Detection of a knockdown mutation in the voltage-sensitive sodium channel associated with permethrin tolerance in Sarcoptes scabiei var. hominis mites.

BACKGROUND: Increasing evidence has sparked a debate on the loss of sensitivity of scabies mites to conventional permethrin therapy. Mutations in the voltage-sensitive sodium channels (VSSC) were associated with knockdown resistance (kdr) in many arthropods, but have never been identified in Sarcoptes scabiei variatio (var.) hominis mites. OBJECTIVES: To identify factors contributing to therapy failure. METHODS: Sixty-seven mites were collected from 64 scabies-infested patients in Vienna, Austria, of whom 85.9% were refractory to prior permethrin-based treatments, and genotyped for the presence of nucleotide polymorphisms in Domain II of the VSSC, known to be associated with kdr. Information regarding previous antiscabietic therapies, decontamination procedures and possible re-infestations by contacts as well as the response to re-imposed therapies were obtained. RESULTS: Sequence alignment comparisons revealed previously unidentified mutations in the coding region of Domain II of the VSSC. A novel A1663T transversion was detected in 97.0% of the mites, resulting in a non-synonymous substitution from methionine to leucine, M918L, a mutation known to confer kdr in other arthropods. In addition, a synonymous G1659A transition was identified in one mite, which otherwise showed a nucleotide sequence identical to the wild-type reference. No major inconsistencies were observed within the previous therapeutic and decontamination procedures, which could have accounted for the observed non-responsiveness to permethrin-based therapies. Subsequent cure of infestation was achieved in 65.6% of the participants, predominantly by combination therapies with topical permethrin and systemic ivermectin. However, in 14.6% of the cured cases, permethrin monotherapy sufficed for eradication of scabies, albeit in some cases prolonged exposure was necessary. CONCLUSIONS: The kdr-associated M918L mutation in the VSSC gene has now emerged in S. scabiei var. hominis mites. Hence, loss of sensitivity to permethrin due to kdr-type resistance may be more prevalent than anticipated and may be decisive for the therapy responsiveness of scabies-infested patients.

Fucosyltransferase and alpha-L-fucosidase activities and fucose levels in normal and malignant endometrial tissue.

Previous immuno- and lectin-histochemical studies using mAbs and Ulex europaeus lectin I, which recognize various fucose-containing blood group antigens, have shown an increased expression of Lewis and H blood group antigens in endometrial carcinoma. We investigated the biochemical basis of aberrant fucose-containing antigen expression by comparing the activity of fucosyltransferases (FTase) and alpha-L-fucosidase in tissue biopsies from normal (n = 18) and malignant (n = 20) endometrium. Alteration of FTase activity in tumor tissue homogenates was evaluated by using a panel of FTase substrates including N-acetyllactosamine (type 2), lacto-N-biose I (type 1), and phenyl-beta-D-galactoside. Based on histological subtyping, the endometrioid group (n = 14) showed a significant (P < 0.05) increase in tumor FTase activity with all three substrates, while no significant increase was detected for the papillary serous group (n = 4). Matched pair analysis of normal and tumor tissue from a subgroup (n = 5) of the patients with increased tumor enzyme activity also showed higher FTase activity (P < 0.05) in the tumor tissue when the type 1 substrate was used. Regression analysis showed a correlation between the FTase activities acting on type 2 or type 1 substrates (r = 0.821 and r = 0.722, respectively) and the endogenous fucose levels in tumor homogenates. Spectrophotometric analysis of alpha-L-fucosidase activity using p-nitrophenyl-alpha-L-fucoside revealed a higher activity in tumor homogenates than in normal homogenates (P < 0.05) and, therefore, could not account for the enhanced expression of fucose-containing antigens. The current study suggests that aberrant expression of fucose-containing antigens, such as the H and the Lewis blood-group antigens, in endometrial carcinoma is consequential to the change in FTase rather than in alpha-L-fucosidase activity. In addition, the investigation suggests that different glycosylation mechanisms are operative in different subtypes of endometrial cancer.

Reduction of background activity through radiolabeling of antifibrin Fab' with 99mTc-dextran.

UNLABELLED: Scintigraphic detection of occult disease is limited by background activity in the blood and in the extravascular space that reduces target-specific contrast. To lower nonspecific background activity, we have studied the in vivo biodistribution kinetics of a clot-targeting molecule (MH1 Fab') attached to (99m)Tc-dextran. We tested the hypothesis that the complex will have better background clearance than the directly radiolabeled clot-targeting molecule. METHODS: Fab' fragments of MH1 Fab' antifibrin antibody were coupled to (99m)Tc-sulfhydryl dextran through disulfide exchange, and clot binding bioreactivity was tested in vitro and in vivo in a rabbit jugular vein thrombus model. To assess the background clearance kinetics and extravascular leakage, we studied (99m)Tc-dextran, (99m)Tc-MH1 Fab', and the (99m)Tc-dextran-labeled MH1 Fab' complexes in rats. RESULTS: (99m)Tc-radiolabeled dextran derivatives were radiochemically stable and retained clot-binding bioreactivity in vivo. In the rat model, blood and tissue clearance of the (99m)Tc-dextran MH1 Fab' constructs was substantially improved relative to directly radiolabeled MH1 Fab'. At 1 h, total and extravascular tracer localizations in lung and muscle were significantly lower for 99mTc-dextranradiolabeled MH1 Fab' than for (99m)Tc-MH1 Fab' (P < 0.05). CONCLUSION: The study observations suggest that radiolabeling through a (99m)Tc-dextran moiety may improve the detection of pulmonary emboli and other clinically important fixed intravascular targets by lowering nonspecific background activity.

Neuronal recordings in human lateral temporal lobe during verbal paired associate learning.

The temporal lobe has a role in memory and learning. The objective of this study was to assess the activity of neurons in the temporal cortex of humans during learning. Single neuron activity was recorded with extracellular microelectrodes in adults undergoing awake craniotomies for resections for medically intractable epilepsy. Participants were administered control tasks of overt reading, silent reading, nonword visual controls and a recent verbal memory task with distracters followed by a verbal paired associates (PA) paradigm. Forty-nine neurons were identified and characterized by their relationship to language and memory. Neurons related to overt speech (Type A neurons) showed increased activity during learning in patients who were good learners compared to patients who were poor learners. Neurons not related to language or to memory (Type D neurons) showed increased activity with exposure to unlearned word pairs in poor learners compared with those who were good learners. Activity in all groups diminished with practice. Levels of activity in specific types of neurons in human temporal lobe differ in patients who learn word pairs rapidly or poorly.

Halifax interlaminar clamp for posterior cervical fusion: a long-term follow-up review.

Fifty consecutive patients requiring posterior cervical fusion for various pathologies were treated with Halifax interlaminar clamps for internal spinal fixation. Fusion involved the C1-2 level in 17 cases, the C1-3 level in one, and the lower cervical area (C2-7) in 32. No patient was lost to follow-up review, which varied from 6 to 40 months (average 21 months). Fusion failed in five patients, three at the C1-2 level, one at the C1-3 level, and one at the C2-3 level. Screw loosening was the cause of failure in four patients, and in one the arch of C-1 fractured. No other complications occurred. Because of the lack of complications, avoidance of the hazards of sublaminar instrumentation, and an excellent fusion rate, this technique is highly recommended for posterior cervical fusion in the lower cervical spine. Atlantoaxial arthrodesis was achieved in only 14 (82%) of 17 patients, however, which might be due to the higher mobility at this multiaxial level. Improved results in this region may be possible by using a new modified interlaminar clamp, by performing adequate bone fusions, and by postoperative external halo immobilization in high-risk patients.

Use of MR imaging-compatible Halifax interlaminar clamps for posterior cervical fusion.

Twenty-one patients requiring posterior cervical fusion were treated with magnetic resonance (MR) imaging-compatible Halifax interlaminar clamps for internal fixation. Various levels were involved: the C1-2 level in eight cases, the C4-5 level in four, the C5-6 level in three, the C6-7 level in three, the C4-6 level in two, and the C5-7 level in one. Bilateral clamps were used in 18 cases and unilateral clamps in three. Autogenous iliac bone grafting was performed in all cases but one. Follow-up periods ranged from 1 to 18 months (average 9.2 months), with no complications or mechanical failures occurring thus far. Follow-up diagnostic studies revealed rigid fixation and fusion in all cases. The MR imaging-compatibility of the clamps allowed excellent follow-up studies with minimal artifact. Because of their ease of use, rigid stabilization, good results, lack of complications, and compatibility with MR imaging, the Halifax interlaminar clamp with bone grafting provides an ideal method for posterior cervical stabilization.

Mesenchymal proliferation with decidual-like morphology in seminal vesicles of aging mice.

Lesions characterized by spindle and epithelioid cells and nuclear progesterone receptors are described in seminal vesicles of four aging mice. The lesions of two mice also contain granular metrial gland (GMG)-like cells. The same cellular details are seen in the uterine decidual reaction and the similar urinary bladder lesion in mice, also called mesenchymal tumor. Therefore, it is hypothesized that these lesions in male accessory sex glands and the urinary bladders of aging male and female mice are of mesenchymal origin with the potential for differentiation along several pathways, leading especially to lesions with decidual-like morphology, but also to lesions which contain only spindle cells. The decidual hypothesis is further supported by the occurrence of round eosinophilic granules and focal necrosis, interpreted as a sign of regression in all these lesion types. The bilateral lesions of a fifth mouse consist of spindle cells and scar-like tissue, the latter suggesting regression, and lack epithelioid and GMG-like cells. In this case, verification of the diagnosis depends on the demonstration of progesterone receptors, absent in normal glands. Uterine decidual reactions during pregnancy are brought about by priming with progesterone/estrogen, initiation through the blastocyst, and maintenance through progesterone. Experiments by others show that priming may also occur through growth factors/growth hormone, initiation through prostaglandins, and maintenance through testosterone in mice. It is hypothesized that upon such stimulation, certain cells in male accessory sex glands and the urinary bladder, possibly derived from the Muellerian ducts or other subperitoneal tissue, appear to have the potential in mice of developing into spindle and epithelioid cells, including decidual-like cells. All published uterine decidual reactions and lesions with decidual-like morphology in other organs of mice stayed within the peritoneal coverage of their respective organ and did not metastasize despite their "anaplastic", tumor-like appearance. Thus, they should be considered non-neoplastic. It is proposed to name above lesions in male accessory sex glands and urinary bladders "mesenchymal proliferation, decidual type" or "mesenchymal proliferation, spindle-cell type", depending on their cellular characteristics.

Cleavage of a model peptide at its glycine residue by alkaline mercuric oxycyanide.

Treatment of peptides with excess HgO in the presence of alkaline cyanide leads to cleavage of the peptides at glycine residues. The reaction appears to involve both C- and N-mercuration with subsequent release of 2 mol mercury per mol of glycine. An intermediate glyoxylic acid residue in Schiff base linkage is postulated. Treatment of the heptapeptide Phe-Ala-Lys-Gly-Leu-Asp-Val with alkaline HgO and KCN for 6 h at 25 degrees resulted in greater than 90% cleavage, and the resultant reaction products were separated by reverse phase chromatography and identified by amino acid analysis. N-terminal products were approximately equimolar Phe-Ala-Lys, Phe-Ala-Lys-Gly, and Phe-Ala-Lys-amide. C-terminal products were predominantly Leu-Asp-Val (63%), plus Gly-Leu-Asp-Val (9%), and oxalyl-Leu-Asp-Val (8%). This method may be useful for cleavage of peptides or proteins containing glycine residues.

Left ventricular function during lethal and sublethal endotoxemia in swine.

Our previous studies suggested that after a median lethal dose (LD50) of endotoxin, cardiac contractility was depressed in nonsurviving dogs. The canine cardiovascular system is unlike humans in that dogs have a hepatic vein sphincter that is susceptible to adrenergic stimulation capable of raising hepatic and splanchnic venous pressures. We retested the hypothesis that lethality after endotoxin administration is associated with cardiac contractile depression in pigs, because the hepatic circulation in this species is similar to that of humans. We compared cardiac mechanical function of pigs administered a high dose (250 micrograms/kg) or a low dose (100 micrograms/kg) endotoxin by use of the slope of the end-systolic pressure-diameter relationship (ESPDR) as well as other measurements of cardiac performance. In all the pigs administered a high dose, ESPDR demonstrated a marked, time-dependent depression, whereas we observed no significant ESPDR changes after low endotoxin doses. The other cardiodynamic variables were uninterpretable, due to the significant changes in heart rate, end-diastolic diameter (preload), and aortic diastolic pressure (afterload). Plasma myocardial depressant factor activity accumulated in all endotoxin-administered animals, tending to be greater in the high-dose group. In this group, both subendocardial blood flow and global function were depressed, whereas pigs administered the low dose of endotoxin demonstrated slight, but nonsignificant, increases in flow and function. These observations indicate that myocardial contractile depression is associated with a lethal outcome to high doses of endotoxin. One possible mechanism for this loss of contractile function may be a relative hypoperfusion of the subendocardium.

Delayed cutaneous hypersensitivity reactions to concanavalin A-binding tumour membrane components.

The use of affinity chromatography has permitted the isolation of those tumour membrane fractions possessing affinity for the lectin concanavalin A. That these fractions are rich in tumour-associated antigens is supported by their ability to induce delayed cutaneous hypersensitivity reactions. The tumour extracts possessing concanavalin affinity resulted in skin test reactivity of considerably greater frequency and magnitude than normal tissue fractions, disrupted unfractionated tumour membrane extracts, or tumour membrane fractions not possessing concanavalin A affinity. Although the present data does not permit the correlation of skin reactivity with patient or disease parameters, the isolation of augmented concentrations of tumour-associated antigens may lead to improved diagnostic and prognostic tests and vaccines for patients with cancer.

Canine left ventricular performance during LD50 endotoxemia.

Previous reports of the effect of endotoxin shock on cardiac performance have not achieved uniform results. These discrepancies have possibly been caused by the use of indices of cardiac performance that may have been sensitive to altered heart rate or preconditions of cardiac contraction as well as altered cardiac performance. We tested the hypothesis that, following a median lethal dose (LD50) of E. coli endotoxin, cardiac performance would be diminished in nonsurviving animals and maintained in surviving animals. We elected to employ the analysis of the end-systolic pressure-diameter relationship (sigma ES) as well as other measurements of cardiac performance to test this hypothesis. We established that the sigma ES measurement was independent of increased and decreased afterload and relatively insensitive to altered heart rate. In the nonsurviving animals, sigma ES exhibited a marked depression following endotoxin a administration. In the surviving animals, sigma ES exhibited a nonsignificant decrease followed by a return toward preendotoxin values. All other cardiodynamic measurements were uninterpretable due to the marked changes in heart rate, peripheral vascular function, aortic pressure, and cardiac output. We conclude that, following endotoxin administration, those animals that exhibited a diminished myocardial contractility failed to survive more than 2.5 h postendotoxin, whereas the surviving animals were able to restore normal cardiac contractility. Thus survival of endotoxin administration is associated with the maintenance of normal cardiac contractility.

Segmentation of magnetic resonance images using an artificial neural network.

Signal intensities from intermediate and T2 weighted spin echo images of the brain were used as inputs into an artificial neural network (ANN). The signal intensities were used to train the network to recognize anatomically-important segments. The ANN was a self-organizing map (SOM) neural network which develops a continuous topographical map of the signal intensities within the two images. The neural network segmented images demonstrated good correlation with white matter, gray matter, and cerebral spinal fluid (CSF) spaces. This technique was rated better than manual thresholding of the intermediate images, but not as good as manual thresholding of the T2 weighted images.

Canine left ventricular function during experimental pancreatitis.

Left ventricular contractility following induction of experimental pancreatitis (EP) was studied. Contractility was evaluated by analyzing the left ventricular end systolic pressure-diameter relationship (sigma ES). Sigma ES is independent of large changes in preload, afterload, and heart rate, but sensitive to changes in ventricular contractility. Following injection of 100,000 IU trypsin in 4% taurocholate into the pancreas to induce EP, seven of eight dogs survived 5 hr. These dogs exhibited an initial significant reduction in mean arterial pressure (MABP) which stabilized at 90% of control at 3-5 hr post-EP. Cardiac output (CO) dropped slowly after EP induction (from 3.08 +/- 0.43 to 2.22 +/- 0.22 liters/min) associated with no significant change in peripheral resistance. Stroke work and stroke volume were markedly depressed reflecting the changes in MABP and CO. No consistent changes in +dP/dt or -dP/dt were observed. The ratio of endo/epicardial blood flow was unchanged as was blood Ca2+ levels throughout the experiment. Ventricular contractility as reflected by sigma ES tended to improve (from 49.7 to 69.6 mm Hg/mm at 4 hr following EP). Therefore, it was concluded that these animals exhibited no loss of ventricular contractility during EP.

Premortem biodistribution of radioactivity in the rat: measurement of blood and tissue activity of tracers used in clinical imaging studies.

Radioactive tracers are used in nuclear medicine imaging studies to detect sites of human disease. Use of animal models helps to establish tracer biodistribution kinetics and, thus, is critical to the early testing of radiopharmaceuticals. We developed a method to characterize the premortem temporal, spatial, and compartmental biodistribution of tracer molecules in the rat and used this method to study three tracers of potential value in detecting thromboembolic disease. Dynamic gamma scintigraphy was used to determine the spatial and temporal distribution of 99mTc-labeled IgG antifibrin antibody, Fab' fragment of antifibrin, and oxidized human serum albumin (OHSA). The blood pool compartment within each tissue was determined from the biodistribution of 131I-labeled bovine serum albumin injected prior to termination. The biodistribution of the blood compartment was maintained by immediately freezing the rat carcass in isotonic saline. Three-dimensional maps of tracer distribution in the tissue and blood compartments were then constructed from cross sections of the frozen tissue. These maps were used to relate necropsy tissue counts to premortem scintigraphic images. Over a 60-min interval after administration of tracer via a tail vein, significant differences in biodistribution were evident. The IgG remained within the blood pool, but there was rapid blood clearance of the OHSA molecules by the kidney and liver. The Fab' molecules were cleared more slowly by the kidney; Fab' molecules were found in the extravascular spaces, whereas IgG and OHSA were not found. The kinetics of OHSA and Fab' in organ regions paralleled changes in the blood compartment.(ABSTRACT TRUNCATED AT 250 WORDS)