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Biomedical subjects

J M Howell

Publications and source records attributed to J M Howell.

At least 19 recordsLinked to original sources
PubMed1999

Myophosphorylase gene transfer in McArdle's disease myoblasts in vitro.

McArdle's disease is due to a genetic deficiency of glycogen phosphorylase and results in a lack of glucose mobilization from glycogen during anaerobic exercise. A genetic defect in Merino sheep produces a similar picture. We constructed a first-generation adenoviral recombinant containing the full-length human phosphorylase cDNA under the control of the Rous sarcoma virus promoter. Primary myoblast cultures from phosphorylase-deficient human and sheep muscle were efficiently transduced with this vector, resulting in restoration of the phosphorylase activity. A similar correction of the genetic defect in muscles of McArdle's patients in vivo appears feasible, preferably with the use of an adeno-associated viral vector.

Animals↗
PubMed1999

Is there a future for gene therapy?

The requirements for successful gene therapy are stated and brief details are given of the gene therapy trials in humans which have been approved by the NIH during the years 1989-1997. An indication is given of the gene therapy trials that have been carried out in animal models of Duchenne muscular dystrophy with emphasis on the Golden Retriever dog model. Problems facing somatic gene therapy for inherited muscle diseases are predominantly the following: the extent of the spread of expression from the injection site, the duration of expression and the need for systemic delivery. Brief details of the problems are given and possible ways of overcoming the difficulties are outlined. These include the use of multiple intramuscular injections, increasing the permeability of the extracellular matrix of muscle, inducing mitosis in myoblasts, the use of ex vivo gene transfer, using modified viruses as vectors or synthesized transporter molecules, the use of mechanisms which combat the action of killer T cells, upregulation of isoforms or of alternative proteins such as utrophin for dystrophin and the use of genetic correction methods such as the use of antisense oligonucleotides. It is concluded that there is a potential future for somatic gene therapy in the inherited muscle diseases.

Animals↗
PubMed1999

Development of a snapback method of single-strand conformation polymorphism analysis for genotyping Golden Retrievers for the X-linked muscular dystrophy allele.

OBJECTIVE: To develop a snapback method of single-strand conformation polymorphism (SSCP) analysis for genotyping Golden Retrievers for the X-linked muscular dystrophy allele. ANIMALS: 20 Golden Retriever puppies from a colony with X-linked muscular dystrophy. PROCEDURE: DNA spanning the canine dystrophin mutation was amplified by means of a polymerase chain reaction (PCR), using a primer modified to have an additional sequence at the 5' terminus. The primer was designed so that 1 terminus of the single-stranded PCR product could anneal to the normal sequence flanking the region of the mutation in the allele but not in the mutant allele. True disease status of the dogs was determined by means of a PCR and restriction digest protocol. RESULTS: Snapback SSCP analysis allowed for accurate and unambiguous genotyping of unaffected, carrier, and affected dogs, whereas conventional SSCP analysis, using the unmodified primer, did not. Creatine kinase activities measured within 24 hours after birth were not consistent with genotype. CONCLUSION AND CLINICAL RELEVANCE: Snapback SSCP analysis provided a simple, fast, and accurate method for genotyping Golden Retrievers for the mutation known to cause X-linked muscular dystrophy.

Animals↗
PubMed1998

Suppression of antigen-specific T cell proliferation by measles virus infection: role of a soluble factor in suppression.

Measles virus infection causes a profound immunosuppression. The basis for this immunosuppression is not known. This immunosuppression could be due to virus acting directly on lymphoid cells, the production of an immunosuppressive viral product, or a lymphoid product. We have developed an antigen-specific T cell system to study measles virus-T-cell interactions. We demonstrate that as few as five infectious viral particles added to 1000 T cells results in profound inhibition of antigen-specific T cell proliferation. Supernates taken from measles virus-infected T cells suppress the proliferation of uninfected T cells. Measles-virus-infected HeLa or Vero cells do not produce the factor. The antiproliferative effects of the supernates cannot be attributed to infectious virus, IL-10 or TGF-beta. The soluble factor appears to be larger than 100 kDa, yet retains antiproliferative activity following trypsin digestion with a size less than 10 kDa. Loss of activity is seen following heat treatment at 56 degrees C. The factor is lymphoid cell specific and exhibits cytokine-like behavior yet appears not to be a known cytokine. This soluble factor may be responsible for the overt clinical immunosuppression seen in man and a previously undescribed cytokine induced by measles virus infection of human lymphocytes.

Animals↗
PubMed1998

High-level dystrophin expression after adenovirus-mediated dystrophin minigene transfer to skeletal muscle of dystrophic dogs: prolongation of expression with immunosuppression.

Replication-deficient adenovirus vectors (AdV) have been successfully used to transfer a truncated human dystrophin cDNA to skeletal muscle of dystrophin-deficient mdx mice. A dystrophin-deficient golden retriever dog model (GRMD) has been identified, which, unlike the mouse model, leads to a clinicopathological phenotype similar to that of Duchenne muscular dystrophy (DMD). We show for the first time that high-level dystrophin expression in skeletal muscle of GRMD dogs can be achieved by AdV-mediated gene transfer. However, a humoral and cellular immune response of the host against antigens of viral and transgene origin (similar to that occurring in mdx mice after AdV-mediated dystrophin gene transfer) leads to a decline of dystrophin expression over a 2-month period. Immunosuppression by cyclosporin significantly prolonged transgene expression. The GRMD model may help to solve the open questions pertaining to dystrophin gene transfer such as systemic delivery and improvement of muscle function before human trials for gene replacement therapy in DMD may be considered.

Adenoviridae↗
PubMed1998

Direct dystrophin and reporter gene transfer into dog muscle in vivo.

Bacterial beta-galactosidase cDNA was injected without lipofectin into 41 sites in dog muscle and expression was seen in 22 of them. The cDNA and lipofectin was injected into 35 similar sites and expression was seen in 21. Expression was seen in a maximum of 2.5% of muscle fibers and 23.21% of nonmuscle cells. A total of 106 muscle sites were injected with the minigene with and without lipofectin. In 4 of the 45 sites injected with the minigene without lipofectin human dystrophin was expressed around the periphery of 0.3% of the fibers. Bacterial beta-galactosidase cDNA was injected into the peritoneal cavity of 4 pups, 2 of which also received lipofectin. In all 4, expression was seen in liver, spleen, and mesenteric lymph node. In the 2 pups that received lipofectin, expression was also seen in the diaphragm, intercostal, and abdominal muscles of 1 and in the diagphragm and intercostal muscles of the other. These experiments show that human dystrophin transgene expression can be obtained in dog muscle. However, other methods will be required to increase the degree of expression before gene therapy trials can be undertaken.

Animals↗
PubMed1998

Follow-up compliance in febrile children: a comparison of two systems.

OBJECTIVES: Follow-up compliance is critical in febrile children because they may harbor unrecognized life-threatening illnesses. This study compares follow-up rates between 2 systems: Wilford Hall Medical Center (WHMC), with preset appointments after ED release, and free medical care; and Fairfax Hospital (FFX), where parents must arrange follow-up appointments after ED release, and are responsible for payment for their follow-up visits. The study also investigated factors associated with follow-up compliance. METHODS: This was a prospective, observational study of febrile children seen in 2 EDs with different systems for patient follow-up. From ED records and parental phone calls, diagnosis, follow-up compliance, and demographics were collected. Data were analyzed using logistic regression and chi2. RESULTS: 423 children met entrance criteria, and 330 parents were successfully contacted after the child's ED release (146 from WHMC; 184 from FFX). The WHMC children were more likely to comply with follow-up than were the children in the FFX system (92% vs 67% follow-up, odds ratio 2.5, 95% CI 1.1-5.3). Other factors associated with noncompliance with recommended follow-up were: Hispanic ethnicity, non-English-speaking parents, and follow-up suggested for >24 hours after ED release. For FFX, self-pay, lack of a follow-up physician, parents' dissatisfaction with the ED medical care, and diagnosis of otitis media were also significant factors found associated with noncompliance. CONCLUSION: Febrile children evaluated in a medical system with prearranged follow-up appointments and free medical care are more likely to comply with recommended follow-up than are those evaluated in a system where payment and appointments are the responsibility of the parents. Efforts should be made to improve follow-up compliance by modeling the WHMC system.

Child, Preschool↗
PubMed1998

Modulation of immune system function by measles virus infection: role of soluble factor and direct infection.

Measles virus infection can result in a variety of immunologic defects. We have begun studies to determine the basis for the lack of immune responsiveness to antigen and mitogen following infection. Here we present data showing that Epstein-Barr virus-transformed B-cell lines infected with measles virus produce a soluble factor that can inhibit antigen-specific T-cell proliferation and inhibit the proliferation of uninfected B cells. The soluble factor was neither interleukin-10, transforming growth factor beta, nor alpha/beta interferon. B cells infected with measles virus or treated with the soluble factor were unable to present antigen to T cells in a manner that supported antigen-specific proliferation. This could represent one mechanism of how measles virus limits T-cell expansion. However, we found that once CD4(+) or CD8(+) T cells were activated, their cytolytic activity was intact whether infected with measles virus or treated with soluble factor. Thus, while slow to be generated these cytoxic cells could participate in viral clearance.

Antigen Presentation↗
PubMed1998

A potential role for glucagon in the treatment of drug-induced symptomatic bradycardia.

Nine cases of symptomatic bradycardia are presented in which treatment with intravenous glucagon was administered when atropine failed to improve the patient's condition significantly. Although the cause often was not obvious at presentation, all nine subjects took oral medications that could have contributed to the development of symptomatic bradycardia. Eight of nine patients demonstrated clinical improvement 5 to 10 min after glucagon administration, which was consistent with its peak clinical action. Beta-blockers, calcium channel blockers, and digoxin were ultimately thought to have contributed to the majority of these presentations. This report suggests that glucagon may have a role in the treatment of symptomatic bradycardia, particularly in the presence of beta-adrenergic blockade and perhaps calcium channel blockade. Furthermore, the results in these cases suggest that future clinical trials should not be limited to drug-induced symptomatic bradycardia.

Adrenergic beta-Antagonists↗
PubMed1997

Glucagon therapy in the treatment of symptomatic bradycardia.

Symptomatic bradycardia is commonly seen in the emergency department. Effective drug therapy for this clinical scenario is limited. Although glucagon has been used in no clinical trial in this setting, its cardiac activity may prove useful, particularly in the setting of beta-adrenergic blockade. We report a case series comprising three patients taking maintenance beta-blocker therapy who presented to the ED with symptomatic bradycardia and hypotension and in whom glucagon therapy obviated the need for further treatment. Further study is warranted to evaluate and define the role of glucagon in the treatment of symptomatic bradycardia.

Adrenergic beta-Antagonists↗
PubMed1997

Wound care.

Soft-tissue injuries remain one of the most common problems encountered in the emergency department. This article discusses techniques to minimize pain during the evaluation and repair process, methods to decrease healing complications, and repair considerations.

Aftercare↗
PubMed1997

Use of the dog model for Duchenne muscular dystrophy in gene therapy trials.

Golden retriever muscular dystrophy (GRMD) is an excellent model for the study of the efficacy of gene therapy in dystrophin deficient myopathies for there are many similarities between affected dogs and Duchenne muscular dystrophy (DMD) in boys. GRMD is not caused by deletion mutation but results from a point mutation in the consensus splice acceptor in intron 6 of the canine dystrophin gene. As a result exon 7 is skipped during processing of the GRMD dystrophin messenger RNA. We have developed a rapid test which makes direct use of exon 7 specific genomic PCR products. We have undertaken preliminary experiments on gene therapy using the mini-gene and the full length gene alone and in combination with lipofectin and/or the bacterial beta-galactosidase reporter gene Lac Z. Following direct injection of the Lac Z plasmid, either alone or with lipofectin, about 50% of the sites showed expression when biopsied some 14 days later. The beta-galactosidase activity was present in muscle and granulation tissue but was never abundant. Pups injected intraperitoneally with Lac Z were found to have positive material in their mesenteric lymph nodes, liver and spleen. Those injected with Lac Z and lipofectin also had positive material in the diaphragm, intercostal muscles and abdominal muscles, but again only a small amount of positive material was present at any of the sites. In animals directly injected into the muscle with the dystrophin mini-gene, half had positive staining for dystrophin in biopsies taken 14 days later. Of the 6 sites in the muscles of animals given the mini-gene and lipofectin only one had fibres positive for dystrophin when examined 14 days later. Six pups were injected directly with full-length gene construct and when biopsies were taken 10 days later two of the animals had strongly stained peripheries to a small number of fibres.

Adenoviridae↗
PubMed1997

Characterization of fatal beta blocker ingestion: a review of the American Association of Poison Control Centers data from 1985 to 1995.

OBJECTIVE: To characterize beta blocker-related deaths. METHODS: This is a retrospective review of beta blocker-related exposure data and fatality case abstracts reported to the American Association of Poison Control Centers Toxic Exposure Surveillance System during the 11 year period, 1985 to 1995. Historical and laboratory data were used to determine those fatalities which resulted primarily from beta blocker intoxication. RESULTS: Of 52,156 reported beta blocker exposures, 164 were fatal. In 38 cases, beta blockers were implicated as the primary cause of death. Propranolol was responsible for the greatest number of exposures (44%) and implicated as the cause of death in a disproportionately high percentage of fatalities (71%). Patients were generally young women; 63% were female and 92% were less than 50 years old. The dysrhythmias most often noted in fatal cases were bradycardia and asystole. Cardiopulmonary arrest did not develop until patients were in the care of health care personnel in 59% of cases. Though glucagon was initiated more often than any other intervention in fatal intoxications (83%), optimal dosing and maintenance infusions appear to have been underutilized. CONCLUSIONS: The predominance of fatalities associated with propranolol compared to other beta blockers reflects both its greater frequency of use over the time period studied and its greater toxicity. Since 59% developed. cardiac arrest after reaching health care personnel, further study should focus on identifying medical intervention that can reduce mortality in this group.

Administration, Oral↗
PubMed1996

Hemodynamic effects of calcium chloride in a canine model of acute propranolol intoxication.

STUDY OBJECTIVE: To evaluate the hemodynamic effects of calcium chloride in a canine model of acute propranolol toxicity. METHODS: Two minutes after the completion of a propranolol infusion (10 mg/kg), a bolus of .125 mL/kg 10% CaCl solution followed by an infusion of .375 mL/kg over the next 30 minutes or a bolus and subsequent infusion of an equivalent volume of normal saline solution was administered to each dog. RESULTS: CaCl yielded significant improvements in propranolol-induced decreases in cardiac index and stroke volume compared with saline solution-treated control animals (overall alpha = .05). Furthermore, CaCl administration resulted in earlier improvement in propranolol-induced alterations in mean arterial pressure, maximal left ventricular pressure change over time, and peripheral vascular resistance compared with saline solution (overall alpha = .05). We observed no difference between treatment groups in response to propranolol-induced bradycardia or QRS-interval prolongation. CONCLUSION: In this model of acute propranolol toxicity, CaCl therapy improved depressed hemodynamic status, mainly by a positive inotropic action.

Acute Disease↗