Peripheral ghrelin interacts with orexin neurons in glucostatic signalling

Ghrelin interactions with glycemia in appetite control as well as the potential mechanisms involving the orexin and melanin-concentrating hormone (MCH) neurons in the orexigenic ghrelin signals were investigated by using a specific anti-ghrelin antibody (AGA). Our results confirm that peripheral ghrelin is an important signal in meal initiation and appetite. Employing immunohistochemistry techniques, we found that c-fos positive neurons in the lateral hypothalamus (LH) and perifornical area (PFA) increased after insulin or 2-deoxyglucose administration. Moreover, we have also demonstrated that peripheral ghrelin blockade by the AGA, reduces the orexigenic signal induced by insulin and 2-DG administration probably partly producing a decrease of c-fos immunoreactivity in the LH and PFA as well as a lower activation of orexin neurons. In contrast, the c-fos positive MCH neurons were not apparently affected. In summary, our findings suggest that peripheral ghrelin plays an important role in regulatory "glucostatic" feeding mechanisms by means of its role as a "hunger" signal affecting the LH and PFA areas, which may contribute to energy homeostasis through orexin neurons.     

Astroglial-Conditioned Media and Growth Factors Modulate Proliferation and Differentiation of Astrocytes in Primary Culture

Astroglial conditioned media (ACM) influence the development and maturation of cultured nerve cells and modulate neuron-glia interaction. To clarify mechanisms of astroglial cell proliferation/differentiation in culture, incorporation of [methyl-³H]-thymidine or [5,6-³H]-uridine in cultured astrocytes was assessed. Cultures were pre-treated with epidermal growth factor (EGF), insulin (INS), insulin-like growth factor-I (IGF-I), and basic fibroblast growth factor (bFGF) and subsequently with ACM. DNA labeling revealed a marked stimulatory effect of ACM from 15 days in vitro (DIV) cultures in 30 DIV astrocytes after12 h pre-treatment with growth factors. The main effects were found after INS or EGF pre-treatment in 30 DIV cultures. ACM collected from 15 or 60 or 90 DIV increased RNA labeling of 15 and 30 DIV astrocyte cultures, being the highest value that of 30 DIV cultures added with ACM from 90 DIV. The findings of increased DNA labeling after EGF or INS pre-treatment in 30 DIV cultures, followed by addition of ACM from 15 DIV cultures, suggest that these phenomena may depend by extra cellular signal-regulated kinase 1 (ERK1) activation.     

Inflammation may modulate IL-6 and C-reactive protein gene expression in the adipose tissue: the role of IL-6 cell membrane receptor

Only few studies have been addressed to the presence and regulation of C-reactive protein (CRP) gene expression in different districts of adipose tissue, and no study has investigated the role of adipose tissue in presence of inflammation. Therefore, the aim of this study was to investigate the inflammatory involvement of either adipose tissue or adipose cells (adipocytes and stromal cells, respectively) in patients with chronic inflammatory disease, focusing on regional adipose tissue CRP gene expression. Eighteen patients with inflammatory disease and 14 healthy controls were enrolled. All subjects underwent specific surgical procedures. Inflamed and noninflamed patients provided samples of subcutaneous and/or omental adipose tissue. All samples were analyzed by RT-PCR and real-time PCR for specific gene expression. In addition, both adipocytes and stromal cells were studied by real-time PCR and immunoprecipitation to evaluate either gene or protein expression of CRP. Our results (real-time PCR) demonstrated a higher gene expression of CRP, IL-6, and both IL-6 membrane receptors in subcutaneous samples of inflamed patients than in healthy controls. Furthermore, in omental fragments of inflamed patients, an enhanced mRNA abundance of the same genes, compared with subcutaneous, was observed. The results obtained at cellular level did not provide evidence of any difference between adipocytes and stromal cell CRP gene expression, whereas immunoprecipitation demonstrated the presence of CRP in inflamed subjects. These results provide first-time evidence of the involvement of adipose tissue in the course of chronic inflammatory diseases, with a different degree of participation of the different adipose tissue districts.     

Trade-offs across life stages: does predator–induced hatching plasticity reduce anuran post-metamorphic performance?

In species with complex life cycles hatching plasticity can provide an effective escape from egg predators, but theoretical studies predict a predation-risk trade-off across egg and larval stages. In this study, we examine whether the presence of an egg predator can alter the timing of hatching in an anuran, Rana temporaria, and the consequences of hatching plasticity after transition to the terrestrial habitat. Predator cues induced earlier hatching, and hatchlings were smaller, less developed and had relatively shorter and deeper tails than control hatchlings. The predator–induced differences in developmental time were compensated throughout the larval period; there was no predator effect on metamorph age or size. Surprisingly, the effects of egg predators were perceptible after metamorphosis. Juveniles emerging from the predator and the no-predator treatments differed in several size-adjusted morphological dimensions. Seemingly these morphological differences were not large enough to give rise to suboptimal growth or locomotor performance after metamorphosis. Thus, our results suggest only a short-term effect on juvenile phenotype, but not a trade-off between hatching time and juvenile performance.     

Tracing past human male movements in northern/eastern Africa and western Eurasia: new clues from Y-chromosomal haplogroups E-M78 and J-M12

Detailed population data were obtained on the distribution of novel biallelic markers that finely dissect the human Y-chromosome haplogroup E-M78. Among 6,501 Y chromosomes sampled in 81 human populations worldwide, we found 517 E-M78 chromosomes and assigned them to 10 subhaplogroups. Eleven microsatellite loci were used to further evaluate subhaplogroup internal diversification. The geographic and quantitative analyses of haplogroup and microsatellite diversity is strongly suggestive of a northeastern African origin of E-M78, with a corridor for bidirectional migrations between northeastern and eastern Africa (at least 2 episodes between 23.9-17.3 ky and 18.0-5.9 ky ago), trans-Mediterranean migrations directly from northern Africa to Europe (mainly in the last 13.0 ky), and flow from northeastern Africa to western Asia between 20.0 and 6.8 ky ago. A single clade within E-M78 (E-V13) highlights a range expansion in the Bronze Age of southeastern Europe, which is also detected by haplogroup J-M12. Phylogeography pattern of molecular radiation and coalescence estimates for both haplogroups are similar and reveal that the genetic landscape of this region is, to a large extent, the consequence of a recent population growth in situ rather than the result of a mere flow of western Asian migrants in the early Neolithic. Our results not only provide a refinement of previous evolutionary hypotheses but also well-defined time frames for past human movements both in northern/eastern Africa and western Eurasia.     

Monte Carlo folding of trans-membrane helical peptides in an implicit generalized Born membrane

An efficient Monte Carlo (MC) algorithm using concerted backbone rotations is combined with a recently developed implicit membrane model to simulate the folding of the hydrophobic transmembrane domain M2TM of the M2 protein from influenza A virus and Sarcolipin at atomic resolution. The implicit membrane environment is based on generalized Born theory and has been calibrated against experimental data. The MC sampling has previously been used to fold several small polypeptides and been shown to be equivalent to molecular dynamics (MD). In combination with a replica exchange algorithm, M2TM is found to form continuous membrane spanning helical conformations for low temperature replicas. Sarcolipin is only partially helical, in agreement with the experimental NMR structures in lipid bilayers and detergent micelles. Higher temperature replicas exhibit a rapidly decreasing helicity, in agreement with expected thermodynamic behavior. To exclude the possibility of an erroneous helical bias in the simulations, the model is tested by sampling a synthetic Alanine-rich polypeptide of known helicity. The results demonstrate there is no overstabilization of helical conformations, indicating that the implicit model captures the essential components of the native membrane environment for M2TM and Sarcolipin.     

The heparin III-binding domain of fibronectin (III4–5 repeats) binds to fibronectin and inhibits fibronectin matrix assembly

Fibronectin matrix assembly involves interactions among various regions of the molecule, which contribute to elongation and stabilization of the fibrils. In this study, we examined the possible role of the heparin III domain of fibronectin (repeats III4-5) in fibronectin fibrillogenesis. We show that a recombinant fragment comprising these repeats (FNIII4-5 fragment) blocked fibronectin fibril formation and the incorporation of 125I-fibronectin into cell layers. Binding assays using a biosensor revealed that FNIII4-5 bound fibronectin and the amino-terminal 70 kDa and 29 kDa fragments. It also bound to itself, indicating a previously unidentified self-association site in repeats III4-5. These interactions were specific since FNIII4-5 did not bind to the FNIII7-10 fragment, representing a central region in fibronectin. The fibronectin-binding property of the III4-5 domain, but not its matrix assembly inhibitory function, was apparently cryptic in larger fragments. By mutating the arginine residues in the WTPPRAQITGYRLTVGLTRR proteoglycan-binding sequence (HBP/III5 site) of FNIII4-5 [Moyano, J.V., Carnemolla, B., Albar, J.P., Leprini, A., Gaggero, B., Zardi, L., Garcia-Pardo, A., 1999. Cooperative role for activated alpha4beta1 integrin and chondroitin sulfate proteoglycans in cell adhesion to the heparin III domain of fibronectin. Identification of a novel heparin and cell binding sequence in repeat III5. J. Biol. Chem. 274, 135-142.], we found that the first two arginine residues in HBP/III5 were involved in the fibronectin-binding property of FNIII4-5, while the last two arginine residues in HBP/III5 were required for inhibition of matrix assembly and the binding of 125I-fibronectin to cell layers. Both properties appear to function independently from each other, depending on the conformation of the fibronectin dimer.     

Contrasting strategies to cope with drought by invasive and endemic species of Lantana in Galapagos

This study compares how Lantana camara, an invasive species, and L. peduncularis, an autochthonous one, cope with drought in Galapagos. Soil surface temperature was the abiotic environmental parameter that best explained variations in photosynthetic stress. Higher soil surface temperatures were recorded in the lowlands and in rain-shadow areas, which were also the driest areas. L. peduncularis, with a shallow root system, behaved as a drought-tolerant species, showing lower relative growth rates, which decreased with leaf water content and higher photosynthetic stress levels in the lowlands and in a northwest rain-shadow area in comparison with higher and wetter locations. Its basal and maximal fluorescences decreased at lower altitudes, reflecting the recorded drops in chlorophyll concentration. In contrast, L. camara with a deep root system behaved as a drought-avoiding species, showing leaf and relative water contents higher than 55% and avoiding permanent damage to its photosynthetic apparatus even in the driest area where it showed very low chlorophyll content. Its relative growth rate decreased more in dry areas in comparison to wetter zones than did that of L. peduncularis, even though it had greater water content. Furthermore, L. camara showed higher water contents, growth rate, and lower photosynthetic stress levels than L. peduncularis in the arid lowlands. Thus, L. peduncularis maintained lower maximum quantum efficiency of photosystem II photochemistry (F _v/F _m) than L. camara even at sunrise, due to higher basal fluorescence values with similar maximal fluorescence, which indicated permanent damage to PSII reaction centres. Our results help to explain the success and limitations of L. camara in the invasion of arid and sub-arid environments.     

Induction of GRP78 by valproic acid is dependent upon histone deacetylase inhibition

Valproic (2-propylpentanoic) acid is a commonly used drug in the treatment of bipolar disorder and epilepsy. The molecular mechanism that underlies its clinical efficacy remains controversial and is complicated by the broad range of intracellular effects of valproic acid, including its ability to inhibit histone deacetylase (HDAC) and induce protein chaperone expression. Here we show that an established HDAC inhibitor, trichostatin A, promotes ER chaperone expression in HEK293 cells. Furthermore, we use chemical derivatives of valproic acid to show that the ability to promote GRP78 levels directly correlates with the induction of histone H4 hyperacetylation. These results suggest that exposure to valproic acid enhances chaperone expression by a mechanism that involves histone hyperacetylation.