Production and characterization of a monoclonal antibody able to discriminate galectin-1 from galectin-2 and galectin-3

Antisera raised against galectin-1 exhibit crossreactivities with other galectins or related molecules. In order to overcome this problem, a monoclonal antibody to human brain galectin-1 was obtained by selecting clones without reactivity toward galectin-3. This mAb specifically bound galectin-1 of various animal origins but neither galectin-2 nor galectin-3. Western-blotting analysis of soluble human brain extracts after 2D gel electrophoresis revealed only the two most acidic isoforms of galectin-1. The ability of this mAb to bind galectin-1/asialofetuin complexes indicates that its epitope is not localized in the carbohydrate recognition domain of galectin-1. This particularity induces with efficiency its monospecificity.      

An anchored restriction-mapping approach applied to the genetic analysis of the Anopheles gambiae malaria vector complex 1

We introduce here a simple approach for rapidly determining restriction maps for a number of regions of a genome; this involves "anchoring" a map with a rare restriction site (in this case the seldom-cutting EagI) followed by partial digestion of a frequent-cutting enzyme (e.g., Sau 3A). We applied this technology to five species of the Anopheles gambiae complex. In a single Southern blot we obtained about a 15-kb restriction map each for the mtDNA, rRNA gene, and a scnDNA region for each of five species. Phylogenetic analyses of these regions yield trees at odds with the more traditional chromosome inversion-based trees. The value of the approach for systematic purposes is the ease with which several large, independent regions of the genome can be quickly assayed for molecular variation.      

Synthesis and possible applications of biotin-linked copper clusters

The trifunctional aziridine XAMA-7 (CAS 57116-45-7) has been used to form crosslinks between a deep red-violet copper cluster of the type Cu(I)8Cu(II)6pen12Cl5- (pen=penicillamine) and molecules with biological activity such as d-biotin and proteins. A complex containing biotin, bovine serum albumin and the copper cluster displayed activity toward affinity columns of avidin on Agarose, and the red-violet pigment was immobilized on the gel. This interaction was completely blocked in gels which had been pretreated with d-biotin carboxylic acid. The free and biologically active versions of the cluster have some potential for biomedical applications. For example, the short-lived positron emitter 64Cu (suitable for positron tomography) may be carried in the cluster's structure. The cluster is paramagnetic, but it is a relatively weak effector of water proton spin-lattice relaxation. Other members of this structural group of inorganic compounds may have better magnetic properties, and the crosslinking reaction with aziridines appears to be generally applicable to the group.     

Analysis of Plasma Biopterin Levels in Psychiatric Disorders Suggests a Common BH<Subscript>4</Subscript> Deficit in Schizophrenia and Schizoaffective Disorder

Tetrahydrobiopterin (BH₄) is an essential cofactor for amine neurotransmitter synthesis. BH₄ also stimulates and modulates the glutamatergic system, and regulates the synthesis of nitric oxide by nitric oxide synthases. A connection between BH₄ deficiencies and psychiatric disorders has been previously reported; major depression and obsessive-compulsive disorder have been found in subjects with a BH₄ deficiency disorder and more recently we have observed a robust plasma deficit of biopterin (a measure of BH₄), in a large group of schizophrenic patients compared to control subjects. To extend our previous finding in schizophrenia, we analyzed plasma biopterin levels from patients with schizoaffective and bipolar disorders. A significant difference in biopterin was seen among the diagnostic groups (P < 0.0001). Post hoc analyses indicated significant biopterin deficits relative to the normal control group for the schizoaffective group, who had biopterin levels comparable to the schizophrenic group. Bipolar disorder subjects had plasma biopterin levels that were higher that the schizoaffective disorder group and significantly higher than the schizophrenic group. The demonstrated significant biopterin deficit in both schizophrenia and schizoaffective disorder, may suggest an etiological role of a BH₄ deficit in these two disorders, via dysregulation of neurotransmitter systems.     

AMP-activated protein kinase mediates preconditioning in cardiomyocytes by regulating activity and trafficking of sarcolemmal ATP-sensitive K(+) channels

Brief periods of ischemia and reperfusion that precede sustained ischemia lead to a reduction in myocardial infarct size. This phenomenon, known as ischemic preconditioning, is mediated by signaling pathway(s) that is complex and yet to be fully defined. AMP-activated kinase (AMPK) is activated in cells under conditions associated with ATP depletion and increased AMP/ATP ratio. In the present study, we have taken advantage of a cardiac phenotype overexpressing a dominant negative form of the alpha2 subunit of AMPK to analyze the role, if any, that AMPK plays in preconditioning the heart. We have found that myocardial preconditioning activates AMPK in wild type, but not transgenic mice. Cardiac cells from transgenic mice could not be preconditioned, as opposed to cells from the wild type. The cytoprotective effect of AMPK was not related to the effect that preconditioning has on mitochondrial membrane potential as revealed by JC-1, a mitochondrial membrane potential-sensitive dye, and laser confocal microscopy. In contrast, experiments with di-8-ANEPPS, a sarcolemmal-potential sensitive dye, has demonstrated that intact AMPK activity is required for preconditioning-induced shortening of the action membrane potential. The preconditioning-induced activation of sarcolemmal K(ATP) channels was observed in wild type, but not in transgenic mice. HMR 1098, a selective inhibitor of sarcolemmal K(ATP) channels opening, inhibited preconditioning-induced shortening of action membrane potential as well as cardioprotection afforded by AMPK. Immunoprecipitation followed by Western blotting has shown that AMPK is essential for preconditioning-induced recruitment of sarcolemmal K(ATP) channels. Based on the obtained results, we conclude that AMPK mediates preconditioning in cardiac cells by regulating the activity and recruitment of sarcolemmal K(ATP) channels without being a part of signaling pathway that regulates mitochondrial membrane potential.     

Mycorrhizal improvement in non-leguminous nitrogen fixing associations with particular reference toHippophaë rhamnoides L.

Summary The roots ofHippophaë rhamnoides which regularly bear actinomycete induced nodules when growing on Scottish sand dunes have also been found to support an endomycorrhizal association withGlomus fasciculatus. Ultrastructural and cytochemical studies carried out on the indigenous infections of establishedHippophaë mycorrhizal roots would support the postulate that transport is indeed occurring between the fungal symbiont and the host plant and vice versa in respect of phosphate and carbohydrate. Experiments using various inoculation regimes, demonstrated the significant improvement in the mycorrhizal/nodulated plants compared to the nodulated-only and the mycorrhizal-only plants with respect to plant growth, uptake of phosphate and nitrogenase activity, when grown in a medium poor in combined nitrogen and soluble phosphate. Preliminary work onAlnus andMyrica species growing in Central Scotland indicates that the mycorrhizae associated with these nodulated root systems exhibit a different interaction pattern which may be dependent on habitat type and associated angiosperm species.