Elicitation of phenylpropanoids in maqui (Aristotelia chilensis [Mol.] Stuntz) plants micropropagated in photomixotrophic temporary immersion bioreactors (TIBs)

Plant Cell, Tissue and Organ Culture (PCTOC) - A biotechnological system for the production of plants biomass and phenylpropanoids of maqui was developed in photomixotrophic TIBs. The in vitro...     

Meta-Fish-Lib: A generalised,dynamic DNA reference library pipeline for metabarcoding of fishes

The accuracy and reliability of DNA metabarcoding analyses depend on the breadth and quality of the reference libraries that underpin them. However, there are limited options available to obtain and curate the huge volumes of sequence data that are available on public repositories such as NCBI and BOLD. Here, we provide a pipeline to download, clean and annotate mitochondrial DNA sequence data for a given list of fish species. Features of this pipeline include (a) support for multiple metabarcode markers; (b) searches on species synonyms and taxonomic name validation; (c) phylogeny assisted quality control for identification and removal of misannotated sequences; (d) automatically generated coverage reports for each new GenBank release update; and (e) citable, versioned DOIs. As an example we provide a ready-to-use curated reference library for the marine and freshwater fishes of the U.K. To augment this reference library for environmental DNA metabarcoding specifically, we generated 241 new MiFish-12S sequences for 88 U.K. marine species, and make available new primer sets useful for sequencing these. This brings the coverage of common U.K. species for the MiFish-12S fragment to 93%, opening new avenues for scaling up fish metabarcoding across wide spatial gradients. The Meta-Fish-Lib reference library and pipeline is hosted at https://github.com/genner-lab/meta-fish-lib .     

A proteomic study of Xanthomonas oryzae pv. oryzae in rice xylem sap

Xanthomonas oryzae pv. oryzae (Xoo) is the second most important rice pathogen, causing a disease called bacterial leaf blight. Xoo colonizes and infects the vascular tissue resulting in tissue necrosis and wilting causing significant yield losses worldwide. In this study Xoo infected vascular fluid (xylem sap) was recovered and analyzed for secreted Xoo proteins. Three independent experiments resulted in the identification of 324 different proteins, 64 proteins were found in all three samples which included many of the known virulence-associated factors. In addition, 10 genes encoding for the identified proteins were inactivated and one mutant displayed statistically a significant loss in virulence when compared to the wild type Xoo, suggesting that a new virulence-associated factor has been revealed. The usefulness of this approach in understanding the lifestyle and unraveling the virulence-associated factors of phytopathogenic vascular bacteria is discussed.     

Liver X Receptor (LXR) activation negatively regulates visfatin expression in macrophages

Adipose tissue macrophages (ATM) are the major source of visfatin, a visceral fat adipokine upregulated during obesity. Also known to play a role in B cell differentiation (pre-B cell colony-enhancing factor (PBEF)) and NAD biosynthesis (nicotinamide phosphoribosyl transferase (NAMPT)), visfatin has been suggested to play a role in inflammation.Liver X Receptor (LXR) and Peroxisome Proliferator-Activated Receptor (PPAR)γ are nuclear receptors expressed in macrophages controlling the inflammatory response. Recently, we reported visfatin as a PPARγ target gene in human macrophages. In this study, we examined whether LXR regulates macrophage visfatin expression. Synthetic LXR ligands decreased visfatin gene expression in a LXR-dependent manner in human and murine macrophages. The decrease of visfatin mRNA was paralleled by a decrease of protein secretion. Consequently, a modest and transient decrease of NAD⁺ concentration was observed. Interestingly, LXR activation decreased the PPARγ-induced visfatin gene and protein secretion in human macrophages.Our results identify visfatin as a gene oppositely regulated by the LXR and PPARγ pathways in human macrophages.     

Modulation of osteoclastogenesis in porcine bone marrow cultures by quercetin and rutin

Flavonols, in contrast to soybean isoflavones, are the most abundant phytoestrogens in western diets, being present in onions, beans, fruits, red wine, and tea. They may protect against atherosclerosis, inhibit certain cancer cell types, and reduce bone resorption. The most widely distributed flavonol is quercetin, which occurs mainly as its glycoside, rutin, but data are very scarce regarding the precise mechanism of action of these compounds on bone-resorbing cells at concentrations similar to those detected in human plasma. We have therefore investigated the effects of nanomolar concentrations of quercetin and rutin on the development and activity of osteoclasts in vitro compared with the effects of 17-estradiol. Nonadherent porcine bone marrow cells were cultured on dentine slices in the presence of 10 nM 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃), with or without 10 nM quercetin, 10 nM rutin or 10 nM 17-estradiol for 11 days. Multinuclear TRAP+ cells that resorbed dentine (osteoclasts) developed in the presence of 1,25(OH)₂D₃, but their number was significantly reduced by quercetin, rutin, and 17-estradiol (P < 0.05). Like 17-estradiol, both flavonols also significantly reduced resorption (P<0.05) as assessed by the size of pits resorbed on dentine slices. Osteoclasts and osteoclast progenitors contained estrogen receptor (ER), ER, and RANK proteins. Both flavonols increased nuclear ER protein and decreased ER protein of osteoclast progenitors. Moreover, rutin reduced RANK protein, whereas 17-oestradiol and quercetin promoted apoptosis by cleavage of caspase-8 and caspase-3. All the effects of flavonols were reversed by 1 M ICI 182,780, an estrogen antagonist. Thus, the anti-resorbing properties of flavonols are mainly mediated by ER proteins through the inhibition of RANK protein or the activation of caspases.C.M.R. was supported by a grant from CAPES (Department of Education, Brasilia, Brasil).     

Osmotic water permeability of rat intestinal brush border membrane vesicles: involvement of aquaporin-7 and aquaporin-8 and effect of metal ions.

Water channels AQP7 and AQP8 may be involved in transcellular water movement in the small intestine. We show that both AQP7 and AQP8 mRNA are expressed in rat small intestine. Immunoblot and immunohistochemistry experiments demonstrate that AQP7 and AQP8 proteins are present in the apical brush border membrane of intestinal epithelial cells. We investigated the effect of several metals and pH on the osmotic water permeability (Pf) of brush border membrane vesicles (BBMVs) and of AQP7 and AQP8 expressed in a cell line. Hg2+, Cu2+, and Zn2+ caused a significant decrease in the BBMV Pf, whereas Ni2+ and Li+ had no effect. AQP8-transfected cells showed a reduction in Pf in the presence of Hg2+ and Cu2+, whereas AQP7-transfected cells were insensitive to all tested metals. The Pf of both BBMVs and cells transfected with AQP7 and AQP8 was not affected by pH changes within the physiological range, and the Pf of BBMVs alone was not affected by phlorizin or amiloride. Our results indicate that AQP7 and AQP8 may play a role in water movement via the apical domain of small intestine epithelial cells. AQP8 may contribute to the water-imbalance-related clinical symptoms apparent after ingestion of high doses of Hg2+ and Cu2+.     

Different factors affecting human ANP amyloid aggregation and their implications in congestive heart failure

Aims

Atrial Natriuretic Peptide (ANP)-containing amyloid is frequently found in the elderly heart. No data exist regarding ANP aggregation process and its link to pathologies. Our aims were: i) to experimentally prove the presumptive association of Congestive Heart Failure (CHF) and Isolated Atrial Amyloidosis (IAA); ii) to characterize ANP aggregation, thereby elucidating IAA implication in the CHF pathogenesis.

Methods and Results

A significant prevalence (85%) of IAA was immunohistochemically proven ex vivo in biopsies from CHF patients. We investigated in vitro (using Congo Red, Thioflavin T, SDS-PAGE, transmission electron microscopy, infrared spectroscopy) ANP fibrillogenesis, starting from α-ANP as well as the ability of dimeric β-ANP to promote amyloid formation. Different conditions were adopted, including those reproducing β-ANP prevalence in CHF. Our results defined the uncommon rapidity of α-ANP self-assembly at acidic pH supporting the hypothesis that such aggregates constitute the onset of a fibrillization process subsequently proceeding at physiological pH. Interestingly, CHF-like conditions induced the production of the most stable and time-resistant ANP fibrils suggesting that CHF affected people may be prone to develop IAA.

Conclusions

We established a link between IAA and CHF by ex vivo examination and assessed that β-ANP is, in vitro, the seed of ANP fibrils. Our results indicate that β-ANP plays a crucial role in ANP amyloid deposition under physiopathological CHF conditions. Overall, our findings indicate that early IAA-related ANP deposition may occur in CHF and suggest that these latter patients should be monitored for the development of cardiac amyloidosis.     

Photophysical and electrochemical properties of a fullerene-stoppered rotaxane.

The photophysical and electrochemical properties of a fumaramide rotaxane stoppered with C(60) are reported. The results evidenced the strong binding interactions between the template and the macrocycle, which are also supported by molecular modelling.