Assessing resistance against macrocyclic lactones in gastro-intestinal nematodes in cattle using the faecal egg count reduction test and the controlled efficacy test.

The main objective of the present study was to evaluate the accuracy of the faecal egg count reduction test (FECRT) to assess the resistance status of ivermectin (IVM)-resistant isolates of the cattle nematodes Ostertagia ostertagi and Cooperia oncophora, using the controlled efficacy test (worm counts) as a reference. The second objective was to investigate whether both IVM-resistant isolates showed side-resistance against moxidectin (MOX) under controlled conditions. Thirty male Holstein calves were experimentally infected with 25,000 L3 of an IVM-resistant O. ostertagi isolate and 25,000 L3 of an IVM-resistant C. oncophora isolate. Twenty-eight days later the calves were randomly divided into 2 treatment groups and 1 untreated control group. Animals in groups 1 and 2 received MOX (Cydectin(®) 1%, Pfizer) and IVM (Ivomec(®) 1%, Merial) respectively, by subcutaneous injection at a dose rate of 0.2mg/kg bodyweight. Faecal samples were collected 7 and 14 days after treatment and animals were necropsied 14/15 days post-treatment. Both the FECRT and the controlled efficacy test demonstrated that the O. ostertagi and C. oncophora isolates were resistant against IVM, with efficacies below 90%. The IVM-resistant O. ostertagia isolate was still susceptible to MOX treatment, as shown by over 99% reduction in egg counts and worm burden. The FECRT suggested borderline resistance against MOX in the IVM-resistant C. oncophora isolate, with egg count reductions between 97% (95% CI: 76; 100) at day 7 and 86% (95% CI: 49; 96) at day 14. However, the controlled efficacy test clearly showed MOX-resistance, with a decrease of only 31% (95% CI: -12; 57) in C. oncophora worm numbers. After MOX treatment, a significantly lower number of eggs per female C. oncophora worms was counted compared to the control group (43% reduction). Due to this reduced fecundity, the FECRT may fail to detect MOX-resistance.

Gene expression analysis of the acute phase response using a canine microarray.

The safety of pharmaceuticals is typically assessed in the dog and rat prior to investigation in humans. As a result, a greater understanding of adverse effects in these preclinical testing species would improve safety assessment. Despite this need, there is a lack of tools to examine mechanisms and identify biomarkers in the dog. To address this issue, we developed an Affymetrix-based oligonucleotide microarray capable of monitoring the expression of thousands of canine genes in parallel. The custom canine array contains 22,774 probe sets, consisting of 13,729 canine and 9045 human-derived probe sets. To improve cross-species hybridization with human-derived probes, the detection region was moved from the variable 3' UTR to the more homologous coding region. Testing of this strategy was accomplished by comparing hybridization of naive dog liver RNA to the canine array (coding region design) and human U133A array (standard 3' design). Although raw signal intensity was greater with canine-specific probe sets, human-derived probes detected the expression of additional liver transcripts. To assess the ability of this tool to detect differential gene expression, the acute phase response was examined in beagle dogs given lipopolysaccharide (LPS). Hepatic gene expression 4 and 24 h post-LPS administration was compared to gene expression profiles of vehicle-treated dogs (n=3/group). Array data was consistent with an acute inflammatory response, with transcripts for multiple cytokines and acute phase proteins markedly induced 4 h after LPS challenge. Robust changes in the expression of transcripts involved with glucose homeostasis, biotransformation, and extracellular matrix remodeling were observed 24 h post-dose. In addition, the canine array identified several potential biomarkers of hepatic inflammation. Strong correlations were found between gene expression data and alterations in clinical chemistry parameters such as serum amyloid A (SAA), albumin, and alkaline phosphatase (ALP). In summary, this new genomic tool successfully detected basal canine gene expression and identified novel aspects of the acute phase response in dog that shed new light on mechanisms underlying inflammatory processes.

Developing empathy to improve patient care: a pilot study of cancer nurses.

Empathy has been identified as an important communication skill that can improve psychological outcomes for patients with cancer and palliative care patients; as such there is a need for cancer nurses to be empathic. The majority of research that has been carried out has been concerned with the definition and measurement of empathy. Exploration into the concept of empathy has shown that there is a need to examine the nature of it and to identify exactly where it succeeds and fails. The questions of how empathy is nurtured and sustained and under what conditions it flourishes and diminishes need to be addressed. This study aimed to investigate how cancer nurses interpret and acquire empathy, and to identify the conditions that influence it. It was also important to establish what cancer nurses considered to be the benefits and disadvantages of expressing empathy. The findings demonstrated that nurses are aware of the concept of empathy and do use their empathic skills; however, there are barriers that could inhibit the expression of empathy, including lack of time, poor environment and communication difficulties. This study highlights the recognition of empathy as a discrete communication skill and the need for more structured courses.

Blood glutathione and cysteine changes in cardiovascular disease.

The need to investigate aminothiols such as glutathione (GSH), cysteine (Cys), and homocysteine (Hcy) in blood is stimulated by the current interest in hyperhomocysteinemia as a risk factor for atherosclerosis. Our current goal was to determine whether various cardiovascular (CV) diseases altered levels of GSH and Cys in blood and the relationships between these two thiols. Blood samples from 96 patients with atherosclerosis and other CV diseases were analyzed and compared with those from 33 control subjects. In CV patients, GSH levels were normal, but free plasma Cys was significantly higher (P < .0001). In patients with atherosclerosis, bound plasma Cys was 21% higher than that in control subjects (P < .0001), and in patients with other CV diseases it was 14% higher (P = .023). Also, in patients with CV diseases, correlations of free GSH with free Cys (P < .007) and total GSH and Cys with age (P < .04) differed from that in control subjects. There were no differences related to functional disability or duration of disease. A key finding was that these abnormal levels of plasma Cys occurred in both atherosclerotic and non-atherosclerotic CV diseases. These results indicate that high levels of oxidized and bound Cys in CV patients create an oxidative environment that may increase susceptibility to vascular damage.

Blood glutathione decreases in chronic diseases.

Previously a high blood glutathione level was correlated with long life span in the mouse and rat and in healthy elderly human beings. This raised the question of whether low glutathione levels occur in unhealthy subjects. To this end, 74 consecutive patients newly admitted to the hospital, with ages ranging from 21 to 89 years and diagnosed with chronic diseases, were studied along with 32 healthy control subjects. Blood samples were analyzed for reduced (GSH) and oxidized (GSSG) glutathione with a high-performance liquid chromatography-dual electrochemical method. The data were integrated with the clinical diagnoses and statistically analyzed. Marked total glutathione decreases from the control levels occurred in over 36% of the patients with chronic diseases including cancer and genitourinary, gastrointestinal, cardiovascular, and musculoskeletal diseases (P < .001). The deficit was due to low GSH concentrations and not to GSSG, which was the same as that in the control subjects. The conclusion is that a decrease in GSH is a risk factor for chronic diseases that may be used to monitor the severity and progress of the diseases. Future work is necessary to elucidate the mechanism of action.

A pilot trial of digital imaging in skin cancer.

We have used inexpensive off-the-shelf equipment for store-and-forward teledermatology and compared the precision and accuracy of digital image consultations with conventional, clinic-based consultations. Thirteen lesions were studied on 12 patients referred to a dermatology clinic for a suspected skin cancer. Patients were examined by two dermatologists. Subsequently, digital images were examined by two different dermatologists. There was almost complete agreement, both among and between the clinical and digital examiners, on different diagnosis and biopsy recommendations. Agreement on the single most likely diagnosis was also good. Digital imaging shows promise in teledermatology.

Isolation of CD34+ cells from blood stem cell components using the Baxter Isolex system.

The CD34 antigen is expressed by human hematopoietic progenitor and stem cells. These cells are capable of reconstituting marrow function after marrow-ablative chemo-radiotherapy. Several different technologies have been developed for the separation of CD34+ cells from bone marrow or peripheral blood stem cell (PBSC) components. We used an immunomagnetic separation technique to enrich CD34+ cells from PBSC components in anticipation of autologous transplantation for patients with B lymphoid malignancies. Twenty-nine patients enrolled on this study and received mobilization chemotherapy followed by G-CSF. Of these, 21 achieved a peripheral blood CD34+ cell level of at least 2.0 x 10(4)/l required by protocol for separation of the stem cell components. A median of three components per patient was collected for processing. The average CD34+ cell concentration in the components after apheresis was 1.0 +/- 1.2%. After the CD34+ cell selection, the enriched components contained 0.6 +/- 0.6% of the starting nucleated cells. The recovery of CD34+ cells, however, averaged 58.4 +/- 19.2% of the starting cell number, with a purity of 90.8 +/- 6.5%. Overall depletion of CD34- cells was 99.96 +/- 0.06%. Nineteen patients were treated with marrow-ablative conditioning regimens and received an average of 6.2 +/- 2.0 x 10(6) CD34+ cells/kg body weight. These patients recovered to an ANC >0.5 x 10(9)/l at a median of 11 days (range 8-14), and platelet transfusion independence at a median of 9 days (range 5-13). Four patients died of transplant-related complications or relapse before 100 days after transplantation. No patient required infusion of unseparated cells because of failure of sustained bone marrow function. These data demonstrate that peripheral blood-derived CD34+ cells enriched by use of an immunomagnetic separation technique are capable of rapid engraftment after autologous transplantation.

Differential distribution of free and bound glutathione and cyst(e)ine in human blood.

The redox status of free and bound glutathione (GSH) and cyst(e)ine (Cys) is altered by oxidative stress, drugs, and disease. Most studies measure only their free forms and not the bound forms, which may have a crucial protective role. For this reason, we determined free and bound, reduced and oxidized GSH and Cys in whole blood, red cells, and plasma of human blood from healthy adults. Distinct compartments of GSH and Cys were found. In whole blood, > 99% GSH was in red cells, of which 16% was bound. GSH values were the same for red cells in whole blood or in cells isolated from the same samples. Only 0.5% of GSH was in plasma, all of which was bound. In contrast, 97% of Cys was in plasma and only 3% in red cells. This was a remarkable separation of these closely related metabolites in the same tissue. In plasma, 60% of Cys was bound. Also, strong correlations were shown of bound vs free Cys and also vs free plus bound Cys. The bound Cys was more constant and suggested that it is a metabolic reserve. Our findings demonstrate the occurrence of significant bound forms of GSH and Cys and have implications for future studies in disease and toxicology.

Glutathione disulfide variability in normal human blood.

A prevailing opinion is that glutathione disulfide (GSSG) levels in human blood are very low, but many studies have reported variable results. Therefore, our objective was to determine valid processing conditions for GSSG measurement and apply them to normal human blood samples. Reproducibility and stability of GSSG were demonstrated in acid extracts of a single sample of fresh whole blood by repetitive measurements during a 6-h period in which the %CVs were < 10. In contrast, in normal subjects tested repeatedly over several years, GSSG values ranging from < 2 to 166 nmol per 10(10) red blood cells were obtained and the overall %CV was 46. Lower GSSG values were obtained in hemolysates and ultrafiltrates. Thus, these results indicate that blood GSSG concentrations differ due to biological variation using optimal processing conditions.

Constrained randomization in a therapeutic efficacy trial.

The use of valid constrained randomization is presented for a therapeutic efficacy trial. The construction and evaluation of randomization schemes are studied for two treatment replicates per block and for hypothesized linear gradients. Two different isomorphism classes of designs are compared using the variance of the mean squared error criterion. The results indicate that valid constrained randomization schemes can be constructed that are superior to ordinary randomization. A mastitis efficacy trial provided the specific problem to be solved.

Bile leak from gallbladder perforation mimicking bowel activity and a false-negative result in a morphine-augmented cholescintigraphy.

Cholescintigraphy of a patient with bile leak demonstrated intra-abdominal activity that mimicked normal bowel activity. Because the gallbladder was not visualized, morphine was injected intravenously. Gallbladder activity after morphine injection was misleading in the finding of chronic cholecystitis. Concurrent abdominal sonography and computerized tomography revealed a thickened gallbladder wall with a gallstone and pericholecystic fluid collection. Exploratory laparotomy confirmed acute and chronic cholecystitis, cholelithiasis, choledocholithiasis, and a pericholecystic abscess. The false-negative conclusion for acute cholecystitis in the patient's morphine-augmented cholescintigraphy resulted from an acceleration of bile leakage due to pre-existing gallbladder perforation.

Low blood glutathione levels in healthy aging adults.

The objective of this investigation was to test the hypothesis that blood glutathione levels are lower in aging human subjects as previously found in blood and tissues of standard rodent models of aging. Thus a study was conducted with 39 men and 130 women, 20 to 94 years old, who were selected by the criteria of being ambulatory, healthy, and free from diabetes mellitus, thyroid disease, anemias, and cancer. The reference group was comprised of the 20- to 39-year-old subjects, whose blood glutathione levels were 547 +/- 53.5 micrograms/10(10) erythrocytes (mean +/- SD) for 40 individuals and defined the reference range (95% confidence limits) of 440 to 654. Based on the 440 micrograms/10(10) erythrocyte cutoff, the incidence of low blood glutathione content in the older subjects increased significantly, particularly in the 60- to 79-year-old group. Their glutathione levels were 452 +/- 86.8 micrograms/10(10) erythrocytes, 17% lower than the reference group (p < 0.001). These findings demonstrate an increased incidence of low glutathione levels in apparently healthy elderly subjects, who thus may be at risk because of a decreased capacity to maintain many metabolic and detoxification reactions mediated by glutathione.

Reversible thallium-201 perfusion defects of the septal and inferoapical segments in a patient with incomplete right bundle branch block and normal coronary angiogram.

Possible causes of reversible perfusion defect in exercise-rest 201Tl myocardial images in a patient with a normal coronary artery angiogram include left bundle branch block, coronary spasm, myocardial bridges, hypertrophic cardiomyopathy, mitral valve prolapse, aortic valve disease and anomalous origin of the left coronary artery arising from the pulmonary artery. This case is a report of a 34-yr-old man with incomplete right bundle branch block and angiographically normal coronary arteries who was found to have reversible defects involving septal and inferoapical walls on stress-rest 201Tl-chloride myocardial imaging.

Urinary excretion of IGF-I and growth hormone in children with IDDM.

OBJECTIVE: To compare the urinary output of insulinlike growth factor I (IGF-I) and growth hormone (GH) in prepubertal and pubertal children with insulin-dependent diabetes mellitus (IDDM) versus nondiabetic subjects and to analyze the relationship between the urinary excretion of these peptides and degree of metabolic control. RESEARCH DESIGN AND METHODS: Group 1 included 30 IDDM patients who had had diabetes for 4.9 +/- 0.7 yr and had normal renal function (mean age 11.6 +/- 0.9 yr); group 2 consisted of 31 control subjects (mean age 9.2 +/- 0.6 yr). Sensitive radioimmunoassays were used to measure IGF-I and GH in urine aliquots from 12-h timed overnight collections that had been dialyzed, concentrated 50-fold, and lyophilized. RESULTS: Significantly lower IGF-I and GH outputs per kilogram body weight per 12 h were observed in IDDM subjects compared with control subjects. When data were expressed per kilogram of body weight, no difference was observed between the urinary output of IGF-I and GH between prepubertal and pubertal subjects within group 1 or group 2. The prepubertal children had significantly lower HbA1 than the pubertal population; however, no correlation was found between urinary output of IGF-I or GH and HbA1. A positive correlation was observed between urinary IGF-I and GH (r = 0.85, P less than .001). CONCLUSIONS: Patients with long-standing IDDM excrete significantly lower urinary levels of IGF-I and GH compared with normal subjects. Serial measurements of these peptides from onset of IDDM are needed to define whether the changes observed are present at diagnosis or are secondary to duration of disease.

Polymeric microspheres for radionuclide synovectomy containing neutron-activated holmium-166.

Poly-L-lactic acid (PLA) microspheres containing neutron-activated 166Ho were investigated as potential agents for radionuclide synovectomy. Stable 165Ho, complexed to acetylacetone (AcAc), was incorporated into PLA spheres by the solvent evaporation technique. Spheres prepared with the optimal mean particle size of 7.2 microns (range 2-13 microns) containing 25.4% 165Ho-AcAc (9.1% 165Ho) were irradiated in a high neutron flux to produce 31.1-36.0 mCi 166Ho. In vitro human plasma studies showed that the irradiated spheres retained 99.0 +/- 0.01% of the 166Ho at 314 hr. In-vivo retention studies were conducted by administering irradiated PLA spheres with 257-591 microCi 166Ho into the joint space of normal rabbits (n = 6). Biodistribution analysis and gamma camera analysis showed 166Ho retention in the joint space after 120 hr of 97.7% +/- 0.8% and 98.2% +/- 2.4%, respectively, with no uptake by the lymph nodes. The ease with which the PLA spheres can be made in the optimal size range for later irradiation and their ability to retain the 166Ho make them attractive agents for radionuclide synovectomy.

Diagnostic significance of urinary growth hormone measurements in children with growth failure: correlation between serum and urine growth hormone.

Twelve-h overnight urine and serum samples obtained simultaneously at 20-min intervals were assayed for growth hormone (GH). Ninety-one children, 5 to 16 y (Tanner stage 1 to 3) participated; group 1 were healthy children, group 2 were children with organic GH deficiency, and group 3 had idiopathic growth failure and normal GH stimulation tests. Serum pool GH concentrations in group 1 were similar to those in group 3 (3.3 +/- 0.3 versus 3.4 +/- 0.2 micrograms/L); group 2 had significantly lower GH concentrations (1.6 +/- 0.2 micrograms/L). Plasma IGF-I levels were significantly greater in groups 1 (14.2 +/- 2.6 nmol/L, p less than 0.001) than in groups 2 and 3 (2.6 +/- 0.5 and 5.5 +/- 0.7 nmol/L, respectively). Urinary GH (mean +/- SEM) standardized for body weight (micrograms/kg) in group 1 (0.31 +/- 0.02) was significantly greater than in group 2 (0.14 +/- 0.01) and group 3 (0.20 +/- 0.01). However, when expressed as microgram/mol creatinine, the output of GH was similar in group 1 (4.0 +/- 0.3) and group 3 (3.4 +/- 0.3); both groups had significantly greater output compared to group 2 (1.3 +/- 0.2). Urinary IGF-I (nmol/kg) in group 1 (0.22 +/- 0.02) was significantly greater than in group 2 (0.12 +/- 0.01) or group 3 (0.07 +/- 0.01). Urinary GH correlated with serum pool GH concentration (r = 0.64, p less than 0.001). Although urinary GH output reflects endogenous GH secretion, the overlap between groups 1 and 3 precludes using urinary GH measurements as a diagnostic test for GH deficiency in children with idiopathic growth failure.

Comparison of urinary growth hormone and IGF-I excretion in small- and appropriate-for-gestational-age infants and healthy children.

The output of urinary growth hormone (GH) and IGF-I were quantitated by RIA in 12-h urine collections obtained from infants who were preterm, small for gestational age (PT-SGA, n = 13); preterm, appropriate for gestational age (PT-AGA, n = 27); full term, small for gestational age (FT-SGA, n = 13); and full term, appropriate for gestational age (FT-AGA, n = 29); and from normal children (n = 33). The amounts of GH and IGF-I (mean +/- SEM) excreted by the PT-SGA and FT-SGA infants were not significantly lower than those excreted by the PT-AGA and FT-AGA groups, respectively [GH (micrograms/kg): PT-SGA 13.7 +/- 3.1 versus PT-AGA 14.0 +/- 2.2, FT-SGA 7.8 +/- 2.4 versus FT-AGA 6.6 +/- 1.8; IGF-I (nmol/kg): PT-SGA 0.52 +/- 0.09 versus PT-AGA 0.53 +/- 0.04, FT-SGA 0.31 +/- 0.05 versus FT-AGA 0.35 +/- 0.04]. All infant groups exhibited significantly greater outputs of urinary GH and IGF-I compared with the children (p less than 0.01). The plasma concentrations of GH in all infant groups were high, whereas the plasma IGF-I levels were low. Microalbumin and beta-2 microglobulin excretion did not correlate with urinary GH and IGF-I output. Despite the higher microalbumin output in FT babies, urinary GH and IGF-I excretion was lower in these groups.(ABSTRACT TRUNCATED AT 250 WORDS)