Neuropathological findings processed by artificial neural networks (ANNs) can perfectly distinguish Alzheimer's patients from controls in the Nun Study

Background  

Many reports have described that there are fewer differences in AD brain neuropathologic lesions between AD patients and control subjects aged 80 years and older, as compared with the considerable differences between younger persons with AD and controls. In fact some investigators have suggested that since neurofibrillary tangles (NFT) can be identified in the brains of non-demented elderly subjects they should be considered as a consequence of the aging process. At present, there are no universally accepted neuropathological criteria which can mathematically differentiate AD from healthy brain in the oldest old.     

Nucleotide Phosphohydrolase in Purified Vaccinia Virus

Purified infectious vaccinia virus has been shown to contain an enzyme or enzymes that remove the terminal phosphate group from adenosine triphosphate (ATP), guanosine triphosphate (GTP), uridine triphosphate (UTP), and cytidine triphosphate (CTP). The K(m) for ATP of this enzyme is 5.5 x 10(-4)m, and the relative rates of the reaction with ATP, GTP, UTP, and CTP are 1.00, 0.34, 0.15, and 0.29, respectively. The virus enzyme does not react with any of the diphosphates. The rate of the reaction is proportional to the amount of virus added and is linear for 130 min. The virus nucleotide phosphohydrolase activity is greatly stimulated by Mg(++) and very slightly stimulated by Ca(++). The small residual activity observed in the absence of divalent cations is completely inhibited by ethylenediaminetetraacetic acid. Neither Na(+) nor K(+) ions, nor any mixture of these, was found to stimulate the reaction significantly, and ouabain, at 10(-4)m, inhibited the reaction by only 27%. The response of the vaccinia enzyme to mono- and divalent cations and to ouabain indicates that the vaccinia enzyme has different properties from those associated with microsomes and mitochondria.     

Delivery of Brain-Derived Neurotrophic Factor by 3D Biocompatible Polymeric Scaffolds for Neural Tissue Engineering and Neuronal Regeneration

Biopolymers are increasingly employed for neuroscience applications as scaffolds to drive and promote neural regrowth, thanks to their ability to mediate the upload and subsequent release of active molecules and drugs. Synthetic degradable polymers are characterized by different responses ranging from tunable distension or shrinkage to total dissolution, depending on the function they are designed for. In this paper we present a biocompatible microfabricated poly-ε-caprolactone (PCL) scaffold for primary neuron growth and maturation that has been optimized for the in vitro controlled release of brain-derived neurotrophic factor (BDNF). We demonstrate that the designed morphology confers to these devices an enhanced drug delivery capability with respect to monolithic unstructured supports. After incubation with BDNF, micropillared PCL devices progressively release the neurotrophin over 21 days in vitro. Moreover, the bioactivity of released BDNF is confirmed using primary neuronal cultures, where it mediates a consistent activation of BDNF signaling cascades, increased synaptic density, and neuronal survival. These results provide the proof-of-principle on the fabrication process of micropatterned PCL devices, which represent a promising therapeutic option to enhance neuronal regeneration after lesion and for neural tissue engineering and prosthetics.     

Dynamic of VE-cadherin-mediated spermatid–Sertoli cell contacts in the mouse seminiferous epithelium

Spermatids are haploid differentiating cells that, in the meantime they differentiate, translocate along the seminiferous epithelium towards the tubule lumen to be just released as spermatozoa. The success of such a migration depends on dynamic of spermatid–Sertoli cell contacts, the molecular nature of which has not been well defined yet. It was demonstrated that the vascular endothelial cadherin (VEC) is expressed transitorily in the mouse seminiferous epithelium. Here, we evaluated the pattern of VEC expression by immunohistochemistry first in seminiferous tubules at different stages of the epithelial cycle when only unique types of germ cell associations are present. Changes in the pattern of VEC localization according to the step of spermatid differentiation were analysed in detail using testis fragments and spontaneously released germ cells. Utilizing the first wave of spermatogenesis as an in vivo model to have at disposal spermatids at progressive steps of differentiation, we checked for level of looser VEC association with the membrane by performing protein solubilisation under mild detergent conditions and assays through VEC-immunoblotting. Being changes in VEC solubilisation paralleled in changes in phosphotyrosine (pY) content, we evaluated if spermatid VEC undergoes Y658 phosphorylation and if this correlates with VEC solubilisation and spermatid progression in differentiation. Altogether, our study shows a temporally restricted pattern of VEC expression that culminates with the presence of round spermatids to progressively decrease starting from spermatid elongation. Conversely, pY658-VEC signs elongating spermatids; its intracellular polarized compartmentalization suggests a possible involvement of pY658-VEC in the acquisition of spermatid cell polarity.     

Contrasting intraspecific foliar trait responses to stressful conditions of two rhizomatous granite outcrop species at different scales in southwestern Australia

Plants respond to changing environmental conditions, and their ability to adjust intra‐specifically to such shifts represents an ecological and evolutionary advantage. We studied seven plant traits for two common, rhizomatous granite outcrop species (the fern Cheilanthes austrotenuifolia, and the herb Stypandra glauca) with seasonal foliage during the cooler, wetter winter months at seven sites across an aridity gradient in southwestern Australia. We investigated trait patterns at regional and habitat scale, by investigating changes in trait values along the aridity gradient, and by comparing two different habitats types (sun‐exposed and sheltered). We expected plants occurring in more arid sites and highly irradiated, shallow‐soil (sun‐exposed) habitats, to exhibit traits indicative of more conservative resource acquisition, retention and use strategies. At the habitat scale, we found support for our prediction, with plants in more stressful, sun‐exposed habitats showing traits’ values associated with more conservative strategies (especially for water), such as smaller plants, denser leaves, higher foliar δ¹³C and C/N. However, at the regional scale many traits displayed the opposite pattern, suggesting less conservative resource acquisition in more arid sites. This evidence was particularly pronounced for specific leaf area (SLA), which exhibited a significant, positive relationship with increasing aridity. We suggest that the unexpected regional trends in foliar traits relate to shorter lived, faster growing leaves linked to highly efficient resource acquisition and use strategies during the shorter growing season in the more arid regions. These highly exploitative strategies may enable plants to avoid climate extremes, that is, hot and dry periods in the more arid sites. Our findings of contrasting foliar traits responses at different scales support the importance of multi‐scale approaches to quantify the role of intraspecific trait variability.     

Exploring interaction of beta-amyloid segment (25-35) with membrane models through paramagnetic probes.

The accumulation of beta-amyloid peptides into senile plaques is one of the hallmarks of Alzheimer's disease (AD). There is mounting evidence that the lipid matrix of neuronal cell membranes plays an important role in the beta-sheet oligomerization process of beta-amyloid. Abeta(25-35), the sequence of which is GSNKGAIIGLM, is a highly toxic segment of amyloid beta (Abeta)-peptides, which forms fibrillary aggregates. In the present work, two spin-labelled Abeta(25-35) analogues containing the nitroxide group of the amino acid TOAC (2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid) as a paramagnetic probe at the N- or the C-terminus of the peptide sequence, respectively, were synthesized in order to investigate the peptide-membrane interaction. The orientation and associated changes of the peptide conformation in the presence of different artificial membrane models (micelles, liposomes) were evaluated by electron paramagnetic resonance and circular dichroism techniques. The results of this study allowed us to propose a model in which the C-terminal portion of the peptide is highly associated to the membrane, while the N-terminal part extends into the aqueous phase with occasional contacts with the lipid head-group region. Interestingly, the interaction of the C-terminal portion of the peptide is particularly enhanced in the presence of sodium dodecyl sulfate (SDS) molecules.     

Rhenium(I) and ruthenium(II) complexes with a crown-linked methanofullerene ligand: synthesis, electrochemistry and photophysical characterization.

A cyclopropanation reaction has been used to prepare two methanofullerenes bearing a 2,2'-bipyridine () or pyridine () ligand separated from the fullerene through an oxyethylene macrocyclic spacer. Derivatives and were, in turn, employed to synthesize two fullerene-based ruthenium(ii) and rhenium(i) donor-acceptor dyads whose molecular structure was confirmed by (1)H NMR, (13)C NMR and exact mass determination. The UV-Vis spectrum of the dyads is the superimposition of those of appropriate model systems, indicating that ground-state electronic interactions between the constituent chromophores, in solution, are negligible, in line also with the electrochemical results. The complex voltammetric pattern was characterized by the superimposition of signals attributed to one moiety or another without significant shifts with respect to their models. Furthermore, both species undergo partial chemical degradation in the time scale of cyclic voltammetry upon their multiple reduction. Photophysical properties of and , namely, excited state interactions between the ruthenium(ii) or rhenium(i) complexes and [60]fullerene have been investigated by steady-state and time-resolved UV-Vis-NIR luminescence spectroscopy that was complemented by nanosecond laser flash photolysis in CH(2)Cl(2) solutions. All experimental findings were set into relation with the corresponding reference compounds. More precisely, excitation of the metal complexes in and gives rise to a notable steady-state and time-resolved luminescence quenching of both metal to ligand charge transfer states (i.e., [Ru(bpy)(3)](2+) and [(bpy)Re(CO)(3)(py)](+)). Conclusive evidence about the nature of the photoproducts came from nanosecond laser flash photolysis. In these experiments, only the long-lived and oxygen-sensitive [60]fullerene triplets were detected. Two pathways are envisioned for this [60]fullerene triplet formation. Firstly, intramolecular transduction of the triplet excited state energy evolving from the photoexcited metal complexes. Secondly, intersystem crossing of directly excited [60]fullerene.     

Evolution of Hypervariable Region 1 of Hepatitis C Virus in Primary Infection

The hypervariable region 1 (HVR-1) of the putative envelope encoding E2 region of hepatitis C virus (HCV) RNA was analyzed in sequential samples from three patients with acute type C hepatitis infected from different sources to address (i) the dynamics of intrahost HCV variability during the primary infection and (ii) the role of host selective pressure in driving viral genetic evolution. HVR-1 sequences from 20 clones per each point in time were analyzed after amplification, cloning, and purification of plasmid DNA from single colonies of transformed cells. The intrasample evolutionary analysis (nonsynonymous mutations per nonsynonymous site [K_a], synonymous mutations per synonymous site [K_s], K_a/K_s ratio, and genetic distances [gd]) documented low gd in early samples (ranging from 2.11 to 7.79%) and a further decrease after seroconversion (from 0 to 4.80%), suggesting that primary HCV infection is an oligoclonal event, and found different levels and dynamics of host pressure in the three cases. The intersample analysis (pairwise comparisons of intrapatient sequences; rK_a, rK_s, rK_a/rK_s ratio, and gd) confirmed the individual features of HCV genetic evolution in the three subjects and pointed to the relative contribution of either neutral evolution or selective forces in driving viral variability, documenting that adaptation of HCV for persistence in vivo follows different routes, probably representing the molecular counterpart of the viral fitness for individual environments.     

Colon cancer cell vaccine prepared with replication-deficient vaccinia viruses encoding B7.1 and interleukin-2 induce antitumor response in syngeneic mice

 A replication-deficient recombinant vaccinia virus, NYVAC, was developed by deleting 18 open reading frames in the vaccinia virus genome. Recombinant NYVAC, encoding the murine T cell co-stimulatory gene B7.1 (CD 80) (NYVAC-B7.1) and the murine interleukin-2 gene (NYVAC-IL-2), were prepared and the expression of B7.1 and the secretion of IL-2 were respectively confirmed in vitro. The use of these viruses to prepare a potent tumor cell vaccine was studied in a syngeneic murine CC-36 colon adenocarcinoma model. Mice were immunized on days 1 and 8 with 10⁶ irradiated CC-36 cells that were infected with 10⁷ plaque-forming units of either NYVAC-B7.1, NYVAC-IL-2 or a control virus, NYVAC-HR, which encodes a vaccinia virus host-range gene. These mice were then challenged with 10⁸ viable CC-36 tumor cells on day 15. All mice (10/10) in a group that had received no vaccination and all mice (20/20) in a group that had received a control vaccine of CC-36/NYVAC-HR developed tumor 4-weeks after tumor cell challenge. Interestingly, only 16/20 mice in a group that had received CC-36/NYVAC-B7.1 showed the development of tumor after the same interval. The protection against tumor development and the reduction in tumor burden (as mean tumor diameter, 4 weeks after tumor challenge) were significant in this group when compared to groups that were either unvaccinated or vaccinated with CC-36/NYVAC-HR (mean tumor diameter = 6.51±3.2 mm compared to 26.5±0.9 mm or 26.2±1.8 mm respectively) (P = < 0.05). The protection against tumor in a group of mice that received CC-36/NYVAC-IL-2 vaccination was similar to that in the unvaccinated group or the group receiving a CC-36/NYVAC-HR control vaccination. However, in a survival experiment, mice that received either CC36/NYVAC-B7.1 or CC-36/NYVAC-IL-2 vaccination on the day of tumor transplantation survived significantly longer than mice that had not been vaccinated (median survival 60+ days, 60+ days or 23.5 days respectively) (P = <0.05). Interestingly, when a therapeutic tumor vaccination was performed on day 4 after tumor transplantation, mice that had been vaccinated with either CC36/NYVAC-B7.1 or CC-36/NYVAC-IL-2 did not show an improved survival when compared to mice in the control that had not been vaccinated (median survival 28 days compared to 26 days or 25 days respectively). However, mice that had received a therapeutic vaccination with CC-36 cells infected with both NYVAC-B7.1 and NYVAC-IL-2, 4 days after tumor transplantation, survived significantly longer than control mice that had not received any vaccination (median survival 29.5 days compared to 25 days respectively) (P<0.05). These results suggest that a replication-deficient recombinant NYVAC encoding the B7.1 gene and NYVAC encoding the IL-2 gene can be used to produce an effective vaccinia-virus-augmented tumor cell vaccine. Received: 2 March 1998 / Accepted 23 March 1998