Effect of age on surface molecules and cytokine expression in human dendritic cells

Dendritic cells (DCs) are central in regulating both innate and acquired immunity, but their possible age-related functional modifications are still unclear. Here we have analyzed the effect of age on LPS-treated monocyte-derived DCs (MDDCs). A negative correlation between age and cell expression of ICAM-1, CD25 and IL-10 was observed in a group of healthy donors. This has been confirmed by a significantly reduced expression of the same molecules in cells of subgrouped elderly versus younger individuals. On the contrary, a positive correlation between age and cell expression of IL-6 and IL-18 has been reported in all the subjects and further supported by a significant increase of the two pro-inflammatory cytokines in cells of elderly versus young subjects.Our data indicate that aging can impair the expression of ICAM-1 and CD25 and skew the production of cytokines, including IL-18, which concur to make a pro-inflammatory milieu. Altogether, the present results point to additional molecules whose role might be relevant in immunosenescence of human DCs, confirming that these cells undergo functional changes during aging.     

WCI,a novel wheat chymotrypsin inhibitor: purification,primary structure,inhibitory properties and heterologous expression

A novel chymotrypsin inhibitor, detected in the endosperm of Triticum aestivum, was purified and characterized with respect to the main physical–chemical properties. On the basis of its specificity, this inhibitor was named WCI (wheat chymotrypsin inhibitor). WCI is a monomeric neutral protein made up of 119 residues and molecular mass value of 12,933.40 Da. Automated sequence and mass spectrometry analyses, carried out on several samples of purified inhibitor, evidenced an intrinsic molecular heterogeneity due to the presence of the isoform [des-(Thr)WCI], accounting for about 40% of the total sample. In vitro, WCI acted as a strong inhibitor of bovine pancreatic chymotrypsin as well as of chymotryptic-like activities isolated from the midgut of two phytophagous insects, Helicoverpa armigera (Hüb.) and Tenebrio molitor L., respectively. No inhibitory activities were detected against bacterial subtilisins, bovine pancreatic trypsin, porcine pancreatic elastase or human leukocyte elastase. The primary structure of WCI was significantly similar (45.7–89.1%) to those of several proteins belonging to the cereal trypsin/α-amylase inhibitor super-family and showed the typical sequence motif of this crowed protein group. The cDNA of the inhibitor (wci-cDNA) was isolated from wheat immature caryopses and employed to obtain a recombinant product in E. coli. Experimental evidences indicated that the recombinant inhibitor was localized in the inclusion bodies from which it was recovered as soluble and partially active protein by applying an appropriate refolding procedure. WCI reactive site localization, as well as its inhibitory specificity, was investigated by molecular modeling approach.     

Overexpression of PhEXPA1 increases cell size, modifies cell wall polymer composition and affects the timing of axillary meristem development in Petunia hybrida

? Expansins are cell wall proteins required for cell enlargement and cell wall loosening during many developmental processes. The involvement of the Petunia hybrida expansin A1 (PhEXPA1) gene in cell expansion, the control of organ size and cell wall polysaccharide composition was investigated by overexpressing PhEXPA1 in petunia plants. ? PhEXPA1 promoter activity was evaluated using a promoter-GUS assay and the protein's subcellular localization was established by expressing a PhEXPA1-GFP fusion protein. PhEXPA1 was overexpressed in transgenic plants using the cauliflower mosaic virus (CaMV) 35S promoter. Fourier transform infrared (FTIR) and chemical analysis were used for the quantitative analysis of cell wall polymers. ? The GUS and GFP assays demonstrated that PhEXPA1 is present in the cell walls of expanding tissues. The constitutive overexpression of PhEXPA1 significantly affected expansin activity and organ size, leading to changes in the architecture of petunia plants by initiating premature axillary meristem outgrowth. Moreover, a significant change in cell wall polymer composition in the petal limbs of transgenic plants was observed. ? These results support a role for expansins in the determination of organ shape, in lateral branching, and in the variation of cell wall polymer composition, probably reflecting a complex role in cell wall metabolism.     

Partial migration in roe deer: migratory and resident tactics are end points of a behavioural gradient determined by ecological factors

Ungulate populations exhibiting partial migration present a unique opportunity to explore the causes of the general phenomenon of migration. The European roe deer Capreolus capreolus is particularly suited for such studies due to a wide distribution range and a high level of ecological plasticity. In this study we undertook a comparative analysis of roe deer GPS location data from a representative set of European ecosystems available within the EURODEER collaborative project. We aimed at evaluating the ecological factors affecting migration tactic (i.e. occurrence) and pattern (i.e. timing, residence time, number of migratory trips). Migration occurrence varied between and within populations and depended on winter severity and topographic variability. Spring migrations were highly synchronous, while the timing of autumn migrations varied widely between regions, individuals and sexes. Overall, roe deer were faithful to their summer ranges, especially males. In the absence of extreme and predictable winter conditions, roe deer seemed to migrate opportunistically, in response to a tradeoff between the costs of residence in spatially separated ranges and the costs of migratory movements. Animals performed numerous trips between winter and summer ranges which depended on factors influencing the costs of movement such as between‐range distance, slope and habitat openness. Our results support the idea that migration encompasses a behavioural continuum, with one‐trip migration and residence as its end points, while commuting and multi‐trip migration with short residence times in seasonal ranges are intermediate tactics. We believe that a full understanding of the variation in tactics of temporal separation in habitat use will provide important insights on migration and the factors that influence its prevalence.     

Hypoxia stimulates the production of the angiogenesis inhibitor 2-methoxyestradiol by swine granulosa cells

We previously demonstrated the presence of 2-methoxyestradiol (2-ME) in swine follicular fluid. Present study was aimed first of all to investigate if swine granulosa cell produce 2-ME; in addition, we tried to assess a potential effect of hypoxia in modulating 2-ME output. Finally, we explored the effect of 2-ME in an angiogenesis bioassay set up in our lab. Our data show that cultured granulosa cells are able to produce 2-ME; interestingly, the secretion of the hormone appeared to be stimulated by hypoxia. Angiogenesis bioassay points out that 2-ME displays an inhibitory effect on neovascularisation. Therefore our data suggest that 2-ME could be a local effector in determining the fine tuning responsible for follicle angiogenesis. These data deserve special attention since the ovary is a valuable experimental model in angiogenesis research.     

Isolation and characterization of a psychropiezophilic alphaproteobacterium

Cultivated psychropiezophilic (low-temperature- and high-pressure-adapted) bacteria are currently restricted to phylogenetically narrow groupings capable of growth under nutrient-replete conditions, limiting current knowledge of the extant functional attributes and evolutionary constraints of diverse microorganisms inhabiting the cold, deep ocean. This study documents the isolation of a deep-sea bacterium following dilution-to-extinction cultivation using a natural seawater medium at high hydrostatic pressure and low temperature. To our knowledge, this isolate, designated PRT1, is the slowest-growing (minimal doubling time, 36 h) and lowest cell density-producing (maximal densities of 5.0 × 10⁶ cells ml⁻¹) piezophile yet obtained. Optimal growth was at 80 MPa, correlating with the depth of capture (8,350 m), and 10°C, with average cell sizes of 1.46 μm in length and 0.59 μm in width. Through detailed growth studies, we provide further evidence for the temperature-pressure dependence of the growth rate for deep-ocean bacteria. PRT1 was phylogenetically placed within the Roseobacter clade, a bacterial lineage known for widespread geographic distribution and assorted lifestyle strategies in the marine environment. Additionally, the gene transfer agent (GTA) g5 capsid protein gene was amplified from PRT1, indicating a potential mechanism for increased genetic diversification through horizontal gene transfer within the hadopelagic environment. This study provides a phylogenetically novel isolate for future investigations of high-pressure adaptation, expands the known physiological traits of cultivated members of the Roseobacter lineage, and demonstrates the feasibility of cultivating novel microbial members from the deep ocean using natural seawater.     

AKT/FOXO signaling enforces reversible differentiation blockade in myeloid leukemias

AKT activation is associated with many malignancies, where AKT acts, in part, by inhibiting FOXO tumor suppressors. We show a converse role for AKT/FOXOs in acute myeloid leukemia (AML). Rather than decreased FOXO activity, we observed that FOXOs are active in ～40% of AML patient samples regardless of genetic subtype. We also observe this activity in human MLL-AF9 leukemia allele-induced AML in mice, where either activation of Akt or compound deletion of FoxO1/3/4 reduced leukemic cell growth, with the latter markedly diminishing leukemia-initiating cell (LIC) function in vivo and improving animal survival. FOXO inhibition resulted in myeloid maturation and subsequent AML cell death. FOXO activation inversely correlated with JNK/c-JUN signaling, and leukemic cells resistant to FOXO inhibition responded to JNK inhibition. These data reveal a molecular role for AKT/FOXO and JNK/c-JUN in maintaining a differentiation blockade that can be targeted to inhibit leukemias with a range of genetic lesions.     

CD14 controls the LPS-induced endocytosis of Toll-like receptor 4

The transport of Toll-like Receptors (TLRs) to various organelles has emerged as an essential means by which innate immunity is regulated. While most of our knowledge is restricted to regulators that promote the transport of newly synthesized receptors, the regulators that control TLR transport after microbial detection remain unknown. Here, we report that the plasma membrane localized Pattern Recognition Receptor (PRR) CD14 is required for the microbe-induced endocytosis of TLR4. In dendritic cells, this CD14-dependent endocytosis pathway is upregulated upon exposure to inflammatory mediators. We identify the tyrosine kinase Syk and its downstream effector PLCγ2 as important regulators of TLR4 endocytosis and signaling. These data establish that upon microbial detection, an upstream PRR (CD14) controls the trafficking and signaling functions of a downstream PRR (TLR4). This innate immune trafficking cascade illustrates how pathogen detection systems operate to induce both membrane transport and signal transduction.