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Biomedical subjects

J E Murphy

Publications and source records attributed to J E Murphy.

At least 37 records · Page 2Linked to original sources
PubMed1998

Adeno-associated virus-mediated delivery of erythropoietin leads to sustained elevation of hematocrit in nonhuman primates.

Recombinant adeno-associated virus encoding the monkey erythropoietin gene (rAAV-cm-Epo) was generated and tested for its potential to confer long-term expression of the gene product following intramuscular injection. A single intramuscular injection of 2 x 10(12) rAAV-cm-Epo particles into two baboons led to sustained high circulating Epo levels and a concomitant increase in hematocrit. The hematocrits reached 62 and 75% by week 10 (from pre-injection values of 38 and 40%, respectively) and remained elevated throughout the study period (28 weeks). Circulating Epo levels were also elevated throughout the study period. Our data demonstrate the potential for long-term gene expression in large animals by a single intramuscular injection of a recombinant adeno-associated virus (rAAV) vector.

Animals↗
PubMed1998

Forms and prevalence of intersexuality and effects of environmental contaminants on sexuality in cricket frogs (Acris crepitans).

Cricket frogs (Acris crepitans) from several different sites in Illinois were collected to assess the effects of environmental contamination on the prevalence of intersex gonads. Of 341 frogs collected in 1993, 1994, and 1995, 2.7% were intersex individuals. There was no statistically significant relationship between the chemical compounds detected and cricket frog intersexuality. However, there was an association approaching significance (p = 0.07) between the detection of atrazine and intersex individuals. A comparison of reference sites with sites that had point polychlorinated biphenyl (PCB) and polychlorinated dibenzofuran (PCDF) contamination revealed a significant relationship between sex-ratio reversal and contamination with PCBs and PCDFs. The sex ratio of juvenile frogs studied from three sites with PCB and PCDF point contamination favored males over females, which was the opposite of the sex ratio in control ponds (p = 0.0007). The statistically significant correlation between organochlorine contamination and sex-ratio reversal suggests PCBs and PCDFs can influence cricket frog sexual differentiation. The current study suggests that in cricket frogs, sex ratios and the prevalence of intersex gonads are altered by environmental contamination.

Aging↗
PubMed1997

Long-term correction of obesity and diabetes in genetically obese mice by a single intramuscular injection of recombinant adeno-associated virus encoding mouse leptin.

The ob/ob mouse is genetically deficient in leptin and exhibits a phenotype that includes obesity and non-insulin-dependent diabetes mellitus. This phenotype closely resembles the morbid obesity seen in humans. In this study, we demonstrate that a single intramuscular injection of a recombinant adeno-associated virus (AAV) vector encoding mouse leptin (rAAV-leptin) in ob/ob mice leads to prevention of obesity and diabetes. The treated animals show normalization of metabolic abnormalities including hyperglycemia, insulin resistance, impaired glucose tolerance, and lethargy. The effects of a single injection have lasted through the 6-month course of the study. At all time points measured the circulating levels of leptin in the serum were similar to age-matched control C57 mice. These results demonstrate that maintenance of normal levels of leptin (2-5 ng/ml) in the circulation can prevent both the onset of obesity and associated non-insulin-dependent diabetes. Thus a single injection of a rAAV vector expressing a therapeutic gene can lead to complete and long-term correction of a genetic disorder. Our study demonstrates the long-term correction of a disease caused by a genetic defect and proves the feasibility of using rAAV-based vectors for the treatment of chronic disorders like obesity.

Animals↗
PubMed1997

Intraperitoneal injection of genetically modified, human mesothelial cells for systemic gene therapy.

An ideal cell type for ex vivo gene therapy should be easy to biopsy, propagate, and genetically engineer in culture, should be transplantable using simple procedures, and should express therapeutic proteins at useful levels. The mesothelial cell appears to satisfy these criteria. Several thousand proliferative mesothelial cells were present in typical specimens of nonpathologic human peritoneal fluid obtained by needle aspiration. These divided rapidly in a specialized medium to yield pure cultures of approximately 10(7) cells within 2 weeks. The replicative lifespan of mesothelial cells cultured from adults was approximately 42-52 population doublings, permitting expansion and cryopreservation of a lifetime supply of autologous cells from one fluid sample. Cells transduced with a human growth hormone (hGH) adenoviral vector secreted 100-300 microg of hGH/10(6) cells per day for at least 6 weeks in culture when maintained at quiescence. Intraperitoneal injection of transduced cells into athymic mice resulted in rapid systemic delivery of hGH, with peak plasma levels of 0.1-1 microg/ml declining over 3 weeks to <1 ng/ml. Mice receiving a second injection of engineered cells displayed the same plasma hGH levels and duration as naive mice. Cells labeled with a beta-galactosidase vector were identifiable by in situ enzymatic staining as clusters attached to peritoneal surfaces at multiple sites for at least 19 days after injection. Cells serially passaged through about three-quarters of their lifespan before transduction and injection were as effective at hGH delivery as earlier-passage cells. These results indicate the clinical potential for ex vivo gene therapy using mesothelial cells.

Adenoviridae↗
PubMed1997

Satisfaction among residents in ASHP-accredited pharmacy residency programs.

The level of work satisfaction among pharmacists in ASHP-accredited residencies was studied. In March 1996 a questionnaire designed to measure residency satisfaction was mailed to 697 individuals in ASHP-accredited pharmacy practice and specialty practice residencies. Subjects responded to 16 statements relating to intrinsic and extrinsic determinants of work satisfaction on a scale of 1 to 5, where 1 = strongly disagree and 5 = strongly agree. Questionnaires were returned by 413 (59%) of the residents. The respondents were predominantly women (76%), and most (86%) had at least a Pharm. D. degree. Hospitals were the primary work setting (88%). Of the 413 residents, 305 were in pharmacy practice residencies and 108 were in specialized residencies. None of the mean scores indicated disagreement (scores < 3) with the positively worded statements or agreement (scores > 3) with the negatively worded statements. The median and mode were equal to 2 (disagree) for the three negatively worded items and 4 (agree) for all but three positively worded items. Only 8% of the residents indicated that they would not accept the residency again if given the chance. Specialized residents tended to rate positively worded statements higher and negatively worded statements lower than pharmacy practice residents. Female residents indicated greater satisfaction than male residents. Pay and benefits were rated slightly better than neutral. Pharmacy residents appeared generally satisfied with their residencies. Specialized pharmacy residents were more satisfied than pharmacy practice residents, and women were more satisfied than men.

Education, Pharmacy↗
PubMed1997

Trapping and visualization of a covalent enzyme-phosphate intermediate.

Using a mutant version of E. coli alkaline phosphatase, we succeeded in trapping and determining the structure of the phospho-enzyme intermediate. The X-ray structure also revealed the catalytic water molecule, bound to one of the active site zinc ions, positioned ideally for the apical attack necessary for the hydrolysis of the intermediate.

Alkaline Phosphatase↗
PubMed1996

National survey of hospital-based pharmacokinetic services.

The status of pharmacokinetic services in the nation's hospitals was studied. A questionnaire was mailed in February 1995 to all 252 respondents to ASHP's 1994 national survey of hospital-based pharmaceutical services who indicated the provision of pharmacokinetic services in their institution. Ninety-eight completed questionnaires were returned (40.2% response rate). The pharmacokinetic services provided required an average of 19.1 person-hours per week, tended to be provided by staff pharmacists, were managed by the pharmacy department, tended to be most focused on aminoglycosides and vancomycin, required an average of 60 notes in patient charts per month, and relied on both computers and calculators for deriving values. There was little contact with patients during the consultation process. Drug concentration measurements tended to be scheduled by service providers, but confidence in the accuracy of the timing of dose administration and blood sampling was limited. Respondents believed that the services tended to be supported by other hospital personnel and that they were successful. There was very little expectation that the workload for providing pharmacokinetic services would increase in the near future. A national survey of hospital-based pharmacokinetic services showed that it took 19 hours per week on average to provide the services, that the focus was on aminoglycosides and vancomycin, and that the services were perceived to be supported by other departments.

Drug Monitoring↗
PubMed1996

Kinetic and structural consequences of replacing the aspartate bridge by asparagine in the catalytic metal triad of Escherichia coli alkaline phosphatase.

In each subunit of the homodimeric enzyme Escherichia coli alkaline phosphatase, two of the three metal cofactors Zn2+ and Mg2+, are bound by an aspartate side-chain at position 51. Using site-specific mutagenesis, Asp51 was mutated both to alanine and to asparagine to produce the D51A and D51N enzymes, respectively. Over the range of pH values examined, the D51A enzyme did not catalyze phosphate ester hydrolysis above non-enzymic levels and was not activated by the addition of millimolar excess Zn2+ or Mg2+. Replacement of Asp51 by asparagine, however, resulted in a mutant enzyme with reduced activity and a higher pH optimum, compared with the wild-type enzyme. At pH 8.0 the D51N enzyme showed about 1% of the activity of the wild-type enzyme, and as the pH was raised to 9.2, the activity of the D51N enzyme increased to about 10% of the value for the wild-type enzyme. Upon the addition of excess Mg2+ at pH 9.2, the D51N enzyme was activated in a time-dependent fashion to nearly the same level as the wild-type enzyme. The affinity for phosphate of the D51N enzyme decreased tenfold as the concentration of Mg2+ increased. Under optimal conditions, the k(cat)/K(m) ratio for the D51N enzyme indicated that it was 87% as efficient as the wild-type enzyme. To investigate the molecular basis for the observed kinetic differences, X-ray data were collected for the D51N enzyme to 2.3 angstroms resolution at pH 7.5, and then to 2.1 angstroms resolution at pH 9.2 with 20 mM MgCl2. The two structures were then refined. The low magnesium, low pH D51N structure showed that the third metal site was unoccupied, apparently blocked by the amide group of Asn51. At this pH the phosphate anion was bound via one oxygen atom, between the zinc cations at the first and second metal sites, which strongly resembled the arrangement previously determined for the D153H enzyme at pH 7.5. In the high magnesium, high pH D51N structure, the third metal site was also vacant, but the phosphate anion bound closer to the surface of the enzyme, coordinated to the first metal site alone. Electron density difference maps provide evidence that magnesium activates the D51N enzyme by replacing zinc at the second metal site.

Alkaline Phosphatase↗
PubMed1996

Academic and training requirements in advertisements for pharmacy management and clinical positions.

The academic and training requirements mentioned in advertisements for various pharmacy management and clinical positions were studied. Position advertisements appearing in five national pharmacy publications between January 1990 and December 1994 were analyzed for requirements or preferences for a particular pharmacy or other degree, postgraduate training (residencies and fellowships), board certification, and experience. The positions were classified as clinical pharmacist, clinical faculty member, pharmacy services manager, clinical manager, or academic manager. A total of 1052 position advertisements were eligible for analysis. Overall, 718 (68%) of the positions either required or preferred postgraduate training; graduates of ASHP-accredited residency programs were sought 10% of the time. Advanced degrees were specified for 934 (89%) of the positions--required for 581 (55%) and preferred for 353 (34%). No degree was specified for 98 (9%) of the positions. Only 20 (2%) of the advertisements indicated that the B.S. degree was a sufficient credential. Board certification was mentioned for 29 (3%) of the positions, although it was never required. Postgraduate pharmacy experience was often acceptable in lieu of advanced degrees and training. Pharmacy management and clinical positions advertised in national pharmacy publications tended to require or prefer residency or fellowship training and advanced degrees.

Advertising↗
PubMed1996

Chain pharmacists' attitudes on and awareness of domestic abuse.

Little has been written to specifically educate pharmacists on the subject of spousal or partner abuse. This project was undertaken to determine whether pharmacists have received training in domestic abuse and to ascertain their attitudes toward participating in abuse intervention. Two hundred twenty-four pharmacists from two chain pharmacies in Arizona received a questionnaire, and 121 (54%) responded. Most (97.5%) had received no training on domestic abuse intervention techniques and did not feel adequately prepared to intervene in a potential situation. Respondents were divided on the extent to which pharmacists should be involved in intervention. Pharmacists who graduated before 1980 were less likely to agree that intervention was an important activity for pharmacists (p < 0.05). More women than men would feel comfortable helping abused patients if they knew more about intervention. More women than men agreed that pharmacists should keep information about domestic abuse on hand (p < 0.025), but most respondents believed that information should be available in pharmacies to give to patients. Opportunities exist for pharmacists to provide educational materials to patients on domestic abuse and abuse intervention.

Adult↗
PubMed1996

Development of physiologically based pharmacokinetic model for methyl tertiary-butyl ether and tertiary-butanol in male Fisher-344 rats.

Methyl tertiary-butyl ether (MTBE) and its metabolite tertiary-butanol (TBA) both cause renal tumors in chronically exposed male rats. Knowledge of the kinetic behavior of MTBE and TBA in rats and its comparison to the kinetics of these chemicals in humans will aid in assessing human risk. The objective of this study was to develop a physiologically based pharmacokinetic (PBPK) model for MTBE and TBA in rats that will form the basis for a human model. Physiological parameters such as blood flows, tissue volumes, and alveolar ventilation were obtained from the literature. Chemical-specific parameters such as the solubility of MTBE and TBA in blood and selected tissues and metabolic rate constants to describe whole-body metabolism of MTBE in rats were measured using vial equilibration and gas uptake techniques, respectively. MTBE metabolism was described in the model as occurring through two saturable pathways. The model was able to predict gas uptake data (100 to 2000 ppm starting concentrations) and levels of MTBE in blood of rats exposed to MTBE by inhalation (400 to 8000 ppm, 6 hr), i.v. (40 mg/kg), and oral (40 or 400 mg/kg) administration. Two different models to describe the dosimetry of TBA in a rat were tested for their ability to predict TBA blood levels after MTBE exposure. TBA blood levels were predicted best at low MTBE exposure concentrations using a two-compartment model. The pharmacokinetics of TBA appear to be far more complex than those of MTBE, and additional experimental data on TBA distribution and elimination will be necessary to refine the submodel. With a quantitative description of the important determinants of MTBE and TBA dosimetry understood, a better assessment of the potential toxic and cancer risk for humans exposed to MTBE can be made.

Administration, Inhalation↗
PubMed1995

Mutations at positions 153 and 328 in Escherichia coli alkaline phosphatase provide insight towards the structure and function of mammalian and yeast alkaline phosphatases.

In order to understand some of the differences between human placental, human, Saccharomyces cerevisiae and Escherichia coli alkaline phosphatases in specific activity, activation by magnesium, and pH versus activity profiles, the X-ray crystal structures of three mutant E. coli alkaline phosphatases have been determined. The aligned sequences of alkaline phosphatases from mammalian, yeast and E. coli show that 25 to 30% of the amino acids are absolutely conserved and the active site residues are completely conserved with the exception of residues 153, 328 and 155. The bacterial enzyme has a salt-bridge, Asp153/Lys328, near the third metal binding site which, based on sequence homology, is apparently absent in the yeast and mammalian enzymes. The human enzymes have histidine at positions 153 and 328, and the yeast enzyme has histidine at position 328. In the E. coli enzyme, Asp153 was replaced by histidine (D153H), Lys328 was replaced by histidine (K328H), and a double mutant (DM) was constructed containing both mutations. The structure of the K328H enzyme was refined using cross-validation to a resolution of 2.3 A with a working R-factor of 0.181 and a free R-factor of 0.249. The DM structure was determined to a resolution of 2.5 A with a working R-factor of 0.166 and a free R-factor of 0.233. The structure of the D135H enzyme, which has been reported to a resolution of 2.4 A, has been re-refined using cross-validation to a working R-factor of 0.179 and a free R-factor of 0.239 for controlled comparisons with the two new structures. In all three structures the most significant changes are related to the bound phosphate inhibitor and the identity of the metal ion in the third binding site. The changes in the position of the phosphate group and the alterations at the third metal binding site indicate the structural basis for the variations in the steady-state kinetic parameters previously reported for these enzymes.

Alkaline Phosphatase↗
PubMed1995

Fluid delivery from infusion-pump syringes.

Fluid-delivery rates of five small-volume infusion-pump syringes were compared. The study consisted of a comparison of the infusion-pump syringes in their respective infusion pumps (1) set for continuous delivery at 1 mL/hr, (2) set for continuous delivery at 3 mL/hr, and (3) set to deliver 1-mL bolus volumes during continuous delivery at 4 mL/hr. The Life-care prefilled 30-mL syringe (Abbott), the DBL 30-mL syringe no. 770205 (DBL Inc.), and the Pump-Jet 30-mL syringe no. 1931 (International Medication Systems) were tested in the Lifecare PCA Plus II infusion pump no. 4100 (Abbott). The 30-mL Pump-Jet syringe no. 1911 (International Medication Systems) and the DBL 30-mL syringe no. 709700 (DBL Inc.) were tested in the Stratofuse PCA infusion pump (Baxter). The infusion pumps were set to deliver fluid continuously at 1 mL/hr for 30 hours, and the solutions were collected separately and weighed. The procedure was repeated with the infusion rate set at 3 mL/hr for 10 hours. For the third part of the study, each syringe was tested to deliver 1-mL boluses with 0, 5, 15, and 25 mL removed from the syringe. The solutions were collected and weighed before and after each bolus was delivered. The volume of solution collected was calculated by using the specific gravity of the solution. The syringes delivered significantly different volumes during the first hour of infusion at both the 1- and 3-mL/hr rates. Differences also existed across time for most of the syringes. Bolus volumes varied greatly after infusion of 0 or 5 mL of fluid but were acceptable for the remainder of the infusions.(ABSTRACT TRUNCATED AT 250 WORDS)

Humans↗