Stabilization and controlled release of micro-encapsulated hydrogen peroxide for wound treatment applications.

AIMS: Hydrogen peroxide (H2 O2 ) disinfection applications are limited by its rapid inactivation. The aims of this study were to (i) micro-encapsulate H2 O2 in silica hydrogels to obtain controlled release up to 72 h, (ii) test hydrogel antimicrobial activity against four common pathogens and (iii) assess H2 O2 release kinetics and antimicrobial activity in 35-65% relative humidity and 37°C to approximate bandaged wound conditions. METHODS AND RESULTS: Hydrogels were characterized using scanning electron microscopy, nitrogen sorption porosimetry, Brunauer Emmet Teller analysis and the Barret-Joyner-Halenda method. Hydrogels formed at lower pH demonstrated increased surface area and decreased pore size, resulting in H2 O2 release lasting 72 h. Using agar well diffusion for antimicrobial activity, statistically significant zones of inhibition (as compared to controls) were seen for Escherichia coli, Staphylococcus aureus, Pseudomona aeruginiosa and Vancomycin-resistant Enterococcus faecalis. Activity remained for hydrogels aged for 72 h in humid, 37°C conditions. CONCLUSIONS: Hydrogels can be synthesized to provide a continuous, controlled release of H2 O2 for up to 72 h. SIGNIFICANCE AND IMPACT OF THE STUDY: Stable, controlled-release H2 O2 hydrogels have potential applications for wound treatment and disinfection of medical equipment, through bonding to bandages or materials such as catheter lumens.

The effectiveness of three home products in cleaning and disinfection of Staphylococcus aureus and Escherichia coli on home environmental surfaces.

AIMS: The objectives of this study were to investigate three products for: (i) cleaning effectiveness on two common household surfaces, and (ii) disinfection effectiveness against two common bacteria. Products included conventional ('bleach'), environmentally preferable (EP), do-it-yourself (DIY: distilled white vinegar, club soda, tea tree oil), 24-h old DIY, and individual DIY components in dilution. METHODS AND RESULTS: For cleaning ceramic, no product was effective (≥85% removal of Hucker's soil), however, DIY performed better than EP and bleach. On stainless, only DIY failed to meet the standard. For disinfection, bleach and EP achieved ≥5·00 log10 reductions under all conditions. DIY and components were more active against Escherichia coli than Staphylococcus aureus but only fresh DIY and 50% vinegar achieved ≥5·00 log10 reductions. CONCLUSIONS: EP is an effective alternative to bleach. DIY may be an adequate alternative for cleaning ceramic and for household use, where complete elimination of micro-organisms is unnecessary; however, it must be freshly prepared each day. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of performance of purportedly safer alternatives for both cleaning and disinfection for use in home health care. The EP product and DIY are potential alternatives for some household uses.

Variation in detection limits between bacterial growth phases and precision of an ATP bioluminescence system.

UNLABELLED: To determine the detection limits of the SystemSure Plus, Escherichia coli and Staphylococcus aureus growth curve samples were taken in lag (1 h), log (6 h), stationary (12 h) and death phases (E. coli 144 h, Staph. aureus 72 h). At each time point, the log10 CFU ml(-1) was determined for the dilution where the SystemSure read 0 relative light units (RLU). Average detection limits were E. coli: lag 6·27, log 5·88, stationary 7·45 and death 6·88; Staph. aureus: lag 4·37, log 5·15, stationary 7·88 and death 7·57. Between-run precision was determined with positive control; within-run precision with positive control, lag and log growth for each bacteria. Within-run precision mean RLU (CV): positive control 274 (12%), E. coli lag 1 (63%), log 2173 RLU (19%), Staph. aureus lag 2 (58%) and log 5535 (18%). Between-run precision was 232 (16%). The precision is adequate with most values within the 95% confidence interval. The detection limit varied by 3·51 log10 for Staph. aureus and 1·47 log10 for E. coli. The lowest detection limits were during E. coli log and Staph. aureus lag phases; the highest was during stationary phase. These results suggest that organism identification and growth phase both impact ATP RLU readings. SIGNIFICANCE AND IMPACT OF THE STUDY: Surface hygiene is a critical component of food safety and infection control; increasingly, ATP detection by bioluminescence is used to evaluate surface hygiene and effective cleaning. This is the first study to show that the number of living and potentially infectious bacteria remaining when the device reads zero varies between the different bacterial life cycle phases: lag, log, stationary and death. ATP device users need to be aware of this information to use the devices appropriately.

Benefits of a statewide high-risk perinatal program.

A study was undertaken to determine whether newborn infants of women in a statewide high-risk program had less perinatal mortality and higher birth weight than newborn infants of high-risk women not in the program. The number of fetal and neonatal deaths was twice as high in the nonprogram group, but there was no statistical difference in birth weight distribution in the two groups. Possible reasons for the program's effectiveness are the facts that a greater number of program women had more prenatal visits and were delivered in a Level II or Level III hospital by appropriate personnel.

Identification of a 37 kDa tacrolimus, sirolimus and cyclosporine binding immunophilin possessing glyceraldehyde 3-phosphate dehydrogenase activity isolated from the Jurkat T cell line.

OBJECTIVE: The isolation and partial characterization of a 37 kDa minor immunophilin from the Jurkat cell line which binds to cyclosporine (CsA), Tacrolimus (FK506) and Sirolimus (RAPA). DESIGN AND METHODS: Using standard protein purification steps including isoelectric focusing and cation exchange chromatography, we have isolated and purified to homogeneity a minor immunophilin from the Jurkat cell line which has a molecular weight of 37 kDa. Binding properties for immunosuppressive drugs CsA, FK506 and RAPA were assessed by Scatchard and displacement analysis. The amino acid analysis and the protein sequences were also studied. RESULTS: The immunophilin was purified to homogeneity and the molecular weight corresponds to 37 kDa. Saturation experiments using 3Hdihydro FK506 gave a Kd of 4.5 nM and the Bmax of 117 nmol/mg protein. Displacement studies using 3Hdihydro FK506 and RAPA gave a Kd of 0.8 nM. For CsA binding, the protein showed somewhat less avid binding. The amino acid composition was in close agreement with the amino acid composition of uracil DNA glycosylase which corresponds to part of the monomer of glyceraldehyde 3 phosphate dehydrogenase (G3PD). Protein digestion gave at least 3 peptides. The primary sequence of the first of these matched 7 of 8 residues of human liver nuclear uracil DNA glycosylase. The 37 kDa immunophilin was found to have G3PD activity not inhibited by FK506. CONCLUSIONS: The amino acid analysis, protein sequences, binding properties and G3PD activity indicate that this 37 kDa immunophilin is different from any other known immunophilins.

Radioreceptor assay for sirolimus in patients with decreased platelet counts.

OBJECTIVE: Sirolimus (RAPA) is a new immunosuppressive drug currently in Phase III clinical trials in combination with cyclosporine A (CsA). The toxicity profiles for CsA and RAPA are only partially overlapping, with RAPA toxicity consisting primarily of hyperlipidemia and myelodepression but without the nephrotoxicity, neurotoxicity, and hepatotoxicity, which are seen with CsA. Patients in the clinical trial are being monitored using HPLC or LC/MS/MS assays; there is no immunoassay for RAPA reported to date. We have previously reported a radioreceptor assay (RRA) for RAPA, which has an excellent correlation with the HPLC assay (r = 0.997). The RRA has several advantages including excellent precision, sensitivity, rapid turnaround time, and a one-step extraction procedure. We report the evaluation of blood samples from patients who were exhibiting RAPA toxicity and comparison of the RRA results with the HPLC results. METHODS: EDTA whole blood specimens (n = 42) were obtained from six renal transplant recipients taking RAPA and CsA and exhibiting decreased platelet counts. Thirty-two samples from patients without decreased platelet counts were also received. The samples were analyzed with the RRA and the results were compared to those obtained with the HPLC assay. RESULTS: By HPLC, the results ranged from 3.2-72.6 micrograms/L RAPA with 43% of the results > or = 30 micrograms/L. With the RRA, the range was 7.7-83.0 micrograms/L RAPA equivalents, with 60% of the results > or = 30 micrograms/L. The RRA results are distinctly higher than the HPLC results all along the range. The correlation between the two assays was 0.861, with a slope of 0.966 and a Y-intercept of 11.1. CONCLUSION: Since the RRA is consistently higher than HPLC concentration in patients with decreased platelet counts, but correlates well in patients with no signs of toxicity, the RRA may be useful for monitoring patients for toxicity, by giving a better indication of increasing degree of immunosuppression than the HPLC assay.

Comparison of binding characteristics of four rapamycin metabolites to the 14 and 52 kDa immunophilins with their pharmacologic activity measured by the mixed-lymphocyte culture assay.

OBJECTIVES: To compare the binding characteristics of four rapamycin (RAPA) metabolites to the 14 and 52 kDa minor immunophilins with their pharmacologic activity, as measured by the mixed-lymphocyte culture (MLC) assay. METHODS: Four RAPA metabolites were isolated by HPLC from the urine of renal transplant patients. Each metabolite was evaluated at 40 micrograms/L for its pharmacologic activity using the MLC assay. The results of the MLC assay were compared to those obtained using the radioreceptor assay (RRA), which measured the binding characteristics of equal concentrations of the metabolites to the 14 and 52 kDa minor immunophilins. RESULTS: Each of the four RAPA metabolites showed low immunosuppressive activity by MLC. RM2 showed the highest activity, with 9% of parent RAPA activity. RM1, 3, and 4 showed 2%, 8%, and 4% activity, respectively. Only RM1 was found to bind significantly to either minor immunophilin, with 21% of parent binding to the 14 kDa protein and 25% of parent binding to the 52 kDa protein. RM2, 3, and 4 bound to both proteins with < or = 2% of parent binding. CONCLUSION: We have demonstrated that the RRA for these four RAPA metabolites shows little cross-reactivity. There is no commercially available immunoassay for RAPA at present. The RRA, therefore, provides an excellent way to rapidly assess efficacy/toxicity of RAPA in patients receiving the drug.

Radioreceptor assay for sirolimus.

OBJECTIVES: To develop a radioreceptor assay (RRA) for sirolimus (rapamycin, RAPA). METHODS: A direct methanol extraction was used to prepare 45 patient samples for the RRA. Results were compared to the results obtained previously using high-performance liquid chromatography (HPLC). Between-run precision, recovery, and drug interference studies were also performed. RESULTS: The RRA is sensitive to 1.0 microgram/L RAPA equivalents in whole blood. Comparison with HPLC yielded a correlation coefficient for 45 patient samples of 0.977. Between-run precision at 2.5, 7.5, 12.5, and 20 micrograms/L showed coefficients of variation (CVs) of 12.9, 9.2, 8.5, and 5.9%, respectively. Recoveries from the extraction procedure were 93% at 7.5 micrograms/L and 103% at 12.5 micrograms/L. Drug interference studies showed no interference in the RRA by cyclosporine (CsA), dexamethasone, prednisone, or methotrexate. CONCLUSION: We have demonstrated that the RRA for RAPA correlates well with HPLC, and has excellent precision and recovery. The procedure is far less time-consuming and complex than HPLC and has potential for automation.

Inflammatory mediators induce sequestration of VE-cadherin in cultured human endothelial cells.

The mechanisms through which inflammatory mediators modify endothelial junctional structure are not well understood. Endothelial cells exposed to 1 mM H2O2, 0.1 mM histamine or 4 mM EDTA displayed decreased amounts of VE-cadherin on the cell surface in a time-dependent manner. H2O2 and EDTA-treated cells showed a sustained reduction in surface VE-cadherin, but histamine (0.1 mM) decreased cell surface VE-cadherin only at 5 and 15 min, not at 30 and 60 min. Sequestering of VE-cadherin could also be visualized as a decrease in immunofluorescent labeling of endothelial junctions in fixed, non-extracted monolayers. However, junctional staining was observed in these cells after membrane extraction. This decreased surface expression of VE-cadherin was actin-filament, but not PKC/MAP kinase dependent. VE-cadherin binding to the cytoskeleton was decreased by EDTA, but was not diminished by histamine or H2O2. Therefore, by promoting sequestration of junctional cadherins, inflammatory mediators may decrease adhesive bonds between apposed endothelial cells and increase solute permeability.

Physical fitness and incidence of hypertension in healthy normotensive men and women.

We measured physical fitness, assessed by maximal treadmill testing in 4,820 men and 1,219 women aged 20 to 65 years. Participants had no history of cardiovascular disease and were normotensive at baseline. We followed up these persons for one to 12 years (median, four years) for the development of hypertension. Multiple logistic risk analysis was used to estimate the independent contribution of physical fitness to risk of becoming hypertensive. After adjustment for sex, age, follow-up interval, baseline blood pressure, and baseline body-mass index, persons with low levels of physical fitness (72% of the group) had a relative risk of 1.52 for the development of hypertension when compared with highly fit persons. Risk of hypertension developing also increased substantially with increased baseline blood pressure.

Comparison of dietary and smoking habit changes in physical fitness improvers and nonimprovers.

Changes in dietary, smoking, and alcohol intake habits of men who voluntarily increased their physical fitness level (n = 532) were compared with men who did not increase physical fitness (n = 390). These men were middle-aged (average age = 43 years), initially sedentary and free of disease, and had been examined twice at a preventive medicine clinic within a 1- to 6-year period. Subjects were retrospectively assigned to two fitness groups--improvers and nonimprovers--based on changes in treadmill performance between baseline and follow-up examinations. Fifteen lifestyle variables were assessed by questionnaire and evaluated for change. At baseline the two groups were similar in demographic variables and diet, differing significantly only in follow-up interval (P less than 0.001), treadmill time (P less than 0.001), whole milk consumption (P less than 0.003), and smoking (P less than 0.001). At follow-up, positive changes in health habits were seen for both groups. Statistically significant differences in health habit change between the groups were seen only for beef (P less than 0.003) and coffee (P less than 0.003) consumption (consumption of both decreasing in more improvers than nonimprovers). Smokers at baseline were less likely to improve their physical fitness. We concluded that individuals who voluntarily increased their physical fitness level were not more likely to change their dietary and alcohol habits than persons who maintained a more sedentary lifestyle.

Immunophilin receptors for immunosuppressive drugs.

The major immunophilins that bind cyclosporine (cyclophilin) and FK-506/rapamycin (FK-BP 12) have been well characterized. They possess rotamase activity, which is inhibited by the immunosuppressant that binds to them. The immunosuppressive action does not appear to be coupled to rotamase activity. The literature is reviewed on some possible mechanisms of immunosuppression. Minor immunophilins of 14, 36, and 52 kDa have also been isolated and partially characterized. Receptor assays employing immunophilins have been developed.

Surrogate measures of physical activity and physical fitness. Evidence for sedentary traits of resting tachycardia, obesity, and low vital capacity.

Studies on physical activity, physical fitness, and health have been hampered because of invalid, unreliable, or impractical measures of physical activity. This report examines the validity of sedentary traits (resting tachycardia, obesity, and low vital capacity) as predictors of physical fitness as assessed by a maximal treadmill exercise test. Study participants were women (n = 3,943) and men (n = 15,627) with at least one visit to the Cooper Clinic in Dallas, Texas. Association of the sedentary traits with physical fitness was examined by multiple regression analyses. Sedentary traits were associated with physical fitness in all age and sex groups, accounting for 12-40% of the variance in treadmill time. When smoking, a simple physical activity index, and sedentary traits were included in a model to predict physical fitness, R2 values ranged from 0.20 to 0.53 in women and 0.45 to 0.61 in men and were significant at p less than 0.0001. These models account for approximately twice as much variance in physical fitness as has been reported previously. The addition of sedentary traits measurements to a simple physical activity index provides a valid estimate of physical fitness in epidemiologic studies.

The impact of previous athleticism on exercise habits, physical fitness, and coronary heart disease risk factors in middle-aged men.

A longitudinal study was performed to determine differences in physiologic variables, health behaviors, risk factors, or clinical status between former athletes (FA) (N = 345) and nonathletes (NA) (N = 75). The subjects, 420 self-referred white males aged 25-60 years old, were examined for prior athleticism and health. Athleticism was determined through self-reported high school or college athletic history. Baseline physiologic and health behavior characteristics were not different between the two groups. Of those study participants (N = 203 FA, 48 NA) who were sedentary at baseline, 208 (N = 167 FA, 41 NA) voluntarily began an exercise program during the followup period (average followup = 56 months). These numbers correspond to exercise adoption rates of 82 and 85% for FA and NA respectively, and were not statistically different (95% CI FA = 0.76, 0.88, NA = 0.73, 0.97). Repeated measures analysis of covariance was performed to determine if physiologic responses to adoption were different between FA and NA. No statistically significant difference was found between the two groups, time effects were similar in the two, and no significant interaction of time by group was observed. We conclude that prior athleticism has little apparent impact on health and health behaviors, thus suggesting that contemporaneous exercise has more impact on clinical variables.

Changes in coronary heart disease risk factors associated with increased treadmill time in 753 men.

Associations between changes in treadmill time and changes in coronary heart disease risk factors were studied in 753 middle-aged men seen between 1978-1981 at the Cooper Clinic in Dallas, Texas. Men were free of known disease at baseline. Significant increases (p less than 0.01) in exercise habits and treadmill time were seen over the course of the study; average follow-up was 1.6 years. In bivariate analyses, increases in treadmill time and weight loss were associated with improvements (p less than 0.01) in risk factors. Multiple regression models were calculated for several risk factors with age, length of follow-up, change in weight, and change in treadmill time as independent variables. After controlling for confounding, increase in treadmill time was independently associated with decreases in the total cholesterol/high density lipoprotein-cholesterol ratio (p less than 0.01) and serum uric acid (p less than 0.05). High density lipoprotein-cholesterol rose with increase in treadmill time (p less than 0.05).