Optimizing DHA levels in piglets by lowering the linoleic acid to alpha-linolenic acid ratio

We examined the effect of altering the linoleic acid (LA, 18:2n-6) to alpha-linolenic acid (ALA, 18:3n-3) ratio in the dietary fats of 3 day old piglets fed formula for 3 weeks. The LA-ALA ratios of the experimental formulas were 0.5:1, 1:1, 2:1, 4:1, and 10:1. The level of LA was held constant at 13% of total fats while the level of ALA varied from 1.3% (10:1 group) to 26.8% (0.5:1 group). Incorporation of the n-3 long chain PUFA EPA and 22:5n-3 into erythrocytes, plasma, liver, and brain tissues was linearly related to dietary ALA. Conversely, incorporation of DHA into all tissues was related to dietary ALA in a curvilinear manner, with the maximum incorporation of DHA appearing to be between the LA-ALA ratios of 4:1 and 2:1. Feeding LA-ALA ratios of 10:1 and 0.5:1 resulted in lower and similar proportions of DHA in tissues despite the very different levels of dietary ALA (1.3 vs. 26.8% of total fats, respectively). These results are relevant to term infant studies in that they confirm our earlier findings of the positive effect on DHA status by lowering the LA-ALA ratio from 10:1 to 3:1 or 4:1, and they predict that ratios of LA-ALA below 4:1 would have little further beneficial effect on DHA status.     

20 S proteasomes are imported as precursor complexes into the nucleus of yeast

The mechanism by which yeast 20 S proteasomes are imported into the nucleus is still unresolved. Here, we provide the first evidence that 20 S proteasomes are imported as precursor complexes into the nucleus. By using the srp1-49 mutant which is deficient in nuclear import of cargos with classical nuclear localization sequences (cNLS), we show that proteasome precursor complexes associate with importin/karyopherin alphabeta, the cNLS receptor, and that they accumulate inside the cytoplasm. Reconstitution assays revealed that only precursor complexes are targeted to the nuclear envelope (NE) by karyopherin alphabeta. In support, the green fluorescent protein (GFP)-labelled maturation factor Ump1, marking precursor complexes, mainly localizes to the nucleus and around the NE. Our data suggest that nuclear 20 S proteasomes are finally matured inside the nucleus.     

Activation of group II mGlu receptors inhibits voltage-gated Ca2+ currents in myenteric neurons

The enteric nervous system (ENS) contains functional ionotropic and group I metabotropic glutamate (mGlu) receptors. In this study, we determined whether enteric neurons express group II mGlu receptors and the effects of mGlu receptor activation on voltage-gated Ca(2+) currents in these cells. (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate (2R,4R-APDC), a group II mGlu receptor agonist, reversibly suppressed the Ba(2+) current in myenteric neurons isolated from the guinea pig ileum. Significant inhibition was also produced by L-glutamate and the group II mGlu receptor agonists, (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG-IV) and (2S,1'S,2'S)-2-(2-carboxycyclopropyl)glycine (L-CCG-I), with a rank order potency of 2R,4R-APDC > DCG-IV > L-glutamate > L-CCG-I, and was reduced by the group II mGlu receptor antagonist LY-341495. Pretreatment of neurons with pertussis toxin (PTX) reduced the action of mGlu receptor agonists, suggesting participation of G(i)/G(o) proteins. Finally, omega-conotoxin GVIA blocked current suppression by DCG-IV, suggesting modulation of N-type calcium channels. mGlu2/3 receptor immunoreactivity was displayed by neurons in culture and in the submucosal and myenteric plexus of the ileum. A subset of these cells displayed a glutamatergic phenotype as shown by the expression of vesicular glutamate transporter 2. These results provide the first evidence for functional group II mGlu receptors in the ENS and show that these receptors are PTX sensitive and negatively coupled to N-type calcium channels. Inhibition of N-type calcium channels produced by activation of group II mGlu receptors may modulate enteric neurotransmission.     

Organization of the <Emphasis Type="Italic">MASP2</Emphasis> locus and its expression profile in mouse and rat

The identification of vertebral fracture in osteoporosis is based mainly on the identification of abnormal variation in vertebral shape, but this can be misleading in the presence of a non-fracture deformity or normal variant of vertebral shape. Qualitative identification of vertebral fracture (Qual) is influenced by the subjectivity of the approach, and although more objective, the semiquantitative method (SQ) can be difficult to apply. In addition, there has been little independent evaluation of SQ in relation to other approaches. We aimed to evaluate a new algorithm-based approach for the qualitative identification of vertebral fracture (ABQ) and to compare it with SQ and Qual. Two radiologists reported spinal radiographs for 372 postmenopausal women using Qual (reader 1), and SQ and ABQ (reader 2). Non-fracture deformities and normal variants were also reported using Qual and ABQ. The prevalence of vertebral fracture by subjects was higher for SQ (24%) than for Qual (11%) and ABQ (7%). Agreement was poor between SQ and the other methods, and moderate between Qual and ABQ. Twenty-two women with vertebral fracture were agreed by all three methods, similar to the total identified by ABQ (25 women). Seventeen women diagnosed with fracture by Qual, had non-fracture deformity or normal variant (but no fracture) according to ABQ. Of the women with SQ fractures, 53% and 70% were identified negative for fracture but positive for non-fracture deformity or normal variant by ABQ and Qual. The main sources of discrepancy between SQ and the other methods were Scheuermanns disease, normal variation, and degenerative change accompanied by short anterior vertebral height. For all methods, bone mineral density (BMD) and BMD Z-scores were lower in women with vertebral fractures than in those with no fractures. Bone mineral density and BMD Z-scores were also lower at the lumbar spine and total body in women with vertebral fractures according to Qual and ABQ than they were for SQ, and were lower in women with SQ fractures agreed by Qual and ABQ, compared with those diagnosed negative for fracture by Qual and ABQ (p<0.01). We conclude that poor agreement between methods arises mainly from difficulties in differentiating true fracture from non-fracture deformity. Our new approach attempts to address this problem but requires further testing in a larger study population.     

Blm3 is part of nascent proteasomes and is involved in a late stage of nuclear proteasome assembly

Proteasomes are multisubunit proteases that are responsible for regulated proteolysis. The degradation of the proteasomal maturation factor, named Ump1 in yeast, completes the autocatalytic processing of inactive precursor complexes into the proteolytically active core particle (CP) of the proteasome. We have identified Blm3, a conserved nuclear protein, as a new component of Ump1-associated precursor complexes. A lack of Blm3 resulted in an increased rate of precursor processing and an accelerated turnover of Ump1, which suggests that Blm3 prevents premature activation of proteasomal CPs. On the basis of biochemical fractionation experiments combined with in vivo localization studies, we propose that Blm3 joins nascent CPs inside the nucleus to coordinate late stages of proteasome assembly in yeast.     

Cell force microscopy on elastic layers of finite thickness

Forces applied by cells to substrates can be measured using soft substrates with embedded displacement markers. Traction forces are retrieved from microscopic images by determining the displacements of these markers and fitting the generating forces. Here we show that using elastic films of 5-10-microm thickness one can improve the spatial resolution of the technique. To this end we derived explicit equations for the mechanical response of an elastic layer of finite thickness to point forces. Moreover, these equations allow highly accurate force measurements on eukaryotic cells on films where finite thickness effects are relevant (below approximately 60 microm).     

A Novel MiRNA-Based Predictive Model for Biochemical Failure Following Post-Prostatectomy Salvage Radiation Therapy

Purpose

To develop a microRNA (miRNA)-based predictive model for prostate cancer patients of 1) time to biochemical recurrence after radical prostatectomy and 2) biochemical recurrence after salvage radiation therapy following documented biochemical disease progression post-radical prostatectomy.

Methods

Forty three patients who had undergone salvage radiation therapy following biochemical failure after radical prostatectomy with greater than 4 years of follow-up data were identified. Formalin-fixed, paraffin-embedded tissue blocks were collected for all patients and total RNA was isolated from 1mm cores enriched for tumor (>70%). Eight hundred miRNAs were analyzed simultaneously using the nCounter human miRNA v2 assay (NanoString Technologies; Seattle, WA). Univariate and multivariate Cox proportion hazards regression models as well as receiver operating characteristics were used to identify statistically significant miRNAs that were predictive of biochemical recurrence.

Results

Eighty eight miRNAs were identified to be significantly (p<0.05) associated with biochemical failure post-prostatectomy by multivariate analysis and clustered into two groups that correlated with early (≤ 36 months) versus late recurrence (>36 months). Nine miRNAs were identified to be significantly (p<0.05) associated by multivariate analysis with biochemical failure after salvage radiation therapy. A new predictive model for biochemical recurrence after salvage radiation therapy was developed; this model consisted of miR-4516 and miR-601 together with, Gleason score, and lymph node status. The area under the ROC curve (AUC) was improved to 0.83 compared to that of 0.66 for Gleason score and lymph node status alone.

Conclusion

miRNA signatures can distinguish patients who fail soon after radical prostatectomy versus late failures, giving insight into which patients may need adjuvant therapy. Notably, two novel miRNAs (miR-4516 and miR-601) were identified that significantly improve prediction of biochemical failure post-salvage radiation therapy compared to clinico-histopathological factors, supporting the use of miRNAs within clinically used predictive models. Both findings warrant further validation studies.     

Analysis of climate change affecting German forests by combination of meteorological and phenological data within a GIS environment

The regional assessment of global change effects on plant phenology usually relies on local observations that need to be up-scaled. Therefore, methodological difficulties mostly related to data spatial resolution and congruency arise while performing broader-scale evaluations. Geostatiscs could be a useful tool to solve this type of problem, provided that a database with adequate spatial and temporal resolution is available. An assessment of variations in air temperature and plant phenology was carried out at the country level by using two German datasets regarding spring phenological phases of 15 plant species and air temperature. The data were collected from 1961-2002 at 1,279 and 675 sites, respectively. The annual mean air temperature in Germany was found to rise from 8.3 degrees C in the 1961-1990 period to 9.1 degrees C in the 1991-2002 term. The overall 15-species mean for the start of spring was found to be 6 days earlier in the latter period. The geostatistical analysis of the data revealed the suitability of Syringa vulgaris to be used as an indicator species to detect phenological changes in German forests. Moreover, their spatial patterns were found to be related to altitude and latitude. Therefore, geostatistics proved to be a useful tool to overcome some of the methodological problems related to the regional assessments of global change impacts on terrestrial ecosystems.