Neurotrophic factor therapy for nervous system degenerative diseases

The ability of neurotrophic factors to regulate developmental neuronal survival and adult nervous system planticity suggests the use of these molecuales to treat neurodegeneration associated with human diseases. Solid rationales exist for the use of NGF and neurotrophin-3 in the treatment of neuropathies of the peripheral sensory system, insulin-like growth factor and ciliary neurotrophic factor in motor neuron atrophy, and NGF in Alzheimer's disease. Growth factors have been identified for neurons affected in Parkinson's disease, Huntington's disease, and acute brain and spinal cord injury. Various strategies are actively pursued to deliver neurotrophic factors to the brain, and develop therapeutically useful molecules that mimic neurotrophic factor actions or stimulate their production or receptor mechanisms. 1994 John Wiley & Sons, Inc.     

Effects of Chronic Basic Fibroblast Growth Factor Administration to Rats with Partial Flmbrial Transections on Presynaptic Cholinergic Parameters and Muscarinic Receptors in the Hippocampus: Comparison with Nerve Growth Factor

Abstract: The present study compares the effects of chronic administration of basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) on various hippocampal cholinergic parameters in rats with partial unilateral fimbrial transections. Lesions resulted in marked reductions of several presynaptic cholinergic parameters: choline acetyltransferase (ChAT) activity (by 50%), [³H]-acetylcholine ([³H]ACh) synthesis (by 59%), basal and ve-ratridine (1 μM)-evoked [³H]ACh release (by 44 and 57%, respectively), and [³H]vesamicol binding site densities (by 35%). In addition, [³H]AF-DX 116/muscarinic M₂ binding site densities were also modestly decreased (by 23%). In contrast, [³H]pirenzepine/muscarinic M₁ and [³H]AF-DX 384/muscarinic M₂/M₄ binding site densities were not altered by the lesions, nor were they affected by any of the treatments. Intracerebroventricular administration of bFGF (10 ng, every other day, for 21 days) partially prevented the lesion-induced deficit in hippocampal ChAT activity, an effect that was not markedly different from that measured in the NGF-treated (1 μg intracerebroventricularly, every other day, for 21 days) rats. In rats treated with a combination of bFGF and NGF, ChAT activity was not different from that in rats treated with the individual factors alone. In contrast, the lesion-induced deficits in the other cholinergic parameters were not attenuated by bFGF treatment, although they were at least partially prevented by NGF administration. To determine whether higher concentrations of bFGF are necessary to affect cholinergic parameters other than hippocampal ChAT activity, rats were treated with 1 μg (every other day, 21 days) of the growth factor. In this group of rats, detrimental effects of bFGF, manifested by an increased death rate (46%), and marked reductions in body weight of the survivors, were observed. In addition, this concentration of bFGF appeared to exacerbate the lesion-induced reduction in [³H]ACh synthesis by hippocampal slices; [³H]ACh synthesis in lesioned hippocampi represented 36 and 52% of that in contralateral unlesioned hippocampi for the bFGF-treated and control groups, respectively. In conclusion, although bFGF administration attenuates the deficit in hippocampal ChAT activity induced by partial fimbrial transections, this does not appear to translate into enhanced functional capacity of the cholinergic terminals. This is clearly in contrast to NGF, which enhances not only hippocampal ChAT activity, but also other parameters indicative of increased function in the cholinergic terminals.     

Integration of Rous-associated virus type O provirus in susceptible chicken cells.

The number of viral genome equivalents per haploid cell genome was determined in normal chicken embryos from three selected chicken lines and in cultured fibroblasts (CEF) from these embryos. The cellular concentration of endogenous proviral DNA is similar in embryos from chickens of lines SPAFAS, 7, 15, 7 x 15, and 100. The concentration of proviral DNA is not affected by in vitro cultivation in CEF from lines that do not spontaneously produce virus, nor in CEF from line 7, which lacks receptors for Rous-associated virus type 0 (RAV-0). There is, however, a restricted increase in the number of integrated proviral genome equivalents in CEF from line 7 x 15, which produces RAV-0 and can support replication of this virus, and in CEF from line 15 experimentally infected with RAV-0.     

In vitro assembly of gap junctions.

Gap junction structures were assembled in vitro from octyl-beta-D-glucopyranoside-solubilized components of lens fiber cell membranes. Individual pore structures (connexons), short double-membrane structures, and other amorphous material were evident in the solubilized mixture. Following the removal of the detergent by dialysis, these connexons associated to form single- and double-layered, two-dimensional hexagonal arrays (unit cell size a = b = 8.5 nm). The formation of larger arrays was dependent on the lipid-to-protein ratio and the presence of Mg2+ ions. Crystallographic analysis of electron micrographs revealed that lens junctional connexons consisted of six subunits surrounding a stain-filled channel. Upon further detergent treatment, in vitro assembled gap junctions were insoluble and formed three-dimensional stacks while other components were solubilized. SDS-PAGE and mass data from scanning transmission electron microscopy strongly suggest that a 38-kDa polypeptide, which is a processed form of the lens specific gap junction protein MP70, is a major component of the arrays. The in vitro assembly of gap junctions opens new avenues for the structural analysis of gap junctions and for the study of the intermolecular interactions of connexons during junctional assembly.     

Adenosine arrests breast cancer cell motility by A3 receptor stimulation

In neutrophils, adenosine triphosphate (ATP) release and autocrine purinergic signaling regulate coordinated cell motility during chemotaxis. Here, we studied whether similar mechanisms regulate the motility of breast cancer cells. While neutrophils and benign human mammary epithelial cells (HMEC) form a single leading edge, MDA-MB-231 breast cancer cells possess multiple leading edges enriched with A3 adenosine receptors. Compared to HMEC, MDA-MB-231 cells overexpress the ectonucleotidases ENPP1 and CD73, which convert extracellular ATP released by the cells to adenosine that stimulates A3 receptors and promotes cell migration with frequent directional changes. However, exogenous adenosine added to breast cancer cells or the A3 receptor agonist IB-MECA dose-dependently arrested cell motility by simultaneous stimulation of multiple leading edges, doubling cell surface areas and significantly reducing migration velocity by up to 75 %. We conclude that MDA-MB-231 cells, HMEC, and neutrophils differ in the purinergic signaling mechanisms that regulate their motility patterns and that the subcellular distribution of A3 adenosine receptors in MDA-MB-231 breast cancer cells contributes to dysfunctional cell motility. These findings imply that purinergic signaling mechanisms may be potential therapeutic targets to interfere with the motility of breast cancer cells in order to reduce the spread of cancer cells and the risk of metastasis.     

The non-peptidyl fungal metabolite L-783,281 activates TRK neurotrophin receptors

Neurotrophin binding to the extracellular surface of the Trk family of tyrosine kinase receptors leads to the activation of multiple signalling cascades, culminating in neuroregenerative effects, including neuronal survival and neurite outgrowth. Since neurotrophins themselves are not ideal drug candidates due to their poor pharmacokinetic behaviour and bioavailability, small molecule neurotrophin mimetics may be beneficial in treating a number of neurodegenerative disorders. The present study demonstrates that L-783,281, a non-peptidyl fungal metabolite, is capable of stimulating TrkA, B and C phosphorylation to various extents in CHO cells stably expressing human Trk receptors. L-783,281 also stimulated Trk phosphorylation in a number of rat and human primary neuronal cultures, whereas the highly similar compound, L-767,827, was without effect. Mechanistic studies utilizing transiently transfected PDGF/TrkA and TrkA/PDGF chimeras, demonstrated that L-783,281 is likely to interact with the intracellular domain of the TrkA receptor. Further investigations suggested that L-783,281 was nevertheless able to instigate receptor dimerization by binding in a non-covalent manner. Although the cytotoxicity of the compound was shown to preclude its effects in neuronal survival and neurite outgrowth assays, it is a prototype for a small molecule neurotrophin mimetic that activates Trk by interacting at a site different from the neurotrophin-binding site.     

Delayed appearance of high altitude retinal hemorrhages

Background

Retinal hemorrhages have been described as a component of high altitude retinopathy (HAR) in association with altitude illness. In this prospective high altitude study, we aimed to gain new insights into the pathophysiology of HAR and explored whether HAR could be a valid early indicator of altitude illness.

Methodology/Principal Findings

28 mountaineers were randomly assigned to two ascent profiles during a research expedition to Mt. Muztagh Ata (7546 m/24,751 ft). Digital fundus photographs were taken prior to expedition at 490 m (1,607 ft), during expedition at 4497 m (14,750 ft = base camp), 5533 m (18,148 ft), 6265 m (20,549 ft), 6865 m (22,517 ft) and 4.5 months thereafter at 490 m. Number, size and time of occurrence of hemorrhages were recorded. Oxygen saturation (SpO₂) and hematocrit were also assessed. 79% of all climbers exhibited retinal hemorrhages during the expedition. Number and area of retinal bleeding increased moderately to medium altitudes (6265 m). Most retinal hemorrhages were detected after return to base camp from a high altitude. No post-expeditional ophthalmic sequelae were detected. Significant negative (SpO₂ Beta: −0.4, p<0.001) and positive (hematocrit Beta: 0.2, p = 0.002, time at altitude Beta: 0.33, p = 0.003) correlations with hemorrhages were found.

Conclusions/Significance

When closely examined, a very large amount of climbers exhibit retinal hemorrhages during exposure to high altitudes. The incidence of retinal hemorrhages may be greater than previously appreciated as a definite time lag was observed between highest altitude reached and development of retinal bleeding. Retinal hemorrhages should not be considered warning signs of impending severe altitude illness due to their delayed appearance.     

Employment of a noninvasive magnetic method for evaluation of gastrointestinal transit in rats

ABSTRACT: AC Biosusceptometry (ACB) was previously employed towards recording gastrointestinal motility. Our data show a reliable and successful evaluation of gastrointestinal transit of liquid and solid meals in rats, considering the methods scarcity and number of experiments needed to endorsement of drugs and medicinal plants. ACB permits real time and simultaneous experiments using the same animal, preserving the physiological conditions employing both meals with simplicity and accuracy.     

5′ Nucleotide Sequence of Sindbis Viral RNA

The 5′ sequence of Sindbis viral RNA is m ⁷G^5′ pppApUpGp...