Fasciola hepatica: a comparative survey of adult fluke resistance to triclabendazole, nitroxynil and closantel on selected upland and lowland sheep farms in Northern Ireland using faecal egg counting, coproantigen ELISA testing and fluke histology.

In order to investigate the incidence and distribution of adult fluke resistance to the fasciolicide tricalbendazole (TCBZ) amongst populations of Fasciola hepatica in sheep flocks in Northern Ireland (NI), individual rectal faeces samples were collected from 3 groups of 20 sheep, before (pre-dose), and 21 days after (post-dose) treatment of the animals with TCBZ, nitroxynil or closantel, on each of 13 well-managed sheep farms distributed across the province. The efficacy of each flukicide was determined for each farm, using faecal egg count reduction (FECRT) and F. hepatica coproantigen ELISA testing. In certain flocks, 2 sheep with high pre-dose faecal egg counts (FEC) were killed 3 days and 21 days respectively after TCBZ treatment, and the histology of the fluke reproductive organs was compared with that of flukes from untreated sheep, and from sheep treated with nitroxynil or closantel 2 days prior to death, using haematoxylin and eosin (H&E) staining and an in situ hybridisation method (TdT-mediated dUDP nick end labelling [TUNEL]) to demonstrate apoptosis. Results from FECRT revealed that in all flocks with a high fluke burden, TCBZ was ineffective in treating chronic fasciolosis, and this finding was generally supported by the results of the coproantigen reduction test (CRT). The histology of reproductive organs of flukes from TCBZ-treated sheep in these flocks was normal, when compared with untreated flukes, and this, together with the FECRT and CRT findings, indicated a likely diagnosis of TCBZ resistance in all the flocks with a high fluke burden. In contrast, nitroxynil and closantel were found to be fully effective against TCBZ-resistant flukes in each of the flocks bearing a high chronic fluke burden. All of the flocks with a high fluke burden and TCBZ resistance were managed on lowland in the South and East of NI. Upland flocks, in the North and West, had low fluke burdens, or were clear of infection; and FECs were too low to allow valid resistance testing. The study highlights the high level of penetration of TCBZ resistance throughout F. hepatica populations in areas of intensively managed sheep production with a high level of fluke challenge. Further, it emphasises the importance of pre-emptive chemotherapeutic action against chronic fasciolosis, using flukicides effective against the egg-producing adult flukes to minimise pasture contamination for the next season's lamb crop. This study also exemplifies the use of several complementary methods (FECRT; CRT; fluke histology; comparative anthelmintic efficacy testing) for confirmation of a diagnosis of fluke drug resistance.

Effect of supplementation with barium selenate on the fertility, prolificacy and lambing performance of hill sheep.

Six weeks before mating, the ewes on six hill farms were randomly assigned to receive either a subcutaneous injection of a long-acting supplement containing 50 mg/ml selenium as barium selenate, or no injected selenium. Before the treatment, the mean activity of glutathione peroxidase (GSHPx) in the ewes on the six farms ranged from 166 to 592 U/g haemoglobin (Hb) and their plasma selenium concentrations ranged from 0.60 to 1.61 micromol/l. Treated ewes had higher plasma selenium concentrations and higher GSHPx activities than control ewes during the study. Conception rates were higher in the treated ewes than in the control ewes. At six weeks, the lambs born to the treated ewes had higher plasma selenium and GSHPx levels than the controls. The treated ewes reared 9 per cent more lambs than the control ewes. The treated ewes had lower abortion rates, and higher liveweights and body condition scores than the controls. There were weak but positive associations between the plasma selenium and GSHPx levels of the ewes and their reproductive performance.

Rational design of a nonpeptide general chemical scaffold for reversible inhibition of PDZ domain interactions.

Novel small molecules were designed to specifically target the ligand-binding pocket of a PDZ domain. Iterative molecular docking and modeling allowed the design of an indole scaffold 10a as a reversible inhibitor of ligand binding. The 10a scaffold inhibited the interaction between MAGI-3 and PTEN and showed cellular activities that are consistent with the inhibition of NHERF-1 function.

High-dose radioimmunotherapy versus conventional high-dose therapy and autologous hematopoietic stem cell transplantation for relapsed follicular non-Hodgkin lymphoma: a multivariable cohort analysis.

We performed a multivariable comparison of 125 consecutive patients with follicular lymphoma (FL) treated at our centers with either high-dose radioimmunotherapy (HD-RIT) using 131I-anti-CD20 (n = 27) or conventional high-dose therapy (C-HDT) (n = 98) and autologous hematopoietic stem cell transplantation. The groups were similar, although more patients treated with HD-RIT had an elevated pretransplantation level of lactate dehydrogenase (41% versus 20%, P =.03) and elevated international prognostic score (41% versus 19%, P =.02). Patients treated with HD-RIT received individualized therapeutic doses of 131I-tositumomab (median, 19.7 GBq [531 mCi]) to deliver 17 to 31 Gy (median, 27 Gy) to critical organs. Patients treated with C-HDT received total body irradiation plus chemotherapy (70%) or chemotherapy alone (30%). Patients treated with HD-RIT experienced improved overall survival (OS) (unadjusted hazard ratio [HR] for death = 0.4 [95% confidence interval (95% CI), 0.2-0.9], P =.02; adjusted HR, 0.3, P =.004) and progression-free survival (PFS) (unadjusted HR =.6 [95% C.I., 0.3-1.0], P =.06; adjusted HR, 0.5, P =.03) versus patients treated with C-HDT. The estimated 5-year OS and PFS were 67% and 48%, respectively, for HD-RIT and 53% and 29%, respectively, for C-HDT. One hundred-day treatment-related mortality was 3.7% in the HD-RIT group and 11% in the C-HDT group. The probability of secondary myelodysplastic syndrome/acute myeloid leukemia (MDS/AML) was estimated to be.076 at 8 years in the HD-RIT group and.086 at 7 years in the C-HDT group. HD-RIT may improve outcomes versus C-HDT in patients with relapsed FL.

Small molecule affinity fingerprinting. A tool for enzyme family subclassification, target identification, and inhibitor design.

Classifying proteins into functionally distinct families based only on primary sequence information remains a difficult task. We describe here a method to generate a large data set of small molecule affinity fingerprints for a group of closely related enzymes, the papain family of cysteine proteases. Binding data was generated for a library of inhibitors based on the ability of each compound to block active-site labeling of the target proteases by a covalent activity based probe (ABP). Clustering algorithms were used to automatically classify a reference group of proteases into subfamilies based on their small molecule affinity fingerprints. This approach was also used to identify cysteine protease targets modified by the ABP in complex proteomes by direct comparison of target affinity fingerprints with those of the reference library of proteases. Finally, experimental data were used to guide the development of a computational method that predicts small molecule inhibitors based on reported crystal structures. This method could ultimately be used with large enzyme families to aid in the design of selective inhibitors of targets based on limited structural/function information.

High-dose chemo-radioimmunotherapy with autologous stem cell support for relapsed mantle cell lymphoma.

Relapsed mantle cell lymphoma is a radiation-sensitive malignancy that is unlikely to be cured by treatment with conventional high-dose therapy and autologous stem cell transplantation. We tested the safety and efficacy of using a CD20-specific monoclonal antibody conjugated with (131)I to deliver high-dose radiation selectively to all lymphoma sites. Patients with relapsed or refractory mantle cell lymphoma received infusions of (131)I-labeled CD20-specific monoclonal antibody (Tositumomab). The antibody dose was 1.7 mg/kg body weight, and the amount of (131)I was calibrated to deliver 20 to 25 Gy to vital normal organs. This treatment was followed 10 days later by administration of high-dose etoposide (30-60 mg/kg), cyclophosphamide (60-100 mg/kg), and infusion of cryopreserved autologous stem cells. The 16 patients in this study had received a median of 3 prior treatments, and 7 had chemotherapy-resistant disease. The median dose of (131)I was 510 mCi (18.87 GBq). There were no therapy-related deaths. Among the 11 patients with conventionally measurable disease at the time of treatment, the respective complete and overall response rates were 91% and 100%. Fifteen patients remain alive, and 12 have had no progression of lymphoma at 6 to 57 months from transplantation and 16 to 97 months from diagnosis. Overall survival at 3 years from transplantation is estimated at 93%, and progression-free survival is estimated at 61%. High-dose treatment with (131)I-Tositumomab, etoposide, and cyclophosphamide results in a high remission rate and may provide long-term disease-free survival for patients with relapsed or refractory mantle cell lymphoma.

Decreased CRH mRNA expression in the fetal guinea pig hypothalamus following maternal nutrient restriction.

The regulation of corticotropin-releasing hormone (CRH) mRNA expression following maternal nutrient restriction was examined in the fetal hypothalamus. Pregnant guinea pigs were food restricted for 48 h or fed normally during late gestation. After nutrient restriction, CRH mRNA levels in the hypothalamic paraventricular nucleus of the fetus were determined using in situ hybridization and were found to be significantly decreased (P<0.0001) compared to controls. In conclusion, we have successfully sequenced the coding sequence of the guinea pig CRH gene, and have shown that a short period (48 h) of maternal nutrient restriction inhibits CRH mRNA expression in the fetal hypothalamus.

Patient adherence to prescribed potassium supplement therapy.

We have investigated whether patient adherence ratios calculated from prescription refill data for potassium supplement medications differ depending on the type of supplement. By using automated pharmacy claims records from a large managed care organization, an index of adherence to prescribed therapy was calculated for each patient as a ratio of total days of drug supplied to the total number of days between prescription refills. The mean patient adherence to prescribed therapy ratios were compared among different potassium drug regimens. There were 2289 patients eligible for analysis; 65.9% were women, and the mean age was 57.6 years. The mean patient adherence ratio for one brand of extended-release tablet, K-DUR, was 0.81 (a majority of the patients were receiving 20 mEq/day). This was higher than the combined mean patient adherence ratio for all other supplements (0.73); the combined mean ratio for all other extended-release tablets (0.74); the combined mean ratio for all other tablets and capsules (0.74); the combined mean ratio for liquids (0.50); the combined mean ratio for liquids and powders (0.63); and equivalent to the ratio for another extended tablet, Micro-K (0.82). Regression analysis showed that increased patient adherence was seen among patients taking K-DUR tablets as compared with those taking other potassium supplements. Increased adherence among patients taking K-DUR remained statistically significant after controlling for number of prescriptions filled, dose, age, sex, and health plan location. Pharmacy claims data can be used effectively to measure patient adherence with potassium supplement therapy. Future research should relate patient adherence ratios to clinical outcomes.

Phosphorylation of specific rat plasma membrane proteins during promotion of gamma-glutamyl transpeptidase-positive hepatic foci and inhibition by di(2-ethylhexyl)phthalate.

The protein kinase activity of isolated plasma membranes from the livers of rats treated with three promoting regimens was examined using both exogenous proteins and endogenous plasma membrane proteins as substrates. Male rats first received either an initiating dose (30 mg/kg) of the hepatocarcinogen diethylnitrosamine or the 0.9% NaCl solution vehicle by i.p. injection at 18 h following partial hepatectomy. Ten days later, the three promoting regimens were begun. These consisted of 10 weeks of treatment with either (a) a choline-deficient (CD) diet, (b) a choline-supplemented (CS) diet containing 0.06% phenobarbital (PHB) (CS plus PHB), or (c) a CD diet containing 0.06% PHB (CD plus PHB). In addition, two other groups of rats received either (a) a CS diet containing 2% di(2-ethylhexyl)phthalate (DEHP) (CS plus DEHP) or (b) a CD diet containing 2% DEHP (CD plus DEHP). DEHP is a widely used plasticizer and environmental contaminant which we have shown previously inhibits the development of putative preneoplastic gamma-glutamyl transpeptidase (GGT) positive foci in rat liver. Total liver plasma membrane protein kinase activity using both protamine sulfate and histone was cyclic adenosine 3':5'-monophosphate independent and did not appear to be a marker of promotion. Its activity was increased by both DEHP which suppresses the development of GGT positive foci and a CD diet which promotes the appearance of GGT positive foci. The CD, CS plus PHB, and CD plus PHB dietary regimens, which promote the appearance of GGT positive foci, induced the phosphorylation of a Mr 40,000 plasma membrane protein in vitro by endogenous protein kinases. Plasma membranes from DEHP-treated rats did not demonstrate phosphorylation of this Mr 40,000 protein. DEHP dietary treatment also blocked the ability of epidermal growth factor to enhance the phosphorylation of its Mr 175,000 receptor protein in isolated liver plasma membranes. These results suggest that the phosphorylation of a Mr 40,000 plasma membrane protein may be important to the early promotional phase of liver carcinogenesis, and that one mechanism by which DEHP inhibits the emergence of GGT positive foci may be by blocking the response of initiated cells to stimulation by epidermal growth factor.

Some thoughts on swimming horses in a pool.

Several indications for swimming horses are recalled. A satisfactory pool and the technique for its use are described. Some observations on the effect on swimming are offered.

An injury-induced diffuse slow potential from brain.

Three different slow potential (SP) changes resulting from focal brain injury are described. The first is an immediate, high amplitude (in excess of 25 mV)negative shift at the site of injury. The second is a biphasic negative-positive SP wave which spreads throughout the cortex ipsilateral to injury and is similar to spreading depression (SD). The third SP change, called here the injury-induced diffuse slow potential (IDSP)is a prolonged (lasting approx. 2h) negative shift occurring simultaneously in many brain areas, also in those far removed from the injured focus. The SD can be separated from IDSP by the size of focal injury; a 20 mu pucture of the parenchyma will trigger IDSP but not SD. An injury resulting from a larger puncture triggers both, SD and IDSP. IDSPcan not be induced by a re-entry of a previously damaged tissue. The magnitude of IDSP has anatomical specificity in that the largest amplitude occurrs in white as compared to gray of the cortex or of the caudate nucleus. Aso, the magnitude of the hypothalamic IDSP is larger when ipsilateral corpus callosum-commissural regions are injured. Electrical stimulation of the cortex in rats sufficiently strong to result in tonic-clonic convulsions triggers SD and IDSP; these two slow potential changes are similar to those induced by mechanical injury. A transpinnate electrical stimulus strong enough to elicit a grand-mal type of discharge results in a diffuse negative slow potential change similar to IDSP elicited by mechanical damage or direct cortical stimulation.

Spontaneous whole brain slow potential changes during recovery from experimental neurosurgery.

Prolonged, nonlocalized brain slow potential changes, frequently associated with cortical spreading depression, occur spontaneously during 5 days following brain implant surgery in rats. These potentials are accompanied by reductions in multiple nerve cell activity and reductions in behavioral motility. The method used in this study provides a tool for evaluating recovery from neurosurgical trauma or other brain injuries, and for testing procedures that facilitate or impede this process.