglobin formation in a two-compartment and cytotoxicity in a single-compartment model were studied using human erythrocytes and neutrophils, respectively, as target cells. In the two-compartment system using rat microsomes as a generating system and methaemoglobin as an endpoint, the least potent methaemoglobin formers tested were the 2-methyl-4-propylamino (AXDD14), 2-hydroxy-4-4'amino (ABDD5) derivatives and a sulphone/trimethoprim derivative (K-130). Dapsone itself, a 2-methoxy-4-ethylamino (W10) and a 2-hydroxyl-4-ethylamino compound (ABDD39) were the most toxic. In the single-compartment cytotoxicity test using rat microsomes, AXDD14 was again among the least toxic, as was a 2-methyl 4-cyclopentyl derivative (AXDD17) and surprisingly ABDD39. The most cytotoxic compounds again included dapsone itself as well as two 2-trifluoromethyl derivatives. The only significant methaemoglobin formation and cytotoxicity shown with the heterologously expressed human CYP 3A4 was with AXDD14, which was extensively activated. Interestingly, metabolism of dapsone was low using the expressed CYP 3A4. In the two-compartment system using human liver microsomes, AXDD14, K-130 and ABDD5 were oxidized to a significantly lesser extent compared with dapsone and these preliminary findings indicate that future development of these compounds may be worthwhile.

Animals↗

PubMed1996

Asthma prophylaxis agents alter the function of an airway epithelial chloride channel.

A number of recent observations suggest a link between airway Cl-transport and asthma. We have previously described the properties of a voltage- and Ca2+ -dependent chloride channel present in airway epithelium. We now show that agents able to prevent indirectly induced bronchoconstriction (sodium cromoglycate, nedocromil sodium, and furosemide) reduce either the single-channel conductance or the open probability of this channel. The effects of these agents and the Ca2+ dependence of the channel are localized to the same surface, and we show that the channel possesses a specific divalent cation binding site, which responds to concentrations of Ca2+ found on the airway mucosal surface. No alteration of the single-channel properties of this channel were seen in cystic fibrosis epithelium. These data suggest a mechanism by which structurally diverse agents may influence asthma.

Animals↗

PubMed1996

A demonstration using mouse models that successful gene therapy for cystic fibrosis requires only partial gene correction.

Quantifying the level of transgene expression necessary for phenotypic effect is an important consideration in designing somatic gene therapy protocols. A nonlinear relationship between phenotype and gene activity is predicted by control analysis for any autosomal recessive condition. The unaffected phenotype of heterozygotes for autosomal recessive disorders demonstrates that 50% of the normal level of gene expression is sufficient to prevent disease. By extension, an exaggerated and positive effect on the mutant phenotype is predicted to arise from only a small addition of normal transgene expression delivered by gene therapy. We tested this expectation directly by intercrossing mice carrying different Cftr alleles which modulate Cftr gene expression from 0 to 100%. We demonstrate that 5% of the normal level of Cftr gene expression results in a disproportionately large correction of the chloride ion transport defect (50% of normal) and essentially complete rescue of the intestinal disease (100% survival). It follows that even modest levels of transgene expression and only partial correction of CFTR channel activity may have a significant clinical impact.

Animals↗

PubMed1996

Laboratory and clinical studies in support of cystic fibrosis gene therapy using pCMV-CFTR-DOTAP.

The first phase I study of cystic fibrosis gene therapy using cationic liposomes to deliver the cystic fibrosis conductance regulator gene to the nose reported partial and transient correction of the nasal transepithelial ion transport defect, While encouraging, further improvements will be required if this form of treatment is to be of therapeutic value. We tested a new formulation, pCMV-CFTR-DOTAP. The complex is stable for 10 days and effective at correcting the electrophysiological deficit in the trachea of CF mutant mice at 8 or 9 days after intratracheal instillation. Reliable protocols for consistent detection of as few as 10 molecules of CFTR mRNA and DNA in nasal brushing samples are described, Both vector and DNA have been produced to Good Manufacturing Practice standard, Nasal potential difference measurements developed at the National Heart and Lung Institute to assess the CFTR ion channel activity in CF patients replicated well at the Scottish Adult Cystic Fibrosis Service. The SPO fluorescence assay for halide ion conductance in nasal brushings has also been tested. These establish baseline conditions in the Scottish CF cohort from which evidence for correction can be judged under clinical trial conditions. These studies formed the basis for regulatory approval of a randomised, placebo controlled double-blind phase I research study.

Aerosols↗

PubMed1995

Bioelectric characteristics of exon 10 insertional cystic fibrosis mouse: comparison with humans.

Two important issues that can be addressed by animal models are disease pathogenesis and the testing of new treatments, including gene therapy. How closely these models mimic the relevant disorder in humans will determine their usefulness. This study examines how closely the characteristic bioelectric features of cystic fibrosis (CF) are reproduced in the airways and intestinal tract of the exon 10 insertional mutant mouse (cf/cf). In agreement with CF subjects these cf/cf mutant mice demonstrate the following: 1) reduced adenosine 3',5'-cyclic monophosphate-related chloride secretion throughout the respiratory and intestinal tracts both in vivo and in vitro, 2) calcium-related chloride secretion that is preserved in the airways but reduced in the intestine, and 3) a more negative nasal potential difference and increased amiloride response compared with wild-type animals, likely to relate to increased sodium absorption. In contrast to humans, sodium absorption is not increased in the small intestine and is reduced in the trachea of the cf/cf mice. We conclude that the majority of the salient electrophysiological features of CF required for studies of pathogenesis or testing of new treatments are present in these cf/cf mice.

Animals↗

PubMed1993

Non-invasive liposome-mediated gene delivery can correct the ion transport defect in cystic fibrosis mutant mice.

We report gene transfer to the Edinburgh insertional mutant mouse (cf/cf), delivering CFTR cDNA-liposome complexes into the airways by nebulization. We show full restoration of cAMP related chloride responses in some animals and demonstrate, in the same tissues, human CFTR cDNA expression. Overall, a range of correction was seen with restoration of about 50% of the deficit between wild type mice and untreated cf/cf controls. We report modest correction in the intestinal tract following direct instillation and provide initial encouraging safety data for both the respiratory and intestinal tract following the liposome mediated gene delivery. The non-viral nature and potentially lower immunogenicity of DNA-liposomes suggest that this may offer a therapeutic alternative to adenoviral therapies.

Animals↗

PubMed1993

Use of erythropoietin in Jehovah's Witness children following acute gastrointestinal blood loss.

The decision to transfuse children in families practicing the Jehovah's Witness faith with human blood products raises medical, legal, and moral questions. Two cases are presented in which recombinant human erythropoietin was used in pediatric patients as an alternative following acute gastrointestinal hemorrhage. The patients demonstrated increased hematocrit levels obviating the need for blood transfusion. Although erythropoietin is not an alternative to hemotransfusion in the unstable patient, it may be an option in the hemodynamically uncompromised Jehovah's Witness patient following acute blood loss.

Adolescent↗

PubMed1992

Cystic fibrosis in the mouse by targeted insertional mutagenesis.

Cystic fibrosis is a fatal genetic disorder which afflicts 50,000 people worldwide. A viable animal model would be invaluable for investigating and combating this disease. The mouse cystic fibrosis transmembrane conductance regulator gene was disrupted in embryonal stem cells using an insertional gene targeting vector. Germ-line chimaeras were derived and the offspring of heterozygous crosses studied. These homozygous mutant mice survive beyond weaning. In vivo electrophysiology demonstrates the predicted defect in chloride ion transport in these mice and can distinguish between each genotype. Histological analysis detects important hallmarks of human disease pathology, including abnormalities of the colon, lung and vas deferens. This insertional mouse mutation provides a valid model system for the development and testing of therapies for cystic fibrosis patients.

Animals↗

PubMed1992

Ion transport characteristics of the murine trachea and caecum.

1. The basic defect in cystic fibrosis relates to abnormalities of ion transport in affected tissues, such as the respiratory and gastrointestinal tracts. The identification of the cystic fibrosis gene has enabled studies on the production of a cystic fibrosis transgenic mouse to be undertaken. Knowledge of normal ion transport will be necessary for the validation of any such animal model. We have therefore characterized selected responses of the murine trachea and caecum mounted in 'mini' Ussing chambers under open-circuit conditions. 2. Basal values for the trachea were: potential difference, 1.1 mV (SEM 0.2; n = 18); equivalent short-circuit current, 20.4 microA/cm2 (3.6); conductance, 18.2 mS/cm2 (1.7). Corresponding values for the caecum were: potential difference, 0.7 mV (0.1; n = 18); equivalent short-circuit current, 11.0 microA/cm2 (1.6); conductance, 14.5 mS/cm2 (1.4). 3. Amiloride (10 mumol/l) produced a significant (P less than 0.001) fall in potential difference of 43.0% (5.7) in the trachea, but had no significant effect in the caecum. 4. Subsequently, one of three protocols was used to assess the capacity of either tissue for chloride secretion. Addition of a combination of forskolin (1 mumol/l) and zardaverine (10 mumol/l) produced rises in the potential difference of 873% (509) in the trachea and 399% (202) in the caecum. Both A23187 (10 mumol/l) and phorbol dibutyrate (10 nmol/l) increased tracheal potential difference by 350% (182) and 147% (47), respectively. Neither had a significant effect in the caecum. 5. Subsequent addition of bumetanide caused a fall in the stimulated potential difference of between 39.8% and 71.7%, depending on secretagogue and tissue type.(ABSTRACT TRUNCATED AT 250 WORDS)

Amiloride↗