BACE1 inhibition induces a specific cerebrospinal fluid β-amyloid pattern that identifies drug effects in the central nervous system

BACE1 is a key enzyme for amyloid-β (Aβ) production, and an attractive therapeutic target in Alzheimer's disease (AD). Here we report that BACE1 inhibitors have distinct effects on neuronal Aβ metabolism, inducing a unique pattern of secreted Aβ peptides, analyzed in cell media from amyloid precursor protein (APP) transfected cells and in cerebrospinal fluid (CSF) from dogs by immunoprecipitation-mass spectrometry, using several different BACE1 inhibitors. Besides the expected reductions in Aβ1-40 and Aβ1-42, treatment also changed the relative levels of several other Aβ isoforms. In particular Aβ1-34 decreased, while Aβ5-40 increased, and these changes were more sensitive to BACE1 inhibition than the changes in Aβ1-40 and Aβ1-42. The effects on Aβ5-40 indicate the presence of a BACE1 independent pathway of APP degradation. The described CSF Aβ pattern may be used as a pharmacodynamic fingerprint to detect biochemical effects of BACE1-therapies in clinical trials, which might accelerate development of novel therapies.     

Retrospective dosimetry using OSL of tooth enamel and dental repair materials irradiated under wet and dry conditions

Following a radiological or nuclear emergency event, there is a need for quick and reliable dose estimations of potentially exposed people. In situations where dosimeters are not readily available, the dose estimations must be carried out using alternative methods. In the present study, the optically stimulated luminescence (OSL) properties of tooth enamel and different dental repair materials have been examined. Specimens of the materials were exposed to gamma and beta radiation in different types of liquid environments to mimic the actual irradiation situation in the mouth. Measurements were taken using a Ris? TL/OSL reader, and irradiations were made using a ⁹⁰Sr/⁹⁰Y source and a linear accelerator (6 MV photons). Results show that the OSL signal from tooth enamel decreases substantially when the enamel is kept in a wet environment. Thus, tooth enamel is not reliable for retrospective dose assessment without further studies of the phenomenon. Dental repair materials, on the other hand, do not exhibit the same effect when exposed to liquids. In addition, dose–response and fading measurements of the dental repair materials show promising results, making these materials highly interesting for retrospective dosimetry. The minimum detectable dose for the dental repair materials has been estimated to be 20–185?mGy.     

Genetic differences in internal transcribed spacer 1 between Dermanyssus gallinae from wild birds and domestic chickens

We investigated the presence of the poultry red mite or the chicken mite, Dermanyssus gallinae De Geer, Acari: Dermanyssidae, in wild bird populations in four different geographical regions of Sweden. The mites identified as D. gallinae were compared genetically with D. gallinae from egg-producing poultry farms in the same regions. The small subunit (SSU) gene, the 5.8S ribosomal RNA (rRNA) gene and the two internal transcribed spacers (ITS) of the rRNA genes were used in the genetic analysis. All D. gallinae mites had identical SSU rRNA, 5.8S rRNA and ITS2 sequences independent of their origin. By contrast, we identified significant differences in the ITS1 sequences. Based on the differences in the ITS1 sequences, the mites could be divided into two genotypes, of wild and domesticated origin, with no variation within the groups. These results imply that wild bird populations are of low importance, if any, as natural reservoirs of D. gallinae in these four geographical regions of Sweden.     

Aminomethyl-2,6-difluorophenols as a novel class of increased lipophilicity GABA(C) receptor antagonists

3- and 4-(Aminomethyl)-2,6-difuorophenols were tested for activity against the three major classes of GABA receptors. 4-(Amninomethyl)-2,6difluorophenol was shown to be a competitive and somewhat selective antagonist at p1 GABA(C) receptors expressed in Xenopus oocytes (K(B) = 75.5 microM with a 95% Confidence Interval range of 75.2 microM to 75.8 microM). This is the first in a novel class of increased lipophilicity GABA(C) receptor antagonists with little activity at alpha1beta2gamma2 GABA(A) and GABA(B) receptors.     

Acute psychological stress raises plasma ghrelin in the rat

Ghrelin is produced by the A-like cells of the stomach and mobilized by food deprivation. It was reported recently that acute psychological stress increases ghrelin gene expression in rat oxyntic mucosa. The aim of this study was to examine the effect of such stress on circulating ghrelin levels. To this end, we measured plasma ghrelin in Wistar Kyoto (WKY) rats (a high-anxiety strain) and Sprague-Dawley (SPD) rats (a low-anxiety strain), exposed to water avoidance stress for 60 min. Blood was collected before and after the stress. Acute stress increased the plasma ACTH concentration approximately 5-fold (p<0.01) in both strains of rats, while plasma ghrelin increased by 85% (p<0.01) in the SPD rats and by 40% (p<0.001) in the WKY rats. Ghrelin levels after acute stress were higher (p<0.05) in the SPD rats than in the WKY rats. Sham stress did not affect plasma ghrelin. We conclude that acute psychological stress mobilizes ghrelin and that the SPD rats respond with a higher plasma ghrelin concentration than the WKY rats.     

Development of peptides specifically modulating the activity of KLK2 and KLK3

The prostate produces several proteases, the most abundant ones being kallikrein-related peptidase 3 (KLK3, PSA) and KLK2 (hK2), which are potential targets for tumor imaging and treatment. KLK3 expression is lower in malignant than in normal prostatic epithelium and it is further reduced in poorly differentiated tumors, in which the expression of KLK2 is increased. KLK3 has been shown to inhibit angiogenesis, whereas KLK2 may mediate tumor growth and invasion by participating in proteolytic cascades. Thus, it may be possible to control prostate cancer growth by modulating the proteolytic activity of KLK3 and KLK2. We have developed peptides that very specifically stimulate the activity of KLK3 or inhibit that of KLK2. Using these peptides we have established peptide-based methods for the determination of enzymatically active KLK3. The first-generation peptides are unstable in vivo and are rapidly cleared from the circulation. Currently we are modifying the peptides to make them suitable for in vivo applications. We have been able to considerably improve the stability of KLK2-binding peptides by cyclization. In this review we summarize the possible roles of KLK3 and KLK2 in prostate cancer and then concentrate on the development of peptides that modulate the activity of these proteases.     

Multiple MONOPTEROS-dependent pathways are involved in leaf initiation

Initiation of leaves at the flanks of the shoot apical meristem occurs at sites of auxin accumulation and pronounced expression of auxin-inducible PIN-FORMED1 (PIN) genes, suggesting a feedback loop to progressively focus auxin in concrete spots. Because PIN expression is regulated by auxin response factor activity, including MONOPTEROS (MP), it appeared possible that MP affects leaf formation as a positive regulator of PIN genes and auxin transport. Here, we analyze a novel, completely leafless phenotype arising from simultaneous interference with both auxin signaling and auxin transport. We show that mp pin1 double mutants, as well as mp mutants treated with auxin-efflux inhibitors, display synergistic abnormalities not seen in wild type regardless of how strongly auxin transport was reduced. The synergism of abnormalities indicates that the role of MP in shoot meristem organization is not limited to auxin transport regulation. In the mp mutant background, auxin transport inhibition completely abolishes leaf formation. Instead of forming leaves, the abnormal shoot meristems dramatically increase in size, harboring correspondingly enlarged expression domains of CLAVATA3 and SHOOTMERISTEMLESS, molecular markers for the central stem cell zone and the complete meristem, respectively. The observed synergism under conditions of auxin efflux inhibition was further supported by an unrestricted PIN1 expression in mp meristems, as compared to a partial restriction in wild-type meristems. Auxin transport-inhibited mp meristems also lacked detectable auxin maxima. We conclude that MP promotes the focusing of auxin and leaf initiation in part through pathways not affected by auxin efflux inhibitors.     

Growth and development of seminal and crown root systems in N-limited barley,and their contributions to nitrate acquisition during vegetative and generative growth

Barley (Hordeum vulgare L., cvs Golf and Laevigatum) was grown under nitrogen limitation, controlled by the relative rate of nitrate-N addition (RA), in solution culture. The seminal and crown root systems were kept apart, but in contact with the same nutrient solution throughout culturing. Growth, nitrate uptake, and in vitro nitrate reductase (NR) activity in the different root parts were studied at plant ages from 40 (late vegetative stage) to 110 (mid grain-filling) days. The RA was during this time interval stepwise decreased from 0.08 day^–1 to 0.005 day^–1. The ratio between seminal root dry weight and total plant dry weight decreased drastically during post-anthesis growth, whereas the contribution by crown roots remained unchanged. Tissue nitrogen concentrations in seminal roots did not change with time, but decreased in crown roots after day 80. The NR activity decreased with age in both seminal and crown roots. The V_max for net nitrate uptake decreased throughout the experiment in the seminal root system, but not in the crown root system. The kinetic properties (V_max and K_M) were used to calculate the nitrate concentration required to maintain a relative rate of nitrate-N uptake that equals the relative addition rate. These concentrations (2 to 5 mmol m^–3) were found to closely match actually measured nitrate concentrations in the nutrient solution (1 to 6 mmol m^–3). From uptake kinetics, it was deduced that the contribution by seminal roots to total nitrate uptake at these concentrations decreased from more than 50% in vegetative plants, to about 20% just after main shoot anthesis, and to less than 5% during grain-filling. ei]Section editor: H Lambers     

Uptake and degradation of low density lipoproteins in atherosclerotic rabbit aorta: role of local LDL modification

The uptake of native and modified low density lipoprotein (LDL) in foam cells in atherosclerotic tissue was studied in an in vitro perfusion system for rabbit aorta. Experimental atherosclerosis was induced in rabbits by a combination of cholesterol feeding and mechanical injury. The aorta was perfused in an incubation chamber. A trace-label, radioiodinated tyramine-cellobiose, was used to study cellular uptake of lipoproteins. After perfusion, the tissue was digested and cells were isolated by centrifugation in a density gradient. About 40 times more LDL per cell was accumulated in the foam cell fraction than in the smooth muscle cell fraction. When the cellular uptake of LDL and acetylated LDL (AcLDL) was compared, about 4 times more AcLDL than LDL was taken up by the foam cells, suggesting that the scavenger receptor is expressed in these cells. In a competition experiment, the uptake of LDL into foam cells was reduced by 70% when a tenfold excess of AcLDL was added. This experiment suggests that native LDL is taken up by the same mechanism as AcLDL. The accumulation of radiolabeled LDL in plaques and in foam cells was reduced by 30-55% by adding vitamin E (0.1 mg/ml) to the system. These studies show an uptake of LDL by foam cells in the atherosclerotic tissue. Furthermore, these cells seem to express the scavenger receptor. The competition experiment would suggest that native LDL is taken up by the scavenger receptor. The observation that an antioxidant, vitamin E, may decrease this uptake suggests that oxidative modification of LDL is of importance for this process.