Neonatal fluoxetine exposure alters motor performances of adolescent rats

Growing evidence from human and animal studies has shown adverse consequences of maternal usage of antidepressants in their newborn babies. To study the effects of early antidepressant exposure on motor function later in life, we treated neonatal rat pups with fluoxetine (FLX), a selective serotonin reuptake inhibitor (SSRI)-type antidepressant, from the day of birth to postnatal day 4 and examined motor performance during adolescence. FLX-treated rats had reduced locomotor activities in an open field and poorer motor performance on an accelerating rotarod compared to the control group of saline-treated animals. Nevertheless, the poorer motor performance largely improved after repetitive practices. To elucidate the structural alterations in the motor system, we examined the structure of neurons in motor-related brain regions. The shape, density, and soma size of cerebellar Purkinje cells were comparable in the two groups, however, density of dendritic spine in medial spiny neurons of striatum and Layer 5 pyramidal neurons in the primary motor cortex (M1) were reduced in FLX-rats. Furthermore, the basilar dendrites in M1 Layer 5 neurons had reduced dendritic complexity than those of the control animals. The impaired dendritic structure in striatal and cortical neurons in FLX-treated rats might account for their poorer motor performances. Together, the structure and function of the motor system are affected by early FLX exposure, the long-term effects of early exposure to SSRI-type antidepressants should be concerned.     

Screening for enzyme activity in turbid suspensions with scattered light

New screening techniques for improved enzyme variants in turbid media are urgently required in many industries such as the detergent and food industry. Here, a new method is presented to measure enzyme activity in different types of substrate suspensions. This method allows a semiquantitative determination of protease activity using native protein substrates. Unlike conventional techniques for measurement of enzyme activity, the BioLector technology enables online monitoring of scattered light intensity and fluorescence signals during the continuous shaking of samples in microtiter plates. The BioLector technique is hereby used to monitor the hydrolysis of an insoluble protein substrate by measuring the decrease of scattered light. The kinetic parameters for the enzyme reaction (V(max,app) and K(m,app)) are determined from the scattered light curves. Moreover, the influence of pH on the protease activity is investigated. The optimal pH value for protease activity was determined to be between pH 8 to 11 and the activities of five subtilisin serine proteases with variations in the amino acid sequence were compared. The presented method enables proteases from genetically modified strains to be easily characterized and compared. Moreover, this method can be applied to other enzyme systems that catalyze various reactions such as cellulose decomposition.     

Irreversible Electroporation of Malignant Hepatic Tumors - Alterations in Venous Structures at Subacute Follow-Up and Evolution at Mid-Term Follow-Up

Purpose

To evaluate risk factors associated with alterations in venous structures adjacent to an ablation zone after percutaneous irreversible electroporation (IRE) of hepatic malignancies at subacute follow-up (1 to 3 days after IRE) and to describe evolution of these alterations at mid-term follow-up.

Materials and Methods

43 patients (men/women, 32/11; mean age, 60.3 years) were identified in whom venous structures were located within a perimeter of 1.0 cm of the ablation zone at subacute follow-up after IRE of 84 hepatic lesions (primary/secondary hepatic tumors, 31/53). These vessels were retrospectively evaluated by means of pre-interventional and post-interventional contrast-enhanced magnetic resonance imaging or computed tomography or both. Any vascular changes in flow, patency, and diameter were documented. Correlations between vascular change (yes/no) and characteristics of patients, lesions, and ablation procedures were assessed by generalized linear models.

Results

191 venous structures were located within a perimeter of 1.0 cm of the ablation zone: 55 (29%) were encased by the ablation zone, 78 (41%) abutted the ablation zone, and 58 (30%) were located between 0.1 and 1.0 cm from the border of the ablation zone. At subacute follow-up, vascular changes were found in 19 of the 191 vessels (9.9%), with partial portal vein thrombosis in 2, complete portal vein thrombosis in 3, and lumen narrowing in 14 of 19. At follow-up of patients with subacute vessel alterations (mean, 5.7 months; range, 0 to 14 months) thrombosis had resolved in 2 of 5 cases; vessel narrowing had completely resolved in 8 of 14 cases, and partly resolved in 1 of 14 cases. The encasement of a vessel by ablation zone (OR = 6.36, p<0.001), ablation zone being adjacent to a portal vein (OR = 8.94, p<0.001), and the usage of more than 3 IRE probes (OR = 3.60, p = 0.035) were independently associated with post-IRE vessel alterations.

Conclusion

Venous structures located in close proximity to an IRE ablation zone remain largely unaffected by this procedure, and thrombosis is rare.     

Critical role of RelB serine 368 for dimerization and p100 stabilization

In mature B cells RelB-containing complexes are constitutively present in the nucleus, and they are less susceptible to inhibitory kappaB proteins. In most other cell types inhibitory kappaB proteins prevent nuclear translocation and activation of NFkappaB. We reasoned that this characteristic might be because of post-translational modifications of RelB. In Drosophila, signal-dependent phosphorylation of the Rel homologue Dorsal at serine 317 has been shown to be critical for nuclear import. The evolutionary conservation of this serine prompted us to analyze the function of the corresponding site in RelB. As a model system we used the murine S107 plasmacytoma cell line, which lacks endogenous RelB expression. Analysis of S107 cells expressing wild type RelB and serine 368 mutants reveals that serine 368 is not required for nuclear import but that it is critical for RelB dimerization with other members of the NFkappaB family. Similar effects were obtained when the conserved serine in RelA was mutated. We further demonstrate that expression of functional RelB, but not of serine 368 mutants, severely reduces p52 generation and strongly increases expression of the p52 precursor, p100. Wild type RelB, but not mutant RelB, prolonged p100 half-life. We therefore suggest an inhibitory effect of RelB on p100 processing, which is possibly regulated in a signal-dependent manner.     

Inbreeding,immune defence and ectoparasite load in different mockingbird populations and species in the Galápagos Islands

Inbreeding may impair an individual's immune system, render it more susceptible to disease and hence contribute to the extinction risk of small and isolated populations, as often found on islands. So far, surprisingly few studies have assessed the effects of inbreeding on immunocompetence in wild populations. Using 26 microsatellite loci and genetic data from museum specimens and contemporary samples, we calculated short‐term and long‐term inbreeding in 13 different mockingbird populations covering the range of all 4 species in the Galápagos Islands and compared them with three different measures of innate immunity and ectoparasite load. We found no significant effect of either measure of inbreeding on natural antibody or complement enzyme titres, heterophil‐lymphocyte ratio or feather louse abundance. Hence, our results do not support a link between inbreeding and immunocompetence. However, overall statistical power and repeatabilities of antibody and complement enzyme titres were low. Nevertheless, generally, natural antibody titres were high suggesting that the mockingbirds may be equipped with a strong first line of defence, as found in other island species.     

Effect of sweet cherry genes PaLACS2 and PaATT1 on cuticle deposition,composition and permeability in Arabidopsis

The cuticular membrane (CM) of sweet cherry (Prunus avium L.) fruit is severely strained during development. Strain results from a cessation of CM deposition during early development and is possibly caused by a downregulation of genes involved in CM synthesis. The objectives of our study were to investigate the effects of ectopic expression of two sweet cherry genes, PaLACS2 (a putative long-chain acyl-CoA synthetase) and PaATT1 (a putative cytochrome P450 monooxygenase), in Arabidopsis thaliana (L.). Effects on the expression of endogenous LACS2, ATT1 and LACS1 genes, wax and cutin composition, and cuticle permeability were investigated in 13 transgenic lines. Of these, six lines are selected for presentation based on the magnitude of the response. The amount of cutin increased in the PaLACS2 overexpression line C-L-29 and in the complemented lacs2-1 knockout mutant line l-L-14, but overexpression had no effect on cutin composition or wax. Wax deposition decreased in the complemented knockout lines l-L-14 and l-L-21. Overexpressing PaATT1 in A. thaliana line C-A-6 had no significant effect on cutin and wax deposition. In the complemented knockout lines a-A-7 and a-A-12, cutin deposition increased, whereas wax deposition was unaffected. The permeability of the cuticle for water and toluidine blue decreased in the PaLACS2 and PaATT1 complemented knockout lines. The results suggest that (1) PaLACS2 and PaATT1 expressed in A. thaliana are involved in cutin biosynthesis, and (2) their functions are consistent with those of a typical long-chain acyl-CoA synthetase (PaLACS2) and of a cytochrome P450 monooxygenase (PaATT1).     

Cooperative binding of single-stranded telomeric DNA by the Pot1 protein of Schizosaccharomyces pombe

The fission yeast Pot1 (protection of telomeres) protein is a single-stranded telomeric DNA-binding protein and is required to protect the ends of chromosomes. Its N-terminal DNA-binding domain, Pot1pN, shows sequence similarity to the first OB fold of the telomere-binding protein alpha subunit of Oxytricha nova. The minimal-length telomeric ssDNA required to bind Pot1pN was determined to consist of six nucleotides, GGTTAC, by gel filtration chromatography and filter-binding assay (K(D) = 83 nM). Pot1pN is a monomer, and each monomer binds one hexanucleotide. Experiments with nucleotide substitutions demonstrated that the central four nucleotides are crucial for binding. The dependence of Pot1pN-ssDNA binding on salt concentration was consistent with a single ionic contact between the protein and the ssDNA phosphate backbone, such that at physiological salt condition 83% of the free energy of binding is nonelectrostatic. Subsequent binding experiments with longer ssDNAs indicated that Pot1pN binds to telomeric ssDNA with 3' end preference and in a highly cooperative manner that mainly results from DNA-induced protein-protein interactions. Together, the binding properties of Pot1pN suggest that the protein anchors itself at the very 3' end of a chromosome and then fills in very efficiently, coating the entire single-stranded overhang of the telomere.