Spherical aggregates of beta-amyloid (amylospheroid) show high neurotoxicity and activate tau protein kinase I/glycogen synthase kinase-3beta

beta-Amyloid (Abeta) acquires toxicity by self-aggregation. To identify and characterize the toxic form(s) of Abeta aggregates, we examined in vitro aggregation conditions by using large quantities of homogenous, chemically synthesized Abeta1-40 peptide. We found that slow rotation of Abeta1-40 solution reproducibly gave self-aggregated Abeta1-40 containing a stable and highly toxic moiety. Examination of the aggregates purified by glycerol-gradient centrifugation by atomic force microscopy and transmission electron microscopy revealed that the toxic moiety is a perfect sphere, which we call amylospheroid (ASPD). Other Abeta1-40 aggregates, including fibrils, were nontoxic. Correlation studies between toxicity and sphere size indicate that 10- to 15-nm ASPD was highly toxic, whereas ASPD <10 nm was nontoxic. A positive correlation between the toxicity and ASPD >10 nm also appeared to exist when Abeta1-42 formed ASPD by slow rotation. However, Abeta1-42-ASPD formed more rapidly, killed neurons at lower concentrations, and showed approximately 100-fold-higher toxicity than Abeta1-40-ASPD. The toxic ASPD was associated with SDS-resistant oligomeric bands in immunoblotting, which were absent in nontoxic ASPD. Because the formation of ASPD was not disturbed by pentapeptides that break beta-sheet interactions, Abeta may form ASPD through a pathway that is at least partly distinct from that of fibril formation. Inhibition experiments with lithium suggest the involvement of tau protein kinase I/glycogen synthase kinase-3beta in the early stages of ASPD-induced neurodegeneration. Here we describe the identification and characterization of ASPD and discuss its possible role in the neurodegeneration in Alzheimer's disease.     

Hemodynamic Changes by Drug Interaction of Adrenaline With Chlorpromazine

Adrenaline (epinephrine) is included in dental local anesthesia for the purpose of vasoconstriction. In Japan, adrenaline is contraindicated for use in patients receiving antipsychotic therapy, because the combination of adrenaline and an antipsychotic is considered to cause severe hypotension; however, there is insufficient evidence supporting this claim. The purpose of the present study was to clarify the changes in hemodynamics caused by drug interaction between adrenaline and an antipsychotic and to evaluate the safety of the combined use of adrenaline and an antipsychotic in an animal study. Male Sprague-Dawley rats were anesthetized with sodium pentobarbital. A catheter was inserted into the femoral artery to measure blood pressure and pulse rate. Rats were pretreated by intraperitoneal injection of chlorpromazine or chlorpromazine and propranolol, and after 20 minutes, saline or 1 of 3 different doses of adrenaline was administered by intraperitoneal injection. Changes in the ratio of mean arterial blood pressure and pulse rate were measured after the injection of adrenaline. Significant hypotension and tachycardia were observed after the injection of adrenaline in the chlorpromazine-pretreated rats. These effects were in a dose-dependent manner, and 100 μg/kg adrenaline induced significant hemodynamic changes. Furthermore, in the chlorpromazine and propranolol–pretreated rats, modest hypertension was induced by adrenaline, but hypotension and tachycardia were not significantly shown. Hypotension was caused by a drug interaction between adrenaline and chlorpromazine through the activation of the β-adrenergic receptor and showed a dose-dependent effect. Low-dose adrenaline similar to what might be used in human dental treatment did not result in a significant homodynamic change.Key Words: Local anesthesia, Drug interaction, Adrenaline, Epinephrine, AntipsychoticsVasoconstrictors are included in local anesthetics to increase the duration of anesthesia, to prevent local anesthesia systemic toxicity, and to promote hemostasis in the local operative field.¹ Adrenaline (epinephrine) is the most common vasoconstrictor that is added to local anesthetics, and lidocaine and articaine with adrenaline are the most widely used local anesthetics for dental treatment. However, adrenaline has drug interactions with some medicines. It is widely believed that the combination of adrenaline with an antipsychotic can cause hypotension because antipsychotics have α-adrenergic blocker effects that can result in decreased peripheral resistance, and adrenaline has been shown to have significant β2-adrenergic mediated vasodilatory effects that also can cause hypotension. Therefore, the combination of adrenaline and an antipsychotic is contraindicated in Japan for fear of causing significant hypotension. However, there is insufficient evidence supporting this claim, and, in clinical settings, there are many situations in which the use of local anesthetics with adrenaline is appropriate for dental treatment of patients receiving antipsychotic therapy. Thus, the purpose of the present study was to clarify the changes in hemodynamics caused by a drug interaction between adrenaline and an antipsychotic and to evaluate the safety of the combined use of adrenaline with an antipsychotic in an animal study.     

Regulation of activity of P2X7 receptor by its splice variants in cultured mouse astrocytes

Of purinergic receptors, P2X7 receptor (P2X7R, defined as a full‐length receptor) has unique characteristics, and its activation leads to ion channel activity and pore formation, causing cell death. Previously, we demonstrated that P2X7R expressed by nonstimulated astrocyte cultures obtained from SJL‐strain mice exhibits constitutive activation, implying its role in maintenance of cellular homeostasis. To obtain novel insights into its physiological roles, we examined whether constitutive activation of P2X7R is regulated by expression of its splice variants in such resting astrocytes, and whether their distinct expression profiles in different mouse strains affect activation levels of astrocytic P2X7Rs. In SJL‐ and ddY‐mouse astrocytes, spontaneous YO‐PRO‐1 uptake, an indicator of pore activity of P2X7R, was detected, but the uptake by the formers was significantly greater than that by the latter. Between the two mouse strains, there was a difference in their sensitivity of YO‐PRO‐1 uptake to antagonists, but not in the expression levels and sequences of P2X7R and pannexin‐1. Regarding expression of splice variants of P2X7R, expression of P2X7R variant‐3 (P2X7R‐v3) and ‐4 (P2X7R‐v4), but not variant‐2 and ‐k, was lower in SJL‐mouse astrocytes than in ddY‐mouse ones. On transfection of P2X7R‐v3 and ‐v4 into SJL‐mouse astrocytes, the pore activity was attenuated as in the case of the HEK293T cell‐expression system. These findings demonstrate that basal activity of P2X7R expressed by resting astrocytes is negatively regulated by P2X7R‐v3 and ‐v4, and that their distinct expression profiles result in the different activation levels of astrocytic P2X7Rs in different mouse strains. GLIA 2014;62:440–451     

Prostaglandin E2 and its receptor EP2 trigger signaling that contributes to YAP‐mediated cell competition

Cell competition is a biological process by which unfit cells are eliminated from “cell society.” We previously showed that cultured mammalian epithelial Madin‐Darby canine kidney (MDCK) cells expressing constitutively active YAP were eliminated by apical extrusion when surrounded by “normal” MDCK cells. However, the molecular mechanism underlying the elimination of active YAP‐expressing cells was unknown. Here, we used high‐throughput chemical compound screening to identify cyclooxygenase‐2 (COX‐2) as a key molecule triggering cell competition. Our work shows that COX‐2‐mediated PGE₂ secretion engages its receptor EP2 on abnormal and nearby normal cells. This engagement of EP2 triggers downstream signaling via an adenylyl cyclase‐cyclic AMP‐PKA pathway that, in the presence of active YAP, induces E‐cadherin internalization leading to apical extrusion. Thus, COX‐2‐induced PGE₂ appears a warning signal to both abnormal and surrounding normal cells to drive cell competition.     

Age-dependent association of mannose-binding lectin polymorphisms with the development of pulmonary tuberculosis in Viet Nam

Mannose-binding lectin (MBL) binds to pathogens and induces complement-mediated opsonophagocytosis. Although the association between MBL2 polymorphisms and tuberculosis (TB) has been studied in various populations, the results are controversial. We explored the stages of TB associated with MBL2 polymorphisms. X/Y (rs7096206) and A/B (rs1800450) were genotyped in 765 new patients with active pulmonary TB without HIV infection and 556 controls in Hanoi, Viet Nam. The MBL2 nucleotide sequences were further analyzed, and plasma MBL levels were measured in 109 apparently healthy healthcare workers and 65 patients with TB. Latent TB infection (LTBI) was detected by interferon-gamma release assay (IGRA). The YA/YA diplotype, which exhibited high plasma MBL levels, was associated with protection against active TB in younger patients (mean age = 32) ≦ 45 years old (odds ratio, 0.61; 95% confidence interval, 0.46–0.80). The resistant diplotype was less frequently found in the younger patients at diagnosis (P = 0.0021). MBL2 diplotype frequencies and plasma MBL levels were not significantly different between the IGRA-positive and -negative groups. MBL2 YA/YA exhibited a protective role against the development of TB in younger patients, whereas the MBL2 genotype and MBL levels were not associated with LTBI. High MBL levels may protect against the early development of pulmonary TB after infection.     

Imaginal cell-specific accumulation of the multicatalytic proteinase complex (proteasome) during post-embryonic development in the tobacco hornworm,Manduca sexta

The multicatalytic proteinase complex is a multi-subunit, high molecular weight proteinase present in the nucleus and cytoplasm of eukaryotic cells. This catalytic complex is involved in diverse cellular functions as part of the ubiquitin proteolysis system, including non-lysosomal proteolysis, antigen presentation, cell cycle progression, and cell proliferation, and in the programmed death of intersegmental muscles after adult eclosion in the tobacco hornworm moth, Manduca sexta. We have investigated the distribution of the multicatalytic proteinase complex in the central nervous system of this moth. At all stages of post-embryonic development, most cell types exhibited consistent, low levels of cytoplasmic and nuclear immunoreactivity for the multicatalytic proteinase complex. High levels of cell-specific accumulation of the complex were, however, demonstrated in abdominal neurosecretory cells and in imaginal cells in the larval brain, the larval segmental ganglia, and the developing wing discs. Imaginal cells exhibited intense immunoreactivity for the multicatalytic proteinase complex only until the onset of terminal differentiation. Intersegmental muscles undergoing programmed cell death exhibited intense cytoplasmic immunoreactivity for the multicatalytic proteinase, while persisting flight muscles and dying neurons were characterized by basal levels of staining. These staining patterns suggest that the multicatalytic proteinase of Manduca sexta serves multiple functions and is associated with the period of developmental arrest displayed by imaginal cells prior to metamorphosis. © 1996 Wiley-Liss, Inc.     

Transurethral microwave thermotherapy for benign prostatic hyperplasia: Relation between clinical response and prostate histology

The effectiveness of transurethral microwave thermotherapy (TUMT) for BPH has been confirmed. To identify the characteristics of the ideal candidate, retrospective analysis and morphometric study of prostatic tissue were performed. Forty-two patients with symptomatic BPH were included in the study; these comprised 10 patients treated for more than 3 months with anti-androgen pre-TUMT (group A) and 32 fresh cases (group B). Subjective and objective responses were evaluated at 2 months post TUMT. In 12 fresh cases who underwent pre-TUMT biopsy of the prostate, the stromal-to-epithelial ratio was determined via quantitative image analysis on a computer-assisted morphometry system. No significant differences in baseline patient characteristics were found between the two groups: age, prostate volume, peak flow rate (PFR), or International Prostate Symptom Score (I-PSS). However, significant differences in treatment outcome were found between the two groups (group A vs. group B, respectively): total energy delivered to the prostate: 96 kJ vs. 125 kJ; I-PSS decrease from baseline: 5.9 vs. 11.6; PFR increase from baseline: 1.1 vs. 4.7 ml/sec. There was a positive correlation between the I-PSS change from baseline and the stromal-to-epithelial ratio of the prostatic tissue (r = 0.4857). The results suggest that microwave interacts poorly with the prostate due to the artificially created “lack” of glandular tissue. The morphometric study also supports the contention that the histological composition of the prostatic tissue plays an important role in terms of microwave thermal interactions and treatment outcome. © 1996 Wiley-Liss, Inc.