Molecular evidence sterile tissue damage during pathogenesis of pododermatitis aseptica hemorrhagica circumscripta is associated with disturbed epidermal-dermal homeostasis.

Podermatitis aseptica hemorrhagica circumscripta is associated with metalloproteinase 2 weakening of distal phalangeal suspensory structures and sinkage of the distal phalanx in the claw capsule. Pressure from the tuberculum flexorium on the sole epidermis and dermis produces hemorrhagic tissue injury and defective horn production appearing as yellow-red, softened claw horn in region 4 of the sole. A model of the MAPK/ERK signal cascade orchestrating epidermal-dermal homeostasis was employed to determine if sterile inflammatory responses are linked to disturbed signal transduction for epidermal homeostasis in sole epidermis and dermis. The objective was to assess shifts in target genes of inflammation, up- and downstream MAPK/ERK signal elements, and targeted genes supporting epidermal proliferation and differentiation. Sole epidermis and dermis was removed from lateral claws bearing lesions of podermatitis aseptica hemorrhagica circumscripta, medial claws from the same limb and lateral claws from completely normal limbs of multiparous, lactating Holstein cows. The abundance levels of targeted transcripts were evaluated by real-time QPCR. Lesion effects were assessed by ANOVA, and mean comparisons were performed with t-tests to assess variations between mean expression in ulcer-bearing or medial claw dermis and epidermis and completely normal lateral claw dermis and epidermis or between ulcer-bearing dermis and epidermis and medial claw dermis and epidermis. The lesions were sterile and showed losses across multiple growth factors, their receptors, several downstream AP1 transcription components, CMYC, multiple cell cycle and terminal differentiation elements conducted by MAPK/ERK signals and ß 4, α 6 and collagen 17A hemidesmosome components. These losses coincided with increased cytokeratin 6, ß 1 integrin, proinflammatory metalloproteinases 2 and 9, IL1B and physiologic inhibitors of IL1B, the decoy receptor and receptor antagonist. Medial claw epidermis and dermis from limbs with lateral claws bearing podermatitis aseptica hemorrhagica circumscripta showed reductions in upstream MAPK/ERK signal elements and downstream targets that paralleled those in hemorrhagic lesions. Inhibitors of IL1B increased in the absence of real increases in inflammatory targets in the medial claw dermis and epidermis. Losses across multiple signal path elements and downstream targets were associated with negative effects on targeted transcripts supporting claw horn production and wound repair across lesion-bearing lateral claws and lesion-free medial claw dermis and epidermis. It was unclear if the sterile inflammation was causative or a consequence of these perturbations.

Transcriptional responses consistent with perturbation in dermo-epidermal homeostasis in septic sole ulceration.

The aim of this study was to evaluate transcriptional changes in sole epidermis and dermis of bovine claws with septic sole ulceration of the lateral claw. Assessment included changes in transcripts orchestrating epidermal homeostatic processes including epidermal proliferation, differentiation, inflammation, and cell signaling. Sole epidermis and dermis was removed from region 4 of lesion-bearing lateral and lesion-free medial claws of pelvic limbs in multiparous, lactating Holstein cows. Control sole epidermis and dermis was obtained from region 4 of lateral claws of normal pelvic limbs. Transcript abundances were evaluated by real-time QPCR and relative expression analyzed by ANOVA. Relative to normal lateral claws, sole epidermis and dermis in ulcer-bearing claws exhibited downregulation of genes associated with growth factors, growth factor receptors, activator protein 1 (AP-1) and proto-oncogene (CMYC) transcription components, cell cycle elements, lateral cell-to-cell signaling elements and structures of early and late keratinocyte differentiation. These changes were accompanied by upregulation of pro-inflammatory transcripts interleukin 1 α (IL1A), interleukin1 ß (IL1B), interleukin 1 receptor 1 (IL1R1), inducible nitric oxide synthase (NOS2), the inflammasome components NOD like receptor protein 3 (NLRP3), pyrin and caspase recruitment domain (PYCARD), and caspase-1 interleukin converting enzyme (CASPASE), the matrix metalloproteinases (MMP2 and MMP9), and anti-inflammatory genes interleukin 1 receptor antagonist (IL1RN) and interleukin1 receptor 2 (IL1R2). Transcript abundance varied across epidermis and dermis from the ulcer center, margin and epidermis and dermis adjacent to the lesion. Sole epidermis and dermis of lesion-free medial claws exhibited changes paralleling those in the adjacent lateral claws in an environment lacking inflammatory transcripts and downregulated IL1A, interleukin 18 (IL18), tumor necrosis factor α (TNFA) and NOS2. These data imply perturbations in signal pathways driving epidermal proliferation and differentiation are associated with, but not inevitably linked to epidermis and dermis inflammation. Further work is warranted to better define the role of crushing tissue injury, sepsis, metalloproteinase activity, and inflammation in sole ulceration.

Novel Strongly Spin-Orbit Coupled Quantum Dimer Magnet: Yb_{2}Si_{2}O_{7}.

The quantum dimer magnet (QDM) is the canonical example of quantum magnetism. The QDM state consists of entangled nearest-neighbor spin dimers and often exhibits a field-induced triplon Bose-Einstein condensate (BEC) phase. We report on a new QDM in the strongly spin-orbit coupled, distorted honeycomb-lattice material Yb_{2}Si_{2}O_{7}. Our single crystal neutron scattering, specific heat, and ultrasound velocity measurements reveal a gapped singlet ground state at zero field with sharp, dispersive excitations. We find a field-induced magnetically ordered phase reminiscent of a BEC phase, with exceptionally low critical fields of H_{c1}∼0.4 and H_{c2}∼1.4 T. Using inelastic neutron scattering in an applied magnetic field we observe a Goldstone mode (gapless to within δE=0.037 meV) that persists throughout the entire field-induced magnetically ordered phase, suggestive of the spontaneous breaking of U(1) symmetry expected for a triplon BEC. However, in contrast to other well-known cases of this phase, the high-field (µ_{0}H≥1.2 T) part of the phase diagram in Yb_{2}Si_{2}O_{7} is interrupted by an unusual regime signaled by a change in the field dependence of the ultrasound velocity and magnetization, as well as the disappearance of a sharp anomaly in the specific heat. These measurements raise the question of how anisotropy in strongly spin-orbit coupled materials modifies the field induced phases of QDMs.

Rate heterogeneity across Squamata, misleading ancestral state reconstruction and the importance of proper null model specification.

The binary-state speciation and extinction (BiSSE) model has been used in many instances to identify state-dependent diversification and reconstruct ancestral states. However, recent studies have shown that the standard procedure of comparing the fit of the BiSSE model to constant-rate birth-death models often inappropriately favours the BiSSE model when diversification rates vary in a state-independent fashion. The newly developed HiSSE model enables researchers to identify state-dependent diversification rates while accounting for state-independent diversification at the same time. The HiSSE model also allows researchers to test state-dependent models against appropriate state-independent null models that have the same number of parameters as the state-dependent models being tested. We reanalyse two data sets that originally used BiSSE to reconstruct ancestral states within squamate reptiles and reached surprising conclusions regarding the evolution of toepads within Gekkota and viviparity across Squamata. We used this new method to demonstrate that there are many shifts in diversification rates across squamates. We then fit various HiSSE submodels and null models to the state and phylogenetic data and reconstructed states under these models. We found that there is no single, consistent signal for state-dependent diversification associated with toepads in gekkotans or viviparity across all squamates. Our reconstructions show limited support for the recently proposed hypotheses that toepads evolved multiple times independently in Gekkota and that transitions from viviparity to oviparity are common in Squamata. Our results highlight the importance of considering an adequate pool of models and null models when estimating diversification rate parameters and reconstructing ancestral states.

Monopolar and dipolar relaxation in spin ice Ho2Ti2O7.

Ferromagnetically interacting Ising spins on the pyrochlore lattice of corner-sharing tetrahedra form a highly degenerate manifold of low-energy states. A spin flip relative to this "spin-ice" manifold can fractionalize into two oppositely charged magnetic monopoles with effective Coulomb interactions. To understand this process, we have probed the low-temperature magnetic response of spin ice to time-varying magnetic fields through stroboscopic neutron scattering and SQUID magnetometry on a new class of ultrapure Ho2Ti2O7 crystals. Covering almost 10 decades of time scales with atomic-scale spatial resolution, the experiments resolve apparent discrepancies between prior measurements on more disordered crystals and reveal a thermal crossover between distinct relaxation processes. Magnetic relaxation at low temperatures is associated with monopole motion through the spin-ice vacuum, while at elevated temperatures, relaxation occurs through reorientation of increasingly spin-like monopolar bound states. Spin fractionalization is thus directly manifest in the relaxation dynamics of spin ice.

Radioimmunodetection of gastrointestinal neoplasms with antibodies to carcinoembryonic antigen.

Primary and secondary gastrointestinal tumors have been identified using sheep immunoglobulin G antibody to carcinoembryonic antigen radiolabeled with 131I. 99mTc-pertechnetate and 99mTc-human serum albumin were used to identify tissue spaces and blood pool and to facilitate external substraction imaging. In 13 patients with tumors, 4 of 5 primary sites and 8 of 11 secondary sites were successfully demonstrated. Two patients with benign disease had negative scans. Comparison with conventional methods of scanning showed good correlation.

Imatinib for systemic mast-cell disease.

Imatinib has shown to be effective against malignant disease driven by ckit. We prospectively treated 12 adults with symptomatic systemic mast-cell disease at a dose of either 100 mg or 400 mg per day. Of the ten patients who we could assess for response, five (50%) had a measurable response to the drug, four of whom had important mast-cell cytoreduction and two who had complete clinical and histological remission. In the five patients with eosinophilia, three had complete clinical and haematological remission. The other two, who did not respond to treatment, were the only patients with the ckit D816V mutation. Our results suggest that imatinib either inhibits the growth-promoting role of wild type ckit, or targets an oncogenic kinase.

AraC protein can activate transcription from only one position and when pointed in only one direction.

At the araBAD promoter, the RNA polymerase-proximal half-site for AraC binding partially overlaps the -35 region. Random and explicit spacing experiments show that both this partial overlapping and AraC binding to the polymerase-proximal half-site are necessary and sufficient for strong transcriptional activation. Normally, this occupancy is generated by the presence of arabinose, which shifts AraC from a DNA looping interaction involving the polymerase-distal half-site and the araO2 site 210 base-pairs away, to an interaction with the two half-sites adjacent to RNA polymerase. Changing the polymerase-proximal half-site to a higher affinity AraC binding site gives activation in the absence of arabinose. Thus, arabinose is not required to transform AraC into an activating conformation. Because the two half-sites of araI are direct repeats, the RNA polymerase proximal and distal surfaces of AraC are not identical. When the araI site was turned around, no spacings were found from which AraC could activate transcription. In light of the strict spacing and orientation requirements for AraC activation, the interactions between AraC and RNA polymerase are likely to be specific and inflexible.

Transcription activation parameters at ara pBAD.

We studied the formation of open complexes of RNA polymerase and promoter DNA as activated by the AraC protein at the Escherichia coli araBAD promoter pBAD and by the cyclic AMP receptor protein at the galKTE promoter P1. The DNA migration retardation assay was demonstrated to be suitable for the detection and quantitation of open complexes by the correspondence in the properties of open complexes in solution and retarded complexes observed in gels. These included, on the ara promoter, heparin resistance, lifetime, DNAseI footprinting, exonuclease III footprinting, permanganate footprinting and disappearance upon transcription, and on the gal promoter, the correspondence between the kinetic parameters Kd and k2 obtained with established techniques and those obtained with the migration retardation assay. On the pBAD promoter we obtained kinetic parameters of Kd = 0.3 nM and K2 = 1 minute(-1). The unusually tight binding of polymerase in the presence of AraC suggests that AraC binds polymerase tightly.

In vitro antiproliferative activity of the farnesyltransferase inhibitor R115777 in hematopoietic progenitors from patients with myelofibrosis with myeloid metaplasia.

R115777 is an orally bioavailable farnesyltransferase inhibitor (FTI) that has displayed encouraging activity in patients with acute myeloid leukemia. To determine whether R115777 might exert similar activity in myelofibrosis with myeloid metaplasia (MMM), we evaluated its effects on circulating myeloid progenitor cells from patients with MMM (n=25) using in vitro colony-forming assays. The median R115777 concentrations that inhibited colony formation by 50% were 34 and 2.7 nM for myeloid and megakaryocytic colonies from MMM patients, respectively. Progenitors from normal controls and patients with other myeloproliferative disorders demonstrated similar sensitivity. Since the ras polypeptides are one putative target of FTIs, the potential role of ras effectors was examined by incubating parallel progenitor assays with the phosphatidyl-inositol-3 (PI-3) kinase inhibitor LY294002 and the mitogen-activated protein kinase 1 inhibitor PD98059. MMM progenitor colonies (n=7) were highly sensitive to LY294002 but not to PD98059, implying that the PI-3 kinase pathway may be critical for survival and proliferation of these cells. In addition to indicating that MMM progenitors are sensitive to clinically achievable R115777 concentrations in vitro, these results provide a potential explanation for the thrombocytopenia observed with R115777 during the treatment of other hematologic malignancies.

An evaluation of human neurophysin production in Alzheimer's disease: preliminary observations.

The concentrations of human neurophysins in the cerebrospinal fluid (CSF) of nine patients with Alzheimer's disease: Preliminary observations. (AD), and one patients with Pick's disease, were determined using specific radioimmunoassays (RIAs). Concentrations of vasopressin and oxytocin were also measured. Values were compared with those from 20 age-matched mentally normal individuals who were being treated for back pain. CSF levels of vasopressin-associated human neurophysin (VP-HNP) and oxytocin-associated human neurophysin (OT-HNP) in patients with AD (22 +/- 4 fmol/ml and 104 +/- 17 fmol/ml) were only 42% and 58% of those in the control subjects (p less than 0.0001, p less than 0.0004). Vasopressin levels for these patients (3.6 +/- 0.4 fmol/ml) were also significantly reduced to 51% of controls (p less than 0.007) and oxytocin levels were marginally (p = 0.092) reduced to 70% of controls. Because neurophysins and neuropeptides are gene-related products of vasopressin-neurons and oxytocin-neurons, the data indicate that these neurons are functionally impaired in patients with AD. Plasma neurophysin values suggest this impairment is confined to neurons with centrally-directed axons. Data from the one patient with Pick's disease demonstrates that reduced CSF levels of neurophysins and hormones is not confined to Alzheimer-type dementia.

Continuous ICV infusion of scopolamine impairs sustained attention of rhesus monkeys.

Systemic administration of anticholinergic agents impairs cognitive performance in animals and man. The anticholinergic, scopolamine, has profound effects on peripheral and central cholinergic function, making interpretation of its effects on cognitive performance difficult. To circumvent this problem, scopolamine was administered directly to the central nervous system of rhesus monkeys using a subcutaneously implanted infusion pump connected to a cannulae directed toward the right lateral ventricle. Intracerebroventricular (ICV) infusion of scopolamine (0.004, 0.012, 12.5, and 40.0 micrograms/kg/h) produced a dose-dependent decrease in the number of responses on a continuous performance task. Response decrements produced by scopolamine were seen mainly during the last half of the test session and at short stimulus durations. These data suggest that scopolamine produces a deficit in sustained attention or slowing of information processing that is mediated through direct central cholinergic blockade in the rhesus monkey.

The spectrum of imaging and neuropsychological findings in Pick's disease.

To seek improved methods for the diagnosis of Pick's disease, we reviewed imaging studies of four women and two men (ages 48 to 65 years at onset) and psychometric testing of three of them with autopsy or biopsy-proved Pick's. The presence of Pick bodies was required for the diagnosis. Seven patients with biopsy-proved Alzheimer's disease served as a comparison group. In the Pick's patients, CT in five of six showed marked frontal pole or temporal pole atrophy, which clearly differed from the pattern of cerebral atrophy seen in the Alzheimer's patients. Psychometric testing showed performance patterns that tended to differ from those of the Alzheimer's patients in that recent memory was relatively preserved despite marked impairment of executive functions. The distinctive psychometric pattern in the Pick's patients was evanescent, however. Thus, there were imaging and psychometric findings of potential diagnostic value for Pick's disease, but, for different reasons, they were imperfect.

Bone marrow mast cell immunophenotyping in adults with mast cell disease: a prospective study of 33 patients.

The aberrant co-expression of CD2 and CD25 antigens is the immunophenotypic hallmark of neoplastic mast cells, and has been consistently identified on bone marrow mast cells from patients with indolent mast cell disease (MCD). We prospectively analyzed the bone marrow mast cell immunophenotype by multiparametric flow cytometry (FC) for 33 MCD cases, to examine the role of CD2 and CD25 expression in establishing diagnosis, detecting histologically occult bone marrow mast cell infiltration, and assessing treatment response. While CD25 was almost uniformly expressed, only 6 of 13 patients with indolent MCD, 1 of 8 with aggressive MCD, 2 of 7 with MCD and an associated hematological disorder, and none of the 2 patients with either mast cell leukemia or smoldering systemic mastocytosis, expressed CD2. One of three patients with cutaneous mastocytosis had an aberrant CD2+/CD25+ mast cell population suggesting histologically occult bone marrow involvement. CD25 expression was lost in one patient who achieved complete histologic remission with therapy, but not in two patients who achieved a partial remission. In conclusion, CD25, but not CD2, is a reliable marker for neoplastic mast cells, and CD25 expression indicates histologically occult bone marrow infiltration and residual disease after therapy.

Continuous intratumoral infusion of methotrexate for recurrent glioblastoma: a pilot study.

Five patients with documented recurrences of glioblastoma multiforme were given continuous infusions of methotrexate delivered intratumorally using implantable catheters and subcutaneous refillable pumps. A continuous infusion of methotrexate (1 mg/d) was begun with concomitant oral administration of folinic acid. The methotrexate dose was increased every 2 weeks to 3, 10, 30, and, ultimately, 75 mg/d in two patients. Samples of serum and ventricular cerebrospinal fluid (CSF) were obtained to determine the levels of methotrexate and total bioactive folates, and brain tissue was obtained from two patients for determination of methotrexate concentration. The patients survived from 7 to 49 weeks after the implantation of the infusion device. Neither the clinical examination nor sequential radiological studies gave clear evidence of reduction in tumor size. Pneumonia developed in one patient, and mild hepatitis and increased seizure frequency in another. Methotrexate was stable in the delivery system over 12 days, and ventricular CSF reached steady-state levels by 5 days. Steady-state ventricular CSF levels of methotrexate were higher than serum levels in some patients, while the reverse was true in others. Levels of total bioactive folates in the CSF did not increase above the normal range. Methotrexate concentrations were highest at the center of the tumor, but measurable amounts of methotrexate were detectable in all areas of the brain. At autopsy in four patients, variable liquefactive necrosis of the brain tumors was seen, and viable tumor was found at the periphery of the tumor bed. These preliminary results suggest that it is technically feasible to infuse methotrexate into brain tumor cavities, and show that little central nervous system or systemic toxicity was encountered in five patients. Better delineation of the safety and efficacy of this therapeutic approach will require further clinical trials.

Preliminary report: intracranial cholinergic drug infusion in patients with Alzheimer's disease.

After toxicity studies in dogs, a preliminary feasibility trial of the continuous intracranial infusion of a muscarinic agonist was begun in four patients with biopsy-documented Alzheimer's disease. During the last 8 months, a totally implantable infusion system has been used to deliver bethanechol chloride into the cerebrospinal fluid of these patients at doses of 0.05 to 0.7 mg/day. Complications have been few and resolved spontaneously or were easily reversible. The subjective response to this treatment has been encouraging, with reports of improved cognitive and social function during drug infusion and a return to base line function with single-blind saline placebo infusions. Obviously, further evaluation will be necessary to demonstrate the efficacy of this treatment, and a double-blind placebo-controlled crossover trial is now being done. However, we think the preliminary results are encouraging and warrant the consideration of this approach as a potential treatment in patients with Alzheimer's disease.

Intracerebroventricular bethanechol chloride infusion in Alzheimer's disease. Results of a collaborative double-blind study.

The use of intracerebroventricular bethanechol chloride infusion in patients with Alzheimer's disease was first reported in 1984. An initial trial in four patients demonstrated the feasibility of this approach for cholinergic drug delivery to the brain, but objective improvement in cognitive function was not documented. A collaborative placebo-controlled double-blind crossover study has now been carried out in 49 patients with biopsy-documented Alzheimer's disease. The results demonstrate a statistical improvement in Mini-Mental State scores and significantly slower performance on Trails A testing during drug infusion. Other neuropsychological test scores were not similarly affected. The degree of improvement was not sufficient to justify further treatment of Alzheimer's disease patients by intracerebroventricular infusion of bethanechol chloride. The drug delivery system used in the study was well tolerated, with two irreversible complications in more than 50,000 patient days.

When the safety net is unsafe: real-time assessment of the overcrowded emergency department.

Although much work has been done evaluating causes for increased demand for emergency department (ED) services, few ways are available to help determine that an individual ED is overcrowded. Four calculations are proposed using real-time data for accurately diagnosing an ED with potential for failing both as a safety net and as a source for quality health care. The bed ratio (BR) accounts for the number of patients in relation to the available treatment spaces. The BR is obtained by adding the current number of ED patients to the predicted arrivals minus the predicted departures and dividing the result by the total number of treatment spaces. The acuity ratio (AR) measures the relative burden of illness in the ED. The AR is the average triage category of all patients in the ED. The provider ratio (PR) determines the volume of patients that can be evaluated and treated by the physician providers. The PR is found by dividing the arrivals per hour by the sum of the average patients per hour usually disposed for each provider on duty. From these ratios, the demand value (DV) is calculated, which gives an overall measure of current demand. The DV is found by taking the sum of the BR and PR and multiplying by the AR. A DV of more than 7 should initiate a specific assessment of the individual ratios in order to accurately diagnose the problem and institute action. Based on the values, predetermined processes can be instituted to help remedy the overcrowded situation. Trended over time, the ratios can provide the data needed for better resource assessment, planning, and allocation.