Advanced glycation end-products (AGEs) induce concerted changes in the osteoblastic expression of their receptor RAGE and in the activation of extracellular signal-regulated kinases (ERK)

An increase in the interaction between advanced glycation end-products (AGEs) and their receptor RAGE is believed to contribute to the pathogenesis of chronic complications of Diabetes mellitus, which can include bone alterations such as osteopenia. We have recently found that extracellular AGEs can directly regulate the growth and development of rat osteosarcoma UMR106 cells, and of mouse calvaria-derived MC3T3E1 osteoblasts throughout their successive developmental stages (proliferation, differentiation and mineralisation), possibly by the recognition of AGEs moieties by specific osteoblastic receptors which are present in both cell lines. In the present study we examined the possible expression of RAGE by UMR106 and MC3T3E1 osteoblastic cells, by immunoblot analysis. We also investigated whether short-, medium- or long-term exposure of osteoblasts to extracellular AGEs, could modify their affinity constant and maximal binding for AGEs (by 125I-AGE-BSA binding experiments), their expression of RAGE (by immunoblot analysis) and the activation status of the osteoblastic ERK 1/2 signal transduction mechanism (by immunoblot analysis for ERK and P-ERK). Our results show that both osteoblastic cell lines express readily detectable levels of RAGE. Short-term exposure of phenotypically mature osteoblastic UMR106 cells to AGEs decrease the cellular density of AGE-binding sites while increasing the affinity of these sites for AGEs. This culture condition also dose-dependently increased the expression of RAGE and the activation of ERK. In proliferating MC3T3E1 pre-osteoblasts, 24–72 h exposure to AGEs did not modify expression of RAGE, ERK activation or the cellular density of AGE-binding sites. However, it did change the affinity of these binding sites for AGEs, with both higher- and lower-affinity sites now being apparent. Medium-term (1 week) incubation of differentiated MC3T3E1 osteoblasts with AGEs, induced a simultaneous increase in RAGE expression and in the relative amount of P-ERK. Mineralising MC3T3E1 cultures grown for 3 weeks in the presence of extracellular AGEs showed a decrease both in RAGE and P-ERK expression. These results indicate that, in phenotypically mature osteoblastic cells, changes in ERK activation closely follow the AGEs-induced regulation of RAGE expression. Thus, the AGEs-induced biological effects that we have observed previously in osteoblasts, could be mediated by RAGE in the later stages of development, and mediated by other AGE receptors in the earlier pre-osteoblastic stage.     

Analysis of the expression of the <Emphasis Type="Italic">Rhodobacter sphaeroides lexA</Emphasis> gene

The regulation of the Rhodobacter sphaeroides lexA gene has been analyzed using both gel-mobility experiments and lacZ gene fusions. PCR-mediated mutagenesis demonstrated that the second GAAC motif in the sequence GAACN₇GAACN₇GAAC located upstream of the R. sphaeroides lexA gene is absolutely necessary for its DNA damage-mediated induction. Moreover, mutagenesis of either the first or the third GAAC motif in this sequence reduced, but did not abolish, the inducibility of the R. sphaeroides lexA gene. A R. sphaeroides lexA-defective (Def) mutant has also been constructed by replacing the active lexA gene with an inactivated gene copy constructed in vitro. Crude extracts of the R. sphaeroides lexA(Def) strain are unable to form any protein-DNA complex when added to the wild-type lexA promoter of R. sphaeroides. Likewise, the R. sphaeroides lexA(Def) cells constitutively express the recA and lexA genes. All these data clearly indicate that the lexA gene product is the negative regulator of the R. sphaeroides SOS response. Furthermore, the morphology, growth and viability of R. sphaeroides lexA(Def) cultures do not show any significant change relative to those of the wild-type strain. Hence, R. sphaeroides is so far the only bacterial species whose viability is known not to be affected by the presence of a lexA(Def) mutation.     

Variations in competitive mechanisms of captive male hamadryas-like baboons in two feeding situations

Experiments were carried out to determine the influence of food distribution (clumped vs dispersed) on processes of competition among seven captive male hamadryas-like baboons. Spatial cohesion, cohesive behaviours that males performed and received from their females, and aggressive and feeding behaviours of the males were collected during 35 min after food supply. Median values and variability of these variables were compared in both the feeding situations. Food distribution influenced both asymmetries in access and the form of competition. When food was clumped, those with less access fed by means of compensatory or alternative strategies (away from the main source and/or by feeding supplants), and they were more aggressive towards other males. When food was dispersed, all individuals could feed simultaneously, males that were more aggressive were those with less cohesive OMUs, despite the fact that cohesive behaviours remained constant. Females appeared to contribute to variation in the OMU's cohesion through their responses to both food distribution and males' competitive ability.     

Characterisation of the cell walls of loquat (Eriobotrya japonica L.) fruit tissues

Loquat fruit (Eriobotrya japonica L. cv. Algor) was dissected to give the following tissue zones: epidermis or epicarp, flesh or mesocarp, integument (a thin layer surrounding the seed cotyledons), seed testa, kernel and hairy receptacle. The alcohol insoluble residues (AIRs) from all these tissues were proved to be free of starch, except loquat kernel which on a fresh weight basis contained about 34% of starch. AIRs were analysed for moisture, ashes, protein, lignin and the component sugars were released by two hydrolytic procedures which helped to distinguish the sugars from non-cellulosic polysaccharides and cellulose. Their major component polysaccharides were inferred to be pectic polysaccharides since all AIRs were very rich in sugars such as uronic acids, arabinose and galactose. Pectic polysaccharides contributed up to 70% of total cell wall polysaccharides in the edible flesh of the loquat fruit. Important differences in the degree of branching, degree of esterification and in the amounts of Ca and Mg associated with pectic polysaccharides were detected among pectic polymers depending on the loquat tissue zone. These compositional and structural differences may be related to the role that these pectic polymers play within the tissues which form the loquat fruit.     

New antifungal peptides. Synthesis,bioassays and initial structure prediction by CD spectroscopy

The synthesis, in vitro evaluation and conformational study of KKWKMRRNQFWIKIQR-NH₂, HFRWRQIKIWFQNRRMKWKK-NH₂ and RQPKIWFPNRRKPWKK-NH₂ acting as antifungal agents are reported. These peptides displayed a moderate but significant antifungal effect against both pathogenic fungi Candida albicans and Cryptococcus neoformans. The conformational analysis of these peptides was carried out using both theoretical and experimental methods.     

Modulation of lyso-platelet activating factor:acetyl-CoA acetyltransferase from rat splenic microsomes. The role of cyclic AMP-dependent protein kinase

Incubation of rat splenic microsomes with the catalytic subunit of cyclic AMP-dependent protein kinase in the presence of Mg-ATP stimulated 2-3-fold lyso-platelet-activating factor:acetyltransferase activity. This activation was due to an increase in the $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ of the acetylation reaction, whereas the $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for acetyl-CoA was not affected. The ATP derivative, AMPPNP, could not replace ATP and preincubation of the microsomes with the heat-stable inhibitor of protein kinase prevented the activation by Mg-ATP obtained in the presence of the protein kinase. Activation of the acetylation reaction by the protein kinase was reversible. Evidence is provided that the reversal of activation is due to dephosphorylation of the enzyme. These data provide evidence that in vitro lyso-platelet-activating factor:acetyltransferase from splenic microsomes is regulated by phosphorylation.     

Effects of small barriers on habitat use by red deer: Implications for conservation practices

Artificial barriers such as wire fences constitute a common conservation management practice to protect vegetation from the browsing impact of large herbivores. Apart from protecting the fenced area, these barriers may affect the use of adjacent areas by animals. For example, they may interrupt major movement routes. We studied the effect of fences on an area used by red deer in Doñana National Park (Andalucía, south west Spain). We used an observational approach to study the effect of existing permanent barriers, and an experimental approach to investigate the effect of new barriers placed between the main areas through which animals moved from resting to foraging sites. Our study was carried out during the mating season, so we could also observe the effects on the distribution of females among harems. We found that “shadow areas”, where projection lines of the movement from resting to foraging areas were interrupted by the barrier, were used less by deer. In agreement with this result, grasses in shadow areas tended to be longer than in other areas, indicating unequal use of resources depending on the placement of barriers. Also, permanent barriers appeared to have negative, long-term effects on the maintenance of meadows as denoted by a higher proportion of rushes in meadows within shadow areas. Experimental barriers supported the cause-effect relationship by decreasing the number of deer using the experimental shadow areas. Our results demonstrated unexpected ecological effects of small barriers on the landscape, mediated by modification of the spatial behaviour of red deer.     

Phylogeny of Macroptilium (Leguminosae): morphological, biochemical and molecular evidence

Macroptilium (Benth.) Urban (Phaseoleae, Papilionoideae, Leguminosae) is an American genus of legumes, belonging to subtribe Phaseolinae along with other economically important genera, such as Vigna Savi and Phaseolus L. (the common bean genus). Cladistic analyses based on morphological, biochemical (storage seed proteins) and molecular (nuclear and plastid DNA sequences) data were performed on the 18 species currently ascribed to the genus, exploring several character weighting strategies. Equal weights, implied weighting and different transversion/transition costs were applied. The three data sets were first analyzed with separate partitions, and then combined into a single matrix. This study is the first one to analyze all the species of the genus from a cladistic point of view. In all the most parsimonious trees obtained, Macroptilium is monophyletic with excellent support values. Two monophyletic clades are recovered in almost all the analyses. Both are compound by nine species, and they constitute two sections of Macroptilium. Several interspecific relationships inside the genus are discussed. © The Willi Hennig Society 2007.     

Electronic Tagging of Atlantic Bluefin Tuna (Thunnus thynnus,L.) Reveals Habitat Use and Behaviors in the Mediterranean Sea

We analyzed the movements of Atlantic tuna (Thunnus thynnus L.) in the Mediterranean Sea using data from 2 archival tags and 37 pop-up satellite archival tags (PAT). Bluefin tuna ranging in size from 12 to 248 kg were tagged on board recreational boats in the western Mediterranean and the Adriatic Sea between May and September during two different periods (2000 to 2001 and 2008 to 2012). Although tuna migrations between the Mediterranean Sea and the Atlantic Ocean have been well reported, our results indicate that part of the bluefin tuna population remains in the Mediterranean basin for much of the year, revealing a more complex population structure. In this study we demonstrate links between the western Mediterranean, the Adriatic and the Gulf of Sidra (Libya) using over 4336 recorded days of location and behavior data from tagged bluefin tuna with a maximum track length of 394 days. We described the oceanographic preferences and horizontal behaviors during the spawning season for 4 adult bluefin tuna. We also analyzed the time series data that reveals the vertical behavior of one pop-up satellite tag recovered, which was attached to a 43.9 kg tuna. This fish displayed a unique diving pattern within 16 days of the spawning season, suggesting a use of the thermocline as a thermoregulatory mechanism compatible with spawning. The results obtained hereby confirm that the Mediterranean is clearly an important habitat for this species, not only as spawning ground, but also as an overwintering foraging ground.