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Biomedical subjects

R Gilbert

Publications and source records attributed to R Gilbert.

At least 37 records · Page 2Linked to original sources
PubMed2000

Modulation of Starling forces and muscle fiber maturity permits adenovirus-mediated gene transfer to adult dystrophic (mdx) mice by the intravascular route.

Duchenne muscular dystrophy (DMD) and other inherited myopathies lead to progressive destruction of most skeletal muscles in the body, including those responsible for maintaining respiration. DMD is a fatal disorder caused by defects in the dystrophin gene. Recombinant adenovirus vectors (AdV) are considered a promising means for therapeutic delivery of a functional dystrophin gene to DMD muscles. If AdV-mediated dystrophin gene replacement in DMD is to be successful, development of a systemic delivery method for targeting the large number of diseased muscles will be required. In this study we investigated two major factors preventing efficient AdV-mediated gene transfer to skeletal muscles of adult animals after intravascular AdV administration: (1) an inability of AdV particles to breach the endothelial barrier and enter into contact with myofibers, and (2) a relatively nonpermissive myofiber population for AdV infection due at least in part to insufficient levels of the coxsackie/adenovirus attachment receptor (CAR). On the basis of established principles governing the transendothelial flux of macromolecules, we further hypothesized that an alteration in Starling forces (increased hydrostatic and decreased osmotic pressures) within the intravascular compartment would facilitate AdV transendothelial flux via convective transport. In addition, experimental muscle regeneration was employed to increase the prevalence of immature myofibers in which CAR expression is upregulated. Here we report that by employing the above-described strategy, high-level heterologous reporter gene expression was achievable in hindlimb muscles of normal rats as well as dystrophic (mdx) mice (genetic homolog of DMD) after a single intraarterial injection of AdV. Microsphere studies confirmed enhanced transport into muscle of fluorescent tracer particles in the size range of AdV, and there was a high concordance between CAR upregulation and myofiber transduction after intraarterial AdV delivery. Furthermore, in mdx mice examined 10 days after intraarterial AdV delivery, the aforementioned procedures had no adverse effects on the force-generating capacity of targeted muscles. These findings have implications for eventual AdV-mediated gene therapy of generalized skeletal muscle diseases such as DMD using a systemic intraarterial delivery approach.

Adenoviridae↗
PubMed2000

The role of panendoscopy in the management of mucosal head and neck malignancy-a prospective evaluation.

BACKGROUND: It is common practice for a panendoscopy to be included in the evaluation of patients with mucosal head and neck malignancies. Whether this intervention is efficient or cost-effective has not been established in our patient population. METHODS: Two hundred twenty-four patients with squamous cell carcinoma involving the oral cavity, pharynx, larynx, or neck were evaluated prospectively with panendoscopy and chest x-ray with or without barium swallow. One hundred fifty-four patients had newly diagnosed tumors and 70 were previously diagnosed and currently undergoing symptom-directed investigations. RESULTS: The incidence of synchronous primary tumors was 2.6% (4 of 154); pulmonary, 1.3%; head and neck mucosa, 1.3%; and esophageal, 0%. There was no associated morbidity. CONCLUSIONS: Although there was no associated morbidity, our head and neck oncology group is of the opinion that routine panendoscopy is not warranted. Specific indications for this investigation are discussed.

Barium Sulfate↗
PubMed2000

Cisapride treatment for gastro-oesophageal reflux in children.

BACKGROUND: Gastro-oesophageal reflux (GOR) is an extremely common and usually self limiting condition in infants. When treatment is required, Cisapride, a pro-kinetic agent, has been commonly prescribed for the symptomatic management of GOR. There have been recent reports of possibly serious adverse events e.g. an increased QTc interval, cardiac arrhythmias, and death, associated with the use of Cisapride. OBJECTIVES: To determine the effectiveness of Cisapride for symptoms of GOR in children compared with placebo or any other non-surgical treatments. SEARCH STRATEGY: Searches were conducted of the Cochrane Central Trials Register and the specialised Trials register of the Cochrane Upper Gastrointestinal and Pancreatic Diseases Group, MEDLINE and Embase. Reference lists of relevant review articles and identified trials were scrutinised and forward citation searches were performed in the Science Citation Index on all trials identified. SELECTION CRITERIA: Randomised controlled trials that compared oral Cisapride therapy with placebo or with other non-surgical treatments for children with a diagnosis of GOR were included. Only studies in which Cisapride was administered orally for a minimum of one week and which documented at least one of the primary outcomes were included. DATA COLLECTION AND ANALYSIS: The primary outcomes were defined as a change in symptoms at the end of treatment, presence of adverse events, occurrence of clinical complications, and weight gain. The secondary outcomes included physiological measures of GOR or histological evidence of oesophagitis. We dichotomised symptoms into 'same or worse' vs 'improved' and calculated summary odds ratios. Continuous measures of GOR (e.g. reflux index) were summarised as a weighted mean difference. All outcomes were analysed using a random effects method. Sensitivity analyses were also performed. MAIN RESULTS: Searches identified eight trials which met the inclusion criteria. Seven trials (a total of 236 participants) compared Cisapride with placebo. The odds ratio for 'same or worse' vs 'improved symptoms' at the end of treatment was 0.34 (95%CI 0.10, 1.19), thus showing no statistically significant difference between the two interventions. There was significant heterogeneity between the studies and the funnel plot suggested substantial publication bias. In the sensitivity analysis, the definition of outcomes was changed to 'any symptoms' vs 'no symptoms'. This resulted in the exclusion of three trials (one of them the largest, best quality trial). The resulting pooled odds ratio showed a significant effect of Cisapride (OR 0.19, 95%CI 0.08, 0.44). There were fewer adverse events with placebo than with Cisapride, but the difference was not statistically significant (OR 1.80, 95%CI 0.87, 3.70) and the result was based on small numbers. Cisapride compared with placebo produced a statistically significant reduction in the reflux index (weighted mean difference -6.49, 95%CI -10.13, -2.85), but as reflux index and clinical symptoms are poorly correlated, the clinical importance of this finding is uncertain. Other measures of oesophageal pH monitoring did not reach significance. One included study compared Cisapride with Gaviscon (or Gaviscon and Carobel). The odds ratio for 'same or worse' vs 'improvement' in the Cisapride group compared with Gaviscon was 3.26 (95%CI 0.93, 11.38). REVIEWER'S CONCLUSIONS: We found no clear evidence that Cisapride reduces symptoms of GOR. The results suggested substantial publication bias favouring studies showing a positive effect of Cisapride. Due to reports of serious adverse events, the company will stop marketing the drug as of July 14th, 2000 and therefore a larger study to provide more conclusive evidence of the effect of Cisapride is no longer possible.

Alginates↗
PubMed2000

Microscopic polyarteritis with renal and cerebral involvement.

We report a 15-year-old girl who presented with renal failure requiring dialysis and a diagnosis of p-ANCA-associated vasculitis. She subsequently developed cerebral vasculitis during treatment with oral cyclophosphamide and prednisolone. Treatment with intravenous cyclophosphamide and plasma exchange produced a complete recovery.

Acute Kidney Injury↗
PubMed2000

Prevention of the dystrophic phenotype in dystrophin/utrophin-deficient muscle following adenovirus-mediated transfer of a utrophin minigene.

Duchenne muscular dystrophy (DMD) is a progressive muscle wasting disorder caused by the lack of a subsarcolemmal protein, dystrophin. We have previously shown that the dystrophin-related protein, utrophin is able to compensate for the lack of dystrophin in the mdx mouse, the mouse model for DMD. Here, we explore whether utrophin delivered to the limb muscle of dystrophin/utrophin-deficient double knockout (dko) neonatal mice can protect the muscle from subsequent dystrophic damage. Utrophin delivery may avoid the potential problems of an immune response associated with the delivery of dystrophin to a previously dystrophin-deficient host. Dko muscle (tibialis anterior) was injected with a first generation recombinant adenovirus containing a utrophin minigene. Up to 95% of the fibres continued expressing the minigene 30 days after injection. Expression of utrophin caused a marked reduction from 80% centrally nucleated fibres (CNFs) in the uninjected dko TA to 12% in the injected dko TA. Within the region of the TA expressing the utrophin minigene, a significant decrease in the prevelance of necrosis was noted. These results demonstrate that the utrophin minigene delivered using an adenoviral vector is able to afford protection to the dystrophin/utrophin-deficient muscle of the dko mouse. Gene Therapy (2000) 7, 201-204.

Adenoviridae↗
PubMed2000

Electrically mediated plasmid DNA delivery to hepatocellular carcinomas in vivo.

Gene therapy by direct delivery of plasmid DNA has several advantages over viral gene transfer, but plasmid delivery is less efficient. In vivo electroporation has been used to enhance delivery of chemotherapeutic agents to tumors in both animal and human studies. Recently, this delivery technique has been extended to large molecules such as plasmid DNA. Here, the successful delivery of plasmids encoding reporter genes to rat hepatocellular carcinomas by in vivo electroporation is demonstrated.

Animals↗
PubMed2000

Toxicity of anticancer agents mediated by electroporation in vitro.

Electroporation is a physical event that temporarily reduces cell membrane barrier properties. Diminished membrane barrier properties are achieved by exposing cells to pulsed electric fields. When a cell has been treated with electric fields it is possible for extracellular agents to gain access to the cell interior. This process has been used in vivo to increase the uptake of chemotherapeutic agents by tumor cells which results in dramatically higher response rates than when drug is used alone. This type of treatment is called electrochemotherapy (ECT); bleomycin is most often used as the drug for this type of treatment. It was hypothesized that electroporation could be used to augment the cytotoxicity of other anticancer agents. Therefore, this study was performed in order to screen 44 different combinations of drug and cell type in vitro to identify drugs that may have higher cytotoxicity when combined with electroporation. Results from seven cell types indicate that the IC50 of bleomycin can be reduced by a factor of 100-5000 when electroporation is used to facilitate internalization. The IC50 values of cisplatin and carboplatin could be reduced by factors ranging from 3 to 13 in six different cell lines as a result of electroporation. These IC50 reductions in multiple cell lines suggest that cisplatin and carboplatin may be effective in vivo as part of ECT treatment.

Antineoplastic Agents↗
PubMed2000

In vivo electroporation of plasmids encoding GM-CSF or interleukin-2 into existing B16 melanomas combined with electrochemotherapy induces long-term antitumour immunity.

When cancer cells, including melanoma cells, are genetically altered to secrete cytokines, irradiated and injected into subjects, long-term antitumour immunity is induced. Optimally, existing melanomas induced to produce cytokines in vivo could stimulate this same immune response. Although in vivo electroporation enhances plasmid expression, electroporation of plasmids encoding granulocyte-monocyte colony stimulating factor (GM-CSF) and interleukin-2 (IL2) into B16 mouse melanomas did not significantly alter tumour growth at the concentration tested. Electrochemotherapy, which causes short-term, complete regressions of treated tumour but no resistance to challenge, was combined with plasmid delivery. The combination treatment resulted in the induction of long-term immunity to recurrence and resistance to challenge in up to 25% of mice.

Animals↗
PubMed2000

Efficacy of surgical salvage for advanced neck metastases after radiotherapy failure.

OBJECTIVE: To analyze the efficacy of surgical salvage in patients with metastatic squamous cell carcinoma to the neck of stage N2 or N3 initially treated with radiotherapy. DESIGN: Retrospective review. SETTING: Tertiary care institution. METHODS: Four hundred and twenty-five patients received neck dissection at Sunnybrook Health Sciences Centre between 1990 and 1996. Sixty-one patients had neck dissection for salvage after radical radiotherapy for advanced neck disease of stage N2 or N3. These patients were selected from the Sunnybrook Head and Neck Database and reviewed. OUTCOME MEASURES: Survival and recurrence. RESULTS: Sixty-one patients underwent neck dissection for salvage after radiotherapy. Five-year overall survival was 30%. The majority (72%) failed locoregionally after surgical salvage. CONCLUSIONS: Surgical salvage after radiotherapy for advanced neck disease has a 30% 5-year survival rate. Surgical salvage after radiotherapy for advanced neck disease is effective in a minority of patients.

Combined Modality Therapy↗
PubMed1999

Identification of amino acid residues critical for aggregation of human CC chemokines macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and RANTES. Characterization of active disaggregated chemokine variants.

Human CC chemokines macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and RANTES (regulated on activation normal T cell expressed) self-associate to form high-molecular mass aggregates. To explore the biological significance of chemokine aggregation, nonaggregating variants were sought. The phenotypes of 105 hMIP-1alpha variants generated by systematic mutagenesis and expression in yeast were determined. hMIP-1alpha residues Asp26 and Glu66 were critical to the self-association process. Substitution at either residue resulted in the formation of essentially homogenous tetramers at 0.5 mg/ml. Substitution of identical or analogous residues in homologous positions in both hMIP-1beta and RANTES demonstrated that they were also critical to aggregation. Our analysis suggests that a single charged residue at either position 26 or 66 is insufficient to support extensive aggregation and that two charged residues must be present. Solution of the three-dimensional NMR structure of hMIP-1alpha has enabled comparison of these residues in hMIP-1beta and RANTES. Aggregated and disaggregated forms of hMIP-1alpha, hMIP-1beta, and RANTES generally have equivalent G-protein-coupled receptor-mediated biological potencies. We have therefore generated novel reagents to evaluate the role of hMIP-1alpha, hMIP-1beta, and RANTES aggregation in vitro and in vivo. The disaggregated chemokines retained their human immunodeficiency virus (HIV) inhibitory activities. Surprisingly, high concentrations of RANTES, but not disaggregated RANTES variants, enhanced infection of cells by both M- and T-tropic HIV isolates/strains. This observation has important implications for potential therapeutic uses of chemokines implying that disaggregated forms may be necessary for safe clinical investigation.

Amino Acid Sequence↗
PubMed1999

Mother-to-child transmission of toxoplasmosis: risk estimates for clinical counselling.

BACKGROUND: Women who acquire toxoplasmosis infection during pregnancy (in most cases detected through serological screening) require counselling about the risk of congenital infection and its clinical sequelae. Reliable estimates of risk are not currently available. We undertook an analysis of data from women referred to the toxoplasmosis reference laboratory, Lyon, France, between 1987 and 1995. METHODS: Information was collected from clinical notes kept at the laboratory and, where necessary, from the relevant obstetrician or paediatrician via telephone. Methods were developed to derive estimates of the risk of congenital toxoplasmosis by exact duration of gestation at maternal seroconversion. FINDINGS: We analysed obstetric and paediatric data on 603 confirmed maternal toxoplasmosis infections. At least 564 women received antiparasitic drugs according to a standard protocol. Congenital infection status was ascertained in 554 cases, and infected children were followed-up for a median of 54 months. The overall maternal-fetal transmission rate was 29% (95% CI 25-33), which masked a sharp increase in risk with duration of gestation from 6% at 13 weeks to 72% at 36 weeks. However, fetuses infected in early pregnancy were much more likely to show clinical signs of infection. These effects counterbalance, and women who seroconverted at 24-30 weeks of gestation carried the highest risk (10%) of having a congenitally infected child with early clinical signs who was thus at risk of long-term complications. INTERPRETATION: This information will assist the clinical counselling of pregnant women diagnosed with acute toxoplasmosis and may guide individual decisions on investigative and therapeutic options. Further studies are required to determine the long-term risks of clinical symptoms and disability due to congenital toxoplasmosis.

Adult↗
PubMed1999

Adenovirus-mediated utrophin gene transfer mitigates the dystrophic phenotype of mdx mouse muscles.

Utrophin is a close homolog of dystrophin, the protein whose mutations cause Duchenne muscular dystrophy (DMD). Utrophin is present at low levels in normal and dystrophic muscle, whereas dystrophin is largely absent in DMD. In such cases, the replacement of dystrophin using a utrophin gene transfer strategy could be more advantageous because utrophin would not be a neoantigen. To establish if adenovirus (AV)-mediated utrophin gene transfer is a possible option for the treatment of DMD, an AV vector expressing a shortened version of utrophin (AdCMV-Utr) was constructed. The effect of utrophin overexpression was investigated following intramuscular injection of this AV into mdx mice, the mouse model of DMD. When the tibialis anterior (TA) muscles of 3- to 5-day-old animals were injected with 5 microl of AdCMV-Utr (7.0 x 10(11) virus/ml), an average of 32% of fibers were transduced and the transduction level remained stable for at least 60 days. The presence of utrophin restored the normal histochemical pattern of the dystrophin-associated protein complex at the cell surface and resulted in a reduction in the number of centrally nucleated fibers. The transduced fibers were largely impermeable to the tracer dye Evans blue, suggesting that utrophin protects the surface membrane from breakage. In vitro measurements of the force decline in response to high-stress eccentric contractions demonstrated that the muscles overexpressing utrophin were more resistant to mechanical stress-induced injury. Taken together, these data indicate that AV-mediated utrophin gene transfer can correct various aspects of the dystrophic phenotype. However, a progressive reduction in the number of transduced fibers was observed when the TA muscles of 30- to 45-day-old mice were injected with 25 microl of AdCMV-Utr. This reduction coincides with a humoral response to the AV and transgene, which consists of a hybrid mouse-human cDNA.

Adenoviridae↗
PubMed1999

Clinical applications of electrochemotherapy.

Electrochemotherapy is a novel treatment which consists of a combination of a chemotherapeutic agent and pulsed electric fields. This relatively new treatment modality relies on the physical effects of locally applied electric fields to temporarily destabilize cell membranes in the presence of a drug to allow increased uptake of the agent into the cytosol. Electrochemotherapy has been used effectively in preclinical and clinical studies. The therapy was shown to be effective regardless of histologic type of tumor including head and neck squamous cell carcinoma, melanoma, basal cell carcinoma, adenocarcinoma and Kaposi's sarcoma. Objective response rates ranging from 72 percent to 100 percent have been reported from these trials. These responses were obtained with minimal adverse side effects. A review of the clinical data for this novel drug delivery method is presented.

Journal Article↗
PubMed1999

In vivo gene delivery by electroporation.

The physical phenomenon of electroporation has been successfully exploited in vitro for the delivery of genes, drugs, and other molecules with increasing frequency over the past two decades. This type of electrically mediated delivery has been translated into an in vivo setting in more recent years with a focus on therapeutic molecules. One promising area is the delivery of genes as a therapy.Advances in molecular medicine have produced a very large amount of information about genes that translate to therapeutic molecules when expressed in living cells. Current standard methods for transferring genes utilize viruses to deliver DNA into cells. These viral methods have not yielded optimal results in most cases. Therefore, there is an increasing interest in nonviral methods for gene delivery. In vivo electrically mediated gene delivery is an attractive alternative because of the site specific nature of delivery as well as the universal applicability of electroporation. A review of the studies performed to investigate and develop this new gene delivery technology is presented.

Journal Article↗
PubMed1999

Use of alternative medicine among patients with head and neck cancer.

OBJECTIVES: To determine the prevalence of alternative medicine use in the population with head and neck cancer and correlate with demographics and tumor characteristics. DESIGN: Cross-sectional survey study. SETTING: Two tertiary cancer centers. PATIENTS: Two hundred consecutive outpatients with consecutive head and neck cancer. INTERVENTIONS: A 10- to 25-minute patient interview administered by primary investigator. MAIN OUTCOME MEASURES: Demographic markers (sex, age, education, household income, marital status, ethnic background, and geographic location); tumor characteristics (tumor site, pathology, staging, time since diagnosis, and incidence of recurrence); conventional mode of treatment; attitudes regarding alternative medicine, source of exposure to alternative medicine, therapeutic rationale, treatment efficacy, sources of information, and discussions with physicians about alternative medicine. RESULTS: Seventy-seven (38.5%) of 200 patients had used alternative medicine for some purpose, and 45 (22.5%) of 200 did so for head and neck cancer. Increased use of alternative medicine occurred among patients of younger age, having a postsecondary education, higher personal income, and Indo-Asian extraction. Of those patients using alternative anticancer therapy, increased use was noted among patients with tumors of the nasopharynx, nonsquamous cell carcinoma pathology, and recurrent disease. Conventional mode of treatment had no association with alternative medicine use. Physicians were believed to be the most knowledgeable about alternative medicine, while the usual proponents of alternative medicine were identified least frequently. CONCLUSIONS: Alternative cancer therapy use among patients with head and neck cancer was 22.5%, with increased use in younger, affluent, better educated patients, and those of Indo-Asian extraction. Patients view physicians as being knowledgeable about alternative medicine. Otolaryngologists should inform themselves about alternative medicine to counsel patients more effectively.

Adult↗
PubMed1999

Extracranial drainage of cerebrospinal fluid: a study of beta-transferrins in nasal and lymphatic tissues.

OBJECTIVE: To determine whether an alternative route of cerebrospinal fluid (CSF) drainage exists through the nasal mucosa and the cervical lymphatic system. STUDY DESIGN: A prospective study was carried out on 18 patients at a university teaching hospital. METHODS: Ten patients undergoing routine endoscopic sinus surgeries and eight patients undergoing neck dissections were recruited for this study. Tissues were sampled from the middle turbinate, nasopharynx, and upper septum in the first group; jugulodigastric lymph nodes and nasopharyngeal tissues were obtained from the second group. Specimens were subjected to immunofixation electrophoresis in an attempt to identify the presence of beta-1 and beta-2 transferrins. Serum samples were obtained from each subject to serve as controls. RESULTS: All tissue specimens contained beta-1 transferrin; none showed evidence of beta-2 transferrin. CONCLUSION: Using this technique, an alternate route of CSF drainage through the nose and the cervical lymphatic system could not be confirmed. Nevertheless, a new technique of performing immunofixation in solid tissues for the purpose of beta-transferrin identification is described.

Cerebrospinal Fluid Rhinorrhea↗
PubMed1999

Identification of the A-band localization domain of myosin binding proteins C and H (MyBP-C, MyBP-H) in skeletal muscle.

Although major constituents of the thick filaments of vertebrate striated muscles, the myosin binding proteins (MyBP-C and MyBP-H) are still of uncertain function. Distributed in the cross-bridge bearing zone of the A-bands of myofibrils, in a series of transverse 43 nm stripes, the proteins are constructed of a tandem series of small globular domains, each composed of approximately 90-100 amino acids, which have sequence similarities to either the C2-set of immunoglobulins (IgC2) and the fibronectin type III (FnIII) motifs. MyBP-C is composed of ten globular domains ( approximately 130 kDa) whereas MyBP-H is smaller ( approximately 58 kDa) and consists of a unique N-terminal segment followed by four globular domains, the order of which is identical to that of MyBP-C (FnIII-IgC2-FnIII-IgC2). To improve our understanding of this protein family we have characterized the domains in each of these two proteins which are required for targeting the proteins to their native site(s) in the sarcomere during myogenesis. Cultures of skeletal muscle myoblasts were transfected with expression plasmids encoding mutant constructs of the MyBPs bearing an N-terminal myc epitope, and their localization to the A-band examined by immunofluorescence microscopy. Based on the clarity and intensity of the myc A-band signals we concluded that constructs encoding the four C-terminal motifs of MyBP-C and MyBP-H ( approximately 360 amino acids) were all that was necessary to efficiently localize each of these peptides to the A-band. Truncation mutants lacking one of these 4 domains were less efficiently targeted to the C-zone of the sarcomere. Deletion of the last C-terminal motif of MyBP-H, its myosin binding domain, abolished all localization to the A-band. A chimeric construct, HU-3C10, in which the C-terminal motif of MyBP-H was replaced by the myosin binding domain of MyBP-C, efficiently localized to the A-band. Taken together, these observations indicate that MyBP-C and MyBP-H are localized to the A-band by the same C-terminal domain, composed of two IgC2 and two FnIII motifs. A model has been proposed for the interaction and positioning of the MyBPs in the thick filament through a ternary complex of the four C-terminal motifs with the myosin rods and titin.

Animals↗