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Biomedical subjects

M Hilf

Publications and source records attributed to M Hilf.

7 recordsLinked to original sources
PubMed1994

High-level expression and purification of secreted forms of herpes simplex virus type 1 glycoprotein gD synthesized by baculovirus-infected insect cells.

Two forms of herpes simplex virus glycoprotein gD were recombined into Autographa californica nuclear polyhedrosis virus (baculovirus) and expressed in infected Spodoptera frugiperda (Sf9) cells. Each protein was truncated at residue 306 of mature gD. One form, gD-1(306t), contains the coding sequence of Patton strain herpes simplex virus type 1 gD; the other, gD-1(QAAt), contains three mutations which eliminate all signals for addition of N-linked oligosaccharides. Prior to recombination, each gene was cloned into the baculovirus transfer vector pVT-Bac, which permits insertion of the gene minus its natural signal peptide in frame with the signal peptide of honeybee melittin. As in the case with many other baculovirus transfer vectors, pVT-Bac also contains the promoter for the baculovirus polyhedrin gene and flanking sequences to permit recombination into the polyhedrin site of baculovirus. Each gD gene was engineered to contain codons for five additional histidine residues following histidine at residue 306, to facilitate purification of the secreted protein on nickel-containing resins. Both forms of gD-1 were abundantly expressed and secreted from infected Sf9 cells, reaching a maximum at 96 h postinfection for gD-1(306t) and 72 h postinfection for gD-1(QAAt). Secretion of the latter protein was less efficient than gD-1(306t), possibly because of the absence of N-linked oligosaccharides from gD-1(QAAt). Purification of the two proteins by a combination of immunoaffinity chromatography, nickel-agarose chromatography, and gel filtration yielded products that were > 99% pure, with excellent recovery. We are able to obtain 20 mg of purified gD-1(306t) and 1 to 5 mg of purified gD-1(QAAt) per liter of infected insect cells grown in suspension. Both proteins reacted with monoclonal antibodies to discontinuous epitopes, indicating that they retain native structure. Use of this system for gD expression makes crystallization trials feasible.

Amino Acid Sequence↗
PubMed1991

A new dosing schedule for gentamicin in blood pythons (Python curtus): a pharmacokinetic study.

Gentamicin is frequently used in the treatment of aerobic Gram-negative infections in reptiles. Pharmacokinetic data to ensure proper dosing are scant, especially for large snakes. A pharmacokinetic study of gentamicin was therefore conducted in four blood pythons. Snakes were given intramuscular injections of either 2.5 mg kg-1 or 3.0 mg kg-1 loading dose followed by 1.5 mg kg-1 at 72 and 96 hours. A linear pharmacokinetic relationship between gentamicin serum concentrations and time was demonstrated in each of the four snakes studied. Peak serum concentrations occurred six to 10 hours after injection and ranged from 4.6 to 8.9 micrograms ml-1. Half-life was variable and ranged from 32 to 110 hours. Total body clearance and apparent volume of distribution varied little between the individual snakes studied. There was no evidence of renal toxicity. For blood pythons a loading dose of 2.5 mg kg-1 followed by 1.5 mg kg-1 at 96 hour intervals is recommended. If higher concentrations are desired, a loading dose of 3.0 mg kg-1 followed by 1.5 mg kg-1 at 96 hours can be given. These dosing schedules will provide serum concentrations in excess of the minimum inhibitory concentrations for most aerobic Gram-negative bacilli that are pathogenic in snakes; gentamicin accumulation with subsequent renal dysfunction should not occur.

Animals↗
PubMed1991

Use of ciprofloxacin versus use of aminoglycosides for therapy of complicated urinary tract infection: prospective, randomized clinical and pharmacokinetic study.

The efficacy of oral ciprofloxacin was compared with that of parenteral aminoglycoside for therapy of complicated urinary tract infection in a prospective randomized trial. The setting was a chronic-care Veterans Administration facility in which long-term bladder catheterization and resistant bacteria were common. Sixty-five consecutive patients were stratified for presence and type of bladder catheter, the presence of Providencia and Pseudomonas aeruginosa organisms versus other urinary pathogens, and renal dysfunction. A pharmacokinetic study of ciprofloxacin concentrations in serum and urine was performed with selected patients. Urinary catheters were present in 94%. All patients were symptomatic, and 58.5% had fever. Ciprofloxacin, 500 mg every 12 h orally, was compared with parenteral aminoglycoside for 7 to 10 days. Clinical response, defined by resolution of symptoms and fever at 5 to 9 days posttherapy (short-term) and 28 to 30 days posttherapy (long-term), was essentially identical for both treatment groups. Bacteriologic response, defined by sterile urine cultures, showed that ciprofloxacin was significantly more efficacious (P = 0.0005) than aminoglycoside at 5 to 9 days posttherapy. However, by 28 to 30 days, the response rate was essentially identical. Emergence of resistance to the study antibiotic was seen in 62 and 70% of those who did not show a bacteriological response and were receiving ciprofloxacin and aminoglycosides, respectively. Concentrations of ciprofloxacin and aminoglycoside in the urine substantially exceeded the MIC for the urinary pathogens isolated, although these concentrations did not correlate with short-term bacteriologic response for either antibiotic. Ciprofloxacin was efficacious in complicated urinary tract infection compared with the current standard of parenteral aminoglycoside among catheterized patients with relatively resistant bacteria.

Administration, Oral↗
PubMed1989

Antibiotic therapy for Pseudomonas aeruginosa bacteremia: outcome correlations in a prospective study of 200 patients.

PURPOSE AND PATIENTS AND METHODS: We performed a prospective clinical study of 200 consecutive patients with Pseudomonas aeruginosa bacteremias to analyze in vitro susceptibility and synergistic testing of antibiotics the patients received and clinical parameters to assess their relationship to survival. RESULTS: No significant correlation between in vitro susceptibility testing (minimal inhibitory concentrations/minimal bactericidal concentrations) and outcome could be demonstrated. Similarly, improved outcome could not be demonstrated for patients receiving antibiotic combinations that were synergistic in vitro (either time-kill or checker-board) versus those combinations that were not. There was also no correlation between results obtained by time-kill curve and checkerboard synergistic testing, i.e., combinations found to be synergistic by one method were not necessarily synergistic by the other method. Clinical parameters associated with improved survival were a urinary portal of entry and absence of neutropenia. Conversely, survival was significantly decreased when the portal was the respiratory tract. The mortality rate between patients receiving combination therapy (27%) and monotherapy (47%) was significant (p less than 0.02); this significant relationship held true for most subgroups including malignancy, nosocomial infection, and infection site. CONCLUSION: Increasing effort should be placed on ensuring timely administration of combination therapy to patients with P. aeruginosa bacteremia since the use of combination therapy was even more important in determining outcome than was underlying disease.

Adolescent↗
PubMed1987

Susceptibility of 100 blood isolates of Pseudomonas aeruginosa to 19 antipseudomonal antibiotics: old and new.

The in vitro susceptibility of 19 antipseudomonal antibiotics, old and new, were tested against 100 blood isolates of Pseudomonas aeruginosa. The isolates were taken from consecutive bacteremic patients hospitalized in nine Pittsburgh hospitals from 1983 to 1984. Twelve percent of isolates tested were tobramycin-resistant. All of the tobramycin-resistant isolates were sensitive to apalcillin, azlocillin, aztreonam, cefsulodin, ceftazidime, ciprofloxicin, and imipenem. Two isolates were resistant in vitro to all beta-lactam antibiotics tested, but sensitive to all aminoglycosides. One hundred percent of isolates tested were susceptible to imipenem, whereas, 98% were susceptible to apalcillin and cefsulodin. The lowest MIC90 was 2 mg/L for ciprofloxicin. Tolerance of P. aeruginosa (MIC90/MBC90 ratio greater than 32) was not observed for any antimicrobial agent.

Anti-Bacterial Agents↗
PubMed1984

Effects of midazolam on directly stimulated muscle biopsies from control and malignant hyperthermia positive patients.

Midazolam is a water soluble benzodiazepine of interest to the anaesthetist for use as a premedicant and for induction of anaesthesia. The effects of midazolam were observed on the resting tension of directly stimulated muscle biopsied from control and malignant hyperthermia (MH) susceptible patients. In addition, interactions between midazolam and the two most commonly used MH diagnostic agents (halothane and caffeine) were examined. Midazolam, at maximum therapeutic concentrations (ca. 0.5 microgram X ml-1), had no detectable effects on muscle contraction in preparations from control or MH positive patients. Midazolam did elicit a contracture from control and MH positive preparations when used in a concentration range of 160-1280 micrograms X ml-1. There was no significant difference between control and MH positive patients in minimum concentration of midazolam causing contracture or the strength of contracture at the respective eliciting concentration. There appears to be no interaction between midazolam and either halothane or caffeine on the resting tension of the directly stimulated muscle twitch preparation.

Anti-Anxiety Agents↗