Anna Rita Taddei

Immunoglobulin protein and gene transcripts in ovarian follicles throughout oogenesis in the teleost Dicentrarchus labrax

Transfer of immunoglobulins (IgM-like) from the female to the teleost embryo has been demonstrated but mechanisms of uptake into and storage within the eggs remain to be clarified. The monoclonal antibody DLIg3 against Dicentrarchus labrax Ig light chain revealed an active role of both follicle cells and oocytes in the Ig uptake. The primordial follicular cells showed DLIg3 immunoreactivity even at a pre-vitellogenetic stage. Early vitellogenetic oocytes (lipid vesicle stages) had DLIg3 staining of pore canals, plasmalemma and outer cortex and of their follicular cells. In protein yolk granule oocytes, DLIg3 staining was also detected within vesicles of the outer-mid cortex and juxtanuclear yolk granules; therefore, a centripetal transport of Ig throughout oocyte development is apparently carried out. Immunoelectron microscopy confirmed the presence of Ig within thecal and granulosa cells (and in the interposed basement membrane) of pre-vitellogenic and vitellogenic follicles. Thus, the transport of Ig to the egg apparently occurs also by transcytosis across the follicle cells. Igs were localised in the pore canals surronding the microvilli and in vesicles of outer-mid cortex of vitellogenic oocytes. Reverse transcription/polymerase chain reaction with primers designed for the constant region of sea bass Ig light chain detected Ig mRNA in hydrated oocytes, a smaller content in released eggs and no signal in larvae at day two post-hatching. These findings show that a significant level of Ig gene transcription in the oocyte and/or a transfer of transcripts may also occur.

Pd-nanoparticles cause increased toxicity to kiwifruit pollen compared to soluble Pd(II).

In the present study, endpoints including in vitro pollen performance (i.e., germination and tube growth) and lethality were used as assessments of nanotoxicity. Pollen was treated with 5-10 nm-sized Pd particles, similar to those released into the environment by catalytic car exhaust converters. Results showed Pd-nanoparticles altered kiwifruit pollen morphology and entered the grains more rapidly and to a greater extent than soluble Pd(II). At particulate Pd concentrations well below those of soluble Pd(II), pollen grains experienced rapid losses in endogenous calcium and pollen plasma membrane damage was induced. This resulted in severe inhibition and subsequent cessation of pollen tube emergence and elongation at particulate Pd concentrations as low as 0.4 mg L(-1). Particulate Pd emissions related to automobile traffic have been increasing and are accumulating in the environment. This could seriously jeopardize in vivo pollen function, with impacts at an ecosystem level.

Lactobacillus rhamnosus lowers zebrafish lipid content by changing gut microbiota and host transcription of genes involved in lipid metabolism

The microbiome plays an important role in lipid metabolism but how the introduction of probiotic communities affects host lipid metabolism is poorly understood. Using a multidisciplinary approach we addressed this knowledge gap using the zebrafish model by coupling high-throughput sequencing with biochemical, molecular and morphological analysis to evaluate the changes in the intestine. Analysis of bacterial 16S libraries revealed that Lactobacillus rhamnosus was able to modulate the gut microbiome of zebrafish larvae, elevating the abundance of Firmicutes sequences and reducing the abundance of Actinobacteria. The gut microbiome changes modulated host lipid processing by inducing transcriptional down-regulation of genes involved in cholesterol and triglycerides metabolism (fit2, agpat4, dgat2, mgll, hnf4α, scap, and cck) concomitantly decreasing total body cholesterol and triglyceride content and increasing fatty acid levels. L. rhamnosus treatment also increased microvilli and enterocyte lengths and decreased lipid droplet size in the intestinal epithelium. These changes resulted in elevated zebrafish larval growth. This integrated system investigation demonstrates probiotic modulation of the gut microbiome, highlights a novel gene network involved in lipid metabolism, provides an insight into how the microbiome regulates molecules involved in lipid metabolism, and reveals a new potential role for L. rhamnosus in the treatment of lipid disorders.

The cell wall of kiwifruit pollen tubes is a target for chromium toxicity: alterations to morphology, callose pattern and arabinogalactan protein distribution

Trivalent chromium has previously been found to effectively inhibit kiwifruit pollen tube emergence and elongation in vitro. In the present study, a photometric measure of increases in tube wall production during germination showed that 25 and 50 mum CrCl(3) treatment induced a substantial reduction in levels of polysaccharides in walls over those in controls. Moreover, chromium-treated kiwifruit pollen tubes had irregular and indented cell walls. Callose, the major tube wall polysaccharide, was deposited in an anomalous punctuate pattern. Arabinogalactan proteins (AGPs), which are integral in maintaining correct tube growth and shape in kiwifruit pollen, were found to be strongly altered in their distribution after CrCl(3) treatment compared to control tube walls. Transmission electron microscopy-immunogold analysis using four monoclonal antibodies (JIM8, JIM13, JIM14 and MAC207) revealed discontinuous AGP distribution within the treated tube walls. Such clearly discernable alterations in the molecular and morphological architecture of pollen tube walls may be detrimental in vivo for the male gametophyte to accomplish its vital role in the fertilisation process.

In vitro toxicity of silver nanoparticles to kiwifruit pollen exhibits peculiar traits beyond the cause of silver ion release

The vast use of silver nanoparticles (AgNPs) mandates thorough investigation of their impact on biosystems at various levels. The cytotoxicity of PVP coated-AgNPs to pollen, the aploid male gametophyte of higher plants, has been assessed here for the first time. The negative effects of AgNPs include substantial decreases in pollen viability and performance, specific ultrastructural alterations, early changes in calcium content, and unbalance of redox status. Ag⁺ released from AgNPs damaged pollen membranes and inhibited germination to a greater extent than the AgNPs themselves. By contrast, the AgNPs were more potent at disrupting the tube elongation process. ROS deficiency and overproduction were registered in the Ag⁺- and AgNP-treatment, respectively. The peculiar features of AgNP toxicity reflected their specific modes of interaction with pollen surface and membranes, and the dynamic exchange between coating (PVP) and culture medium. In contrast, the effects of Ag⁺ were most likely induced through chemical/physicochemical interactions.

Sortilin Is a Putative Postendocytic Receptor of Thyroglobulin

The Vps10p family member sortilin is involved in various cell processes, including protein trafficking. Here we found that sortilin is expressed in thyroid epithelial cells (thyrocytes) in a TSH-dependent manner, that the hormone precursor thyroglobulin (Tg) is a high-affinity sortilin ligand, and that binding to sortilin occurs after Tg endocytosis, resulting in Tg recycling. Sortilin was found to be expressed intracellularly in thyrocytes, as observed in mouse, human, and rat thyroid as well as in FRTL-5 cells. Sortilin expression was demonstrated to be TSH dependent, both in FRTL-5 cells and in mice treated with methimazole and perchlorate. Plasmon resonance binding assays showed that Tg binds to sortilin in a concentration-dependent manner and with high affinity, with Kd values that paralleled the hormone content of Tg. In addition, we found that Tg and sortilin interact in vivo and in cultured cells, as observed by immunoprecipitation, in mouse thyroid extracts and in COS-7 cells transiently cotransfected with sortilin and Tg. After incubation of FRTL-5 cells with exogenous, labeled Tg, sortilin and Tg interacted intracellularly, presumably within the endocytic pathway, as observed by immunofluorescence and immunoelectron microscopy, the latter technique showing some degree of Tg recycling. This was confirmed in FRTL-5 cells in which Tg recycling was reduced by silencing of the sortilin gene and in CHO cells transfected with sortilin in which recycling was increased. Our findings provide a novel pathway of Tg trafficking and a novel function of sortilin in the thyroid gland, the functional impact of which remains to be established.

Fine structure of the chorion and micropyle of the sea bass egg Dicentrarchus labrax (Teleostea, Percichthydae)

Abstract The fine structure of the chorion and micropyle of mature unfertilized eggs of the sea bass Dicentrarchus labrax was examined by scanning and transmission electron microscopy. The chorion consists of three layers: a thin electrondense outer layer, a paracrystalline middle layer and a thick inner layer consisting of 12 electrondense filamentous lamellae alternating with interlamellar material of lower electrondensity. The two more external layers are crossed by pore canals opening on the egg surface. The funnel‐type micropyle is located at the animal pole of the egg. The walls of the micropylar canal have circular, rib‐like thickenings reflecting the stratified inner layer of the chorion. The results are discussed and compared with those in other Teleostea.

A morphological and biochemical analysis comparative study of the collagen products Biopad, Promogram, Puracol, and Colactive.

ABSTRACT The aim of this study was to compare the capacity of the collagen products Biopad (Euroresearch, Milano, Italy), Promogran (Systagenix Wound Management, Quincy, Massachusetts), Colactive (Smith & Nephew, St Petersburg, Florida), and Puracol (Medline Industries, Mundelein, Illinois) to interact with biological tissues and to start restoring the healing process. These results demonstrate how these products can interact differently with enzymes and cells that characterize the environment of a healing wound.

Structure and membrane interactions of chionodracine, a piscidin-like antimicrobial peptide from the icefish Chionodraco hamatus.

Chionodracine (Cnd) is a 22-residue peptide of the piscidin family expressed in the gills of the Chionodraco hamatus as protection from bacterial infections. Here, we report the effects of synthetic Cnd on both Psychrobacter sp. TAD1 and Escherichia coli bacteria, as well as membrane models. We found that Cnd perforates the inner and outer membranes of Psychrobacter sp. TAD1, making discrete pores that cause the cellular content to leak out. Membrane disruption studies using intrinsic and extrinsic fluorescence spectroscopy revealed that Cnd behaves similarly to other piscidins, with comparable membrane partition coefficients. Membrane accessibility assays and structural studies using NMR in detergent micelles show that Cnd adopts a canonical topology of antimicrobial helical peptides, with the hydrophobic face toward the lipid environment and the hydrophilic face toward the bulk solvent. The analysis of Cnd free energy of binding to vesicles with different lipid contents indicates a preference for charged phospholipids and a more marked binding to native E. coli extracts. Taken with previous studies on piscidin-like peptides, we conclude that Cnd first adsorbs to the membrane, and then forms pores together with membrane fragmentation. Since Cnd has only marginal hemolytic activity, it constitutes a good template for developing new antimicrobial agents.

Fine structure of the eggshell of Ommatissus binotatus Fieber (Homoptera, Auchenorrhyncha, Tropiduchidae)

Abstract The external morphology and fine structure of the eggshell of Ommatissus binotatus Fieber (Homoptera : Tropiduchidae) was investigated by light, scanning and transmission electron microscopy. The egg surface has 2 main regions: a specialized area and an unspecialized egg capsule. The specialized area is characterized by a large respiratory plate containing the operculum and a short respiratory horn. The latter consists of an external hollow tube and an internal coneshaped projection hosting a micropylar canal. The eggshell has 4 layers: the vitelline envelope, a wax layer, the chorion and an outer mucous layer. The chorion has inner, intermediate and outer parts. The functions of the different parts of the eggshell are discussed. Characters useful to define the eggs and the oviposition habit in the family Tropiduchidae were provided. The size and morphology of the egg, plate, respiratory horn and operculum are suggested as useful characters for ootaxonomic analysis.