Speed of signal transfer in the chloroplast accumulation response

Chloroplast photorelocation movement is important for plants to perform efficient photosynthesis. Phototropins were identified as blue-light receptors for chloroplast movement in Arabidopsis thaliana and in the fern Adiantum capillus-veneris, whereas neochrome functions as a dual red/blue light receptor in the latter. However, the signal transduction pathways involved in chloroplast movement remain to be clarified. To investigate the kinetic properties of signalling from these photoreceptors to the chloroplasts, we deduced the speed of signal transfer using Adiantum capillus-veneris gametophytes. When a region of dark-adapted gametophyte cells was subjected to microbeam irradiation, chloroplasts moved towards the irradiated area even in subsequent darkness. We therefore recorded the movement and calculated the speeds of signal transfer by time-lapse imaging. Movement speeds under red or blue light were similar, e.g., about 1.0 μm min⁻¹ in prothallial cells. However, speeds varied according to cell polarity in protonemal cells. The speed of signal transfer from the protonemal apex to the base was approximately 0.7 μm min⁻¹, but roughly 2.3 μm min⁻¹ in the opposite direction. The speed of signal transfer in Arabidopsis thaliana mesophyll cells was approximately 0.8 μm min⁻¹ by comparison. Surprisingly, chloroplasts located farthest away from the microbeam were found to move faster than those in close proximity to the site of irradiation both in Adiantum capillus-veneris and A. thaliana.     

53BP1 and MDC1 foci formation in HT-1080 cells for low- and high-LET microbeam irradiations

An improved assessment of the biological effects and related risks of low doses of ionizing radiation is currently an important issue in radiation biology. Irradiations using microbeams are particularly well suited for precise and localized dose depositions, whereas recombinant cell lines with fluorescent proteins allow the live observation of radiation-induced foci. Living cells of the fibrosarcoma cell line HT-1080 stably expressing 53BP1 or full-length reconstituted MDC1 fused to Green Fluorescent Protein (GFP) were irradiated with protons and α-particles of linear energy transfers (LETs) of 15 and 75 keV/μm, respectively. Using a microbeam, the irradiations were carried out in line patterns, which facilitated the discrimination between undefined background and radiation-induced foci. As expected, foci formation and respective kinetics from α-particle irradiations with a high LET of 75 keV/μm could be detected in a reliable manner by both fusion proteins, as reported previously. Colocalization of γ-H2AX foci confirmed the DSB nature of the detected foci. As a novel result, the application of protons with low LET of 15 keV/μm generated 53BP1- and MDC1-mediated foci of almost equal size and slightly different kinetics. This new data expands the capability of 53BP1 and wild-type MDC1 on visible foci formation in living cells after irradiation with low-LET particles. Furthermore, the kinetics in HT-1080 cells for α-particle irradiation show a delay of about 20 s for 53BP1 foci detection compared to wild-type MDC1, confirming the hierarchical assembly of both proteins. Preliminary data for proton irradiations are shown and also these indicate a delay for 53BP1 versus MDC1.     

A proof of principle experiment for microbeam radiation therapy at the Munich compact light source

Microbeam radiation therapy (MRT), a preclinical form of spatially fractionated radiotherapy, uses an array of microbeams of hard synchrotron X-ray radiation. Recently, compact synchrotron X-ray sources got more attention as they provide essential prerequisites for the translation of MRT into clinics while overcoming the limited access to synchrotron facilities. At the Munich compact light source (MuCLS), one of these novel compact X-ray facilities, a proof of principle experiment was conducted applying MRT to a xenograft tumor mouse model. First, subcutaneous tumors derived from the established squamous carcinoma cell line FaDu were irradiated at a conventional X-ray tube using broadbeam geometry to determine a suitable dose range for the tumor growth delay. For irradiations at the MuCLS, FaDu tumors were irradiated with broadbeam and microbeam irradiation at integral doses of either 3 Gy or 5 Gy and tumor growth delay was measured. Microbeams had a width of 50 µm and a center-to-center distance of 350 µm with peak doses of either 21 Gy or 35 Gy. A dose rate of up to 5 Gy/min was delivered to the tumor. Both doses and modalities delayed the tumor growth compared to a sham-irradiated tumor. The irradiated area and microbeam pattern were verified by staining of the DNA double-strand break marker γH2AX. This study demonstrates for the first time that MRT can be successfully performed in vivo at compact inverse Compton sources.     

Microchambers and video-enhanced light microscopy for monitoring cellular events in living hyphae of arbuscular mycorrhizal fungi

Mycelial elongation and protoplasmic flow rate in vitro were monitored for germinated spores of Gigaspora rosea and Glomus caledonium respectively, growing on membranes in microchambers, by using a combination of time-lapse and video-enhanced light microscopy and image analysis. The microchambers allowed continuous observation of living mycelium over a period of several hours during which protoplasm flow and bidirectional movements of cellular organelles and particles were monitored in individual hyphae. Growth rate of G. rosea hyphae, calculated 8 days after germination, was 2.64 μm/min. Protoplasmic flow rate, measured on the basis of the movement of particles, ranged from 2.98 to 4.27 μm/s in living hyphae of G. caledonium. We showed that G. rosea, when growing in axenic culture in the absence of the host, ceased growth within 8 days of germination and underwent a process of protoplasm retraction from hyphal tips, leading to the formation of empty mycelial segments. A process of resource reallocation was inferred in spores of G. rosea showing multiple germination. Detailed developmental studies of living hyphae by using microchambers could provide useful information on spatio-temporal dimensions of cellular events occurring in arbuscular mycorrhizal fungi. This revised version was published online in June 2006 with corrections to the Cover Date.     

The role of nonhomologous end joining and homologous recombination in the clonogenic bystander effects of mammalian cells after exposure to counted 10 MeV protons and 4.5 MeV α-particles of the PTB microbeam

We have studied the dependence of clonogenic bystander effects on defects in the pathways of DNA double-strand break (DSB) repair and on linear energy transfer (LET). The single-ion microbeam of the Physikalisch-Technische Bundesanstalt (PTB) was used to irradiate parental Chinese hamster ovary cells or derivatives deficient in nonhomologous end joining (NHEJ) or homologous recombination (HR) in the G1-phase of the cell cycle. Cell nuclei were targeted with 10 MeV protons (LET = 4.7 keV/μm) or 4.5 MeV α-particles (LET = 100 keV/μm). During exposure, the cells were confluent, allowing signal transfer through both gap junctions and diffusion. When all cell nuclei were targeted with 10 MeV protons, approximately exponential survival curves were obtained for all three cell lines. When only 10% of all cell nuclei were targeted, a significant bystander effect was observed for parental and HR-deficient cells, but not for NHEJ-deficient cells. For all three cell lines, the survival data after exposure of all cell nuclei to 4.5 MeV α-particles could be fitted by exponential curves. When only 10% of all cell nuclei were targeted, significant bystander effects were obtained for parental and HR-deficient cells, whereas for NHEJ-deficient cells a small, but significant, bystander effect was observed only at higher doses. The data suggest that bystander cell killing is a consequence of un- or misrejoined DSB which occur in bystander cells during the S-phase as a result of the processing of oxidative bistranded DNA lesions. The relative contributions of NHEJ and HR to the repairing of DSB in the late S/G2-phase may affect clonogenic bystander effects.     

Low-dose studies of bystander cell killing with targeted soft X rays

The Gray Cancer Institute ultrasoft X-ray microprobe was used to quantify the bystander response of individual V79 cells exposed to a focused carbon K-shell (278 eV) X-ray beam. The ultrasoft X-ray microprobe is designed to precisely assess the biological response of individual cells irradiated in vitro with a very fine beam of low-energy photons. Characteristic CK X rays are generated by a focused beam of 10 keV electrons striking a graphite target. Circular diffraction gratings (i.e. zone plates) are then employed to focus the X-ray beam into a spot with a radius of 0.25 microm at the sample position. Using this microbeam technology, the correlation between the irradiated cells and their nonirradiated neighbors can be examined critically. The survival response of V79 cells irradiated with a CK X-ray beam was measured in the 0-2-Gy dose range. The response when all cells were irradiated was compared to that obtained when only a single cell was exposed. The cell survival data exhibit a linear-quadratic response when all cells were targeted (with evidence for hypersensitivity at low doses). When only a single cell was targeted within the population, 10% cell killing was measured. In contrast to the binary bystander behavior reported by many other investigations, the effect detected was initially dependent on dose (<200 mGy) and then reached a plateau (>200 mGy). In the low-dose region (<200 mGy), the response after irradiation of a single cell was not significantly different from that when all cells were exposed to radiation. Damaged cells were distributed uniformly over the area of the dish scanned (approximately 25 mm2). However, critical analysis of the distance of the damaged, unirradiated cells from other damaged cells revealed the presence of clusters of damaged cells produced under bystander conditions.     

Assessing the in vitro toxicity of the lunar dust environment using respiratory cells exposed to Al<Subscript>2</Subscript>O<Subscript>3</Subscript> or SiO<Subscript>2</Subscript> fine dust particles

Prior chemical and physical analysis of lunar soil suggests a composition of dust particles that may contribute to the development of acute and chronic respiratory disorders. In this study, fine Al₂O₃ (0.7 μm) and fine SiO₂ (mean 1.6 μm) were used to assess the cellular uptake and cellular toxicity of lunar dust particle analogs. Respiratory cells, murine alveolar macrophages (RAW 264.7) and human type II epithelial (A549), were cultured as the in vitro model system. The phagocytic activity of both cell types using ultrafine (0.1 μm) and fine (0.5 μm) fluorescent polystyrene beads was determined. Following a 6-h exposure, RAW 264.7 cells had extended pseudopods with beads localized in the cytoplasmic region of cells. After 24 h, the macrophage cells were rounded and clumped and lacked pseudopods, which suggest impairment of phagocytosis. A549 cells did not contain beads, and after 24 h, the majority of the beads appeared to primarily coat the surface of the cells. Next, we investigated the cellular response to fine SiO₂ and Al₂O₃ (up to 5 mg/ml). RAW 264.7 cells exposed to 1.0 mg/ml of fine SiO₂ for 6 h demonstrated pseudopods, cellular damage, apoptosis, and necrosis. A549 cells showed slight toxicity when exposed to fine SiO₂ for the same time and dose. A549 cells had particles clustered on the surface of the cells. Only a higher dose (5.0 mg/ml) of fine SiO₂ resulted in a significant cytotoxicity to A549 cells. Most importantly, both cell types showed minimal cytotoxicity following exposure to fine Al₂O₃. Overall, this study suggests differential cellular toxicity associated with exposure to fine mineral dust particles.     

Effects of microbeam light on growth and phototropism ofPilobolus crystallinus sporangiophores

A small blue-light beam (50 μm in diam) was used to examine light-growth response and phototropism inPilobolus crystallinus sporangiophores. Continuous irradiation by microbeam of a region 100–150 μm from the apex promoted the growth of a dark-adapted sporangiophore for about 15 min after a lag period of 1–2 min. After the promotion, the growth rate fell below that before the irradiation. Irradiation of the apex of sporangiophore slightly promoted the growth but strongly inhibited the growth after the promotion. A smaller light beam (10 μm in diam) applied continuously at grazing incidence along one side of the sporangiophore caused bending toward the shaded side, implying that the irradiated side grew more rapidly than the shaded side and that the lens effect is involved in the phototropism of young sporangiophores ofP. crystallinus. The involvement of the lens effect was confirmed by the fact that a carotenoid-less mutant was 1.5–2 times more sensitive to unilateral blue light than the wild type, probably because of a smaller intracellular light attenuation during passage through the mutant cell.     

Accurate detection of carcinoma cells by use of a cell microarray chip

Background

Accurate detection and analysis of circulating tumor cells plays an important role in the diagnosis and treatment of metastatic cancer treatment.

Methods and Findings

A cell microarray chip was used to detect spiked carcinoma cells among leukocytes. The chip, with 20,944 microchambers (105 µm width and 50 µm depth), was made from polystyrene; and the formation of monolayers of leukocytes in the microchambers was observed. Cultured human T lymphoblastoid leukemia (CCRF-CEM) cells were used to examine the potential of the cell microarray chip for the detection of spiked carcinoma cells. A T lymphoblastoid leukemia suspension was dispersed on the chip surface, followed by 15 min standing to allow the leukocytes to settle down into the microchambers. Approximately 29 leukocytes were found in each microchamber when about 600,000 leukocytes in total were dispersed onto a cell microarray chip. Similarly, when leukocytes isolated from human whole blood were used, approximately 89 leukocytes entered each microchamber when about 1,800,000 leukocytes in total were placed onto the cell microarray chip. After washing the chip surface, PE-labeled anti-cytokeratin monoclonal antibody and APC-labeled anti-CD326 (EpCAM) monoclonal antibody solution were dispersed onto the chip surface and allowed to react for 15 min; and then a microarray scanner was employed to detect any fluorescence-positive cells within 20 min. In the experiments using spiked carcinoma cells (NCI-H1650, 0.01 to 0.0001%), accurate detection of carcinoma cells was achieved with PE-labeled anti-cytokeratin monoclonal antibody. Furthermore, verification of carcinoma cells in the microchambers was performed by double staining with the above monoclonal antibodies.

Conclusion

The potential application of the cell microarray chip for the detection of CTCs was shown, thus demonstrating accurate detection by double staining for cytokeratin and EpCAM at the single carcinoma cell level.     

Measurement of particle diameter of <Emphasis Type="Italic">Lactobacillus acidophilus</Emphasis> microcapsule by spray drying and analysis on its microstructure

Lactobacillus acidophilus, as a probiotic, is widely used in many functional food products. Microencapsulation not only increases the survival rate of L. acidophilus during storage and extends the shelf-life of its products, but also optimal size microcapsule makes L. acidophilus have an excellent dispersability in final products. In this paper, L. acidophilus was microencapsulated using spray drying (inlet air temperature of 170°C; outlet air temperature of 85–90°C). The wall materials used in this study were β-cyclodextrin and acacia gum in the proportion of 9:1 (w/w), and microcapsules were prepared at four levels of wall materials (15, 20, 25 and 30% [w/v]) with a core material concentration of 6% (v/v). The microcapsule diameters were measured by Malvern’s Mastersizer-2000 particle size analyzer. The results showed that the particle diameters of microcapsule were mostly within 6.607 μm and 60.256 μm and varied with 2.884–120.226 μm (the standard smaller microcapsule designated as <350 μm). Through comparison of microcapsule size and uniformity with different concentration of wall materials, we concluded that the optimal concentration of wall material was 20% (w/v), which gave microcapsule with a relatively uniform size (averaging 22.153 μm), and the number of surviving encapsulated L. acidophilus was 1.50 × 10⁹ c.f.u./ml. After 8 weeks storage at 4°C, the live bacterial number was above 10⁷ c.f.u./ml, compared with unencapsulated L. acidophilus, 10⁴–10⁵ c.f.u./ml. Through the observation of scanning electron microscopy, we found that the shapes of microcapsule were round and oval, and L. acidophilus cells located in the centre of microcapsule.     

Chromosome aberration measurements in mitotic and G<Subscript>2</Subscript>-PCC lymphocytes at the standard sampling time of 48 h underestimate the effectiveness of high-LET particles

The relationship between heavy-ion-induced cell cycle delay and the time-course of aberrations in first-cycle metaphases or prematurely condensed G₂-cells (G₂-PCC) was investigated. Lymphocytes of the same donor were irradiated with X-rays or various charged particles (carbon, iron, xenon, and chromium) covering an LET range of 2–3,160 keV/μm. Chromosome aberrations were measured in samples collected at 48, 60, 72, and 84 h postirradiation. Linear-quadratic functions were fitted to the data, and the fit parameters α and β were determined. At any sampling time, α values derived from G₂-cells were higher than those from metaphases. The α value derived from metaphase analysis at 48 h increased with LET, reached a maximum around 155 keV/μm, and decreased with a further rise in LET. At the later time-points, higher α values were estimated for particles with LET > 30 keV/μm. Estimates of α values from G₂-cells showed a similar LET dependence, yet the time-dependent increase was less pronounced. Altogether, our data demonstrate that heavily damaged lymphocytes suffer a prolonged G₂-arrest that is clearly LET dependent. For this very reason, the standard analysis of aberrations in metaphase cells 48 h postirradiation will considerably underestimate the effectiveness of high-LET radiation. Scoring of aberrations in G₂-PCC at 48 h as suggested by several authors will result in higher aberration yields. However, when particles with a very high-LET value (LET > 150 keV/μm) are applied, still a fraction of multiple damaged cells escape detection by G₂-analysis 48 h postirradiation.     

Induction of a bystander chromosomal damage of He-ion microbeams in mammalian cells.

We report here a bystander effect in chromosomal damage using He-ion microbeam. Human-hamster hybrid cells were irradiated with a precision He-ion microbeam generated by the Columbia microbeam system. When 20% of the cells were exposed to single He ion, the incidence of cells with chromatid-type breaks detected with the PCC technique was covered wide range from 0 to 6 breaks per cell. In contrast, the distribution showed a mixed two-peak pattern, such as non-exposed and all-cell exposed patterns, under the condition of assuming no bystander effect by treating with an effective inhibitor of cell-cell communication. These findings provide clear evidence that single He-ion irradiated cells can induce bystander chromosomal alterations in neighboring cells not directly hit by He ion.     

DNA Damage and Decreased DNA Repair in Peripheral Blood Mononuclear Cells in Individuals Exposed to Arsenic and Lead in a Mining Site

The aim of this study was to evaluate DNA damage and the capacity for DNA repair in children exposed to arsenic and lead. During 2006, we studied a total of 85 healthy children (aged 4–11 years) who were residents of Villa de la Paz (community A), Matehuala (community B), and Soledad de Graciano Sanchez (community C) in San Luis Potosi, Mexico. The quantification of arsenic in urine (AsU) and lead in blood (PbB) was performed by atomic absorption spectrophotometry. The alkaline comet assay was used to evaluate DNA damage and DNA repair. The highest levels of AsU and PbB in children were found in community A (44.5 μg/g creatinine for arsenic and 11.4 μg/dL for lead), followed by community B (16.8 μg/g creatinine for arsenic and 7.3 μg/dL for lead) and finally by children living in community C (12.8 μg/g creatinine for arsenic and 5.3 μg/dL for lead). When DNA damage was assessed, children living in community A had the highest DNA damage. Analysis of these same cells 1 h after a challenge with H₂O₂ 10 μM showed a dramatic increase in DNA damage in the cells of children living in community B and community C, but not in the cells of children living in community A. Moreover, significantly higher levels of DNA damage were observed 3 h after the challenge ended (repair period) in cells from individuals living in community A. Our results show that children exposed to metals might be more susceptible to DNA alterations.     

Antibacterial activity and mechanism of silver nanoparticles on Escherichia coli

The antibacterial activity and acting mechanism of silver nanoparticles (SNPs) on Escherichia coli ATCC 8739 were investigated in this study by analyzing the growth, permeability, and morphology of the bacterial cells following treatment with SNPs. The experimental results indicated 10 μg/ml SNPs could completely inhibit the growth of 10⁷ cfu/ml E. coli cells in liquid Mueller–Hinton medium. Meanwhile, SNPs resulted in the leakage of reducing sugars and proteins and induced the respiratory chain dehydrogenases into inactive state, suggesting that SNPs were able to destroy the permeability of the bacterial membranes. When the cells of E. coli were exposed to 50 μg/ml SNPs, many pits and gaps were observed in bacterial cells by transmission electron microscopy and scanning electron microscopy, and the cell membrane was fragmentary, indicating the bacterial cells were damaged severely. After being exposed to 10 μg/ml SNPs, the membrane vesicles were dissolved and dispersed, and their membrane components became disorganized and scattered from their original ordered and close arrangement based on TEM observation. In conclusion, the combined results suggested that SNPs may damage the structure of bacterial cell membrane and depress the activity of some membranous enzymes, which cause E. coli bacteria to die eventually.     

Improved normal tissue protection by proton and X-ray microchannels compared to homogeneous field irradiation

The risk of developing normal tissue injuries often limits the radiation dose that can be applied to the tumour in radiation therapy. Microbeam Radiation Therapy (MRT), a spatially fractionated photon radiotherapy is currently tested at the European Synchrotron Radiation Facility (ESRF) to improve normal tissue protection. MRT utilizes an array of microscopically thin and nearly parallel X-ray beams that are generated by a synchrotron. At the ion microprobe SNAKE in Munich focused proton microbeams (“proton microchannels”) are studied to improve normal tissue protection. Here, we comparatively investigate microbeam/microchannel irradiations with sub-millimetre X-ray versus proton beams to minimize the risk of normal tissue damage in a human skin model, in vitro. Skin tissues were irradiated with a mean dose of 2 Gy over the irradiated area either with parallel synchrotron-generated X-ray beams at the ESRF or with 20 MeV protons at SNAKE using four different irradiation modes: homogeneous field, parallel lines and microchannel applications using two different channel sizes. Normal tissue viability as determined in an MTT test was significantly higher after proton or X-ray microchannel irradiation compared to a homogeneous field irradiation. In line with these findings genetic damage, as determined by the measurement of micronuclei in keratinocytes, was significantly reduced after proton or X-ray microchannel compared to a homogeneous field irradiation. Our data show that skin irradiation using either X-ray or proton microchannels maintain a higher cell viability and DNA integrity compared to a homogeneous irradiation, and thus might improve normal tissue protection after radiation therapy.     

Microbeam X-ray Diffraction Study of Granular Crystals Formed in Water-in-Oil Emulsion

Margarine and fat spread contain typical water-in-oil emulsions, including semi-solid fats, as continuous oil phases. The application of palm oil, one of the most promising trans-fat alternatives, for semi-solid fats is increasing. However, granular crystals often occur in palm-oil-based solid fats and cause deterioration. In this study, we carried out differential scanning calorimetry (DSC), optical microscopy, and X-ray diffraction (XRD) experiments on granular crystals in margarine. A conventional laboratory-scale technique was applied to examine thermal properties of high-melting fat fractions in granular crystals. In addition, microstructures of the granular crystal were precisely observed with a synchrotron radiation small-angle XRD (SR-μ-SAXD) technique by using a microbeam having a cross section of 5 × 5 μm². The following results were obtained. (1) DSC indicated that granular crystals are composed of high-melting fractions having a melting temperature of 23 °C. (2) Conventional XRD of granular crystals indicated that β-fat crystals of a triple chain length structure (β-3) melted below the melting of β′-fat of a double chain length structure (β′-2). (3) SR-μ-SAXD indicated that the fat crystals in normal margarine were β′-2. However, the fat crystals of the double and triple chain length structures were simultaneously observed at the center region of a granular crystal, whereas only the fat crystals of β-3 were observed at the outer region of a granular crystal. We analyzed the microstructures and formation processes of granular crystals in relation to the fractional crystallization of the β form of 1,3-dipalmitoyl-2-oleoyl-sn-glycerol promoted by crystallization and transformation of tripalmitin and tristearin fractions.     

The importance and distinctiveness of small-sized phytoplankton in the Magellan Straits

The distribution of summer phytoplankton across the Straits of Magellan (SOM) was studied with the aims of tracing differences among the distinct subregions of the area and contributing to the knowledge of its biodiversity. Samples collected at 25 stations were observed and counted in light microscopy. Selected samples were observed with transmission electron microscopy. The main unifying feature of the phytoplankton in the SOM was the high abundance and numerical dominance of small-sized (<10 μm) eukaryotic species, among which coccoid cells of <3 μm size were predominant (56.2 ± 30.6 of the total phytoplankton abundance). They mostly belonged to the prasinophyte Pycnococcus provasolii, which was abundant (0.8–6,834 cells × 10³ ml⁻¹) at all stations with the exception of those in proximity to the Atlantic entrances, where it was not recorded. Small-sized (<3 and 3–5 μm) diatoms (Minidiscus trioculatus, Lennoxia faveolata and other undetermined centric species) attained high densities (<3,757 cells 10³ ml⁻¹) especially at stations of the Patagonian sectors, whereas microplanktonic diatoms were only found at the two entrances of the Straits. Dinoflagellates were constituted mainly by >10 μm forms in the Andean subregion and <10 μm naked species in the Patagonian subregion, contributing up to 75.9 and 41.8% of the total carbon in these two areas, respectively. In the Patagonian subregion, flagellates mainly constituted by <5 μm forms and by cryptomonads <10 μm comprised up to 53.9% of the total biomass. Several species identified in this study have never been reported in other investigations in the SOM, while others, including Pycnococcus provasolii and Lennoxia faveolata, have rarely been recorded elsewhere. Overall, the summer phytoplankton of the Straits does not resemble that of any other region of the world’s seas. Although some of the predominant species might have been overlooked elsewhere, their abundance and relative importance apparently constitute a distinctive feature of the SOM.     

Distribution and morphology of the ampullary organs of the estuarine long-tailed catfish, <Emphasis Type="Italic">Euristhmus lepturus</Emphasis> (Plotosidae,Siluriformes)

Ampullary organs of Euristhmus lepturus occur in high densities along the head and in four parallel pathways along the trunk of the body. Large ampullary pores (125–130 μm) are easily distinguishable from other sensory epithelial pores due to the differences in size and the presence of a collar-like structure. Simple, singular ampullary organs of the head region consist of an ampullary pore connected to a long canal with a diameter of 115–175 μm before terminating as a simple ampulla with an external diameter of 390–480 μm. The ampullary canal is composed of 1–2 layers of flattened squamous epithelial cells, the basement membrane and an interlocking collagen sheath. The innermost cells lining the canal wall are adjoined via tight junctions and numerous desmosomes, as are those of the receptor and supportive cells. Canal wall tissue gives rise to a sensory epithelium containing between 242 and 285 total receptor cells, with an average diameter of 11.7 ± 5.3 μm, intermixed with medially nucleated supportive cells. Each receptor cell (21.38 ± 4.41 μm, height) has an apically positioned nucleus and a luminal surface covered with numerous microvilli. Neural terminals abut the basal region of receptor cells opposite multiple presynaptic bodies and dense mitochondria. Supportive cells extend from the ampullary lumen to the basement membrane, which is adjacent to the complex system of collagen fibres.     

Effect of green tea powder (<Emphasis Type="Italic">Camellia sinensis</Emphasis> L. cv. Benifuuki) particle size on <Emphasis Type="Italic">O</Emphasis>-methylated EGCG absorption in rats; The Kakegawa Study

Tea polyphenols, e.g., (-)-epigallocatechin-3-O-(3-O-methyl gallate (EGCG3”Me), (-)-epigallocatechin-3-O-gallate (EGCG), (-)-epigallocatechin (EGC), (-)-epicatechin-3-O-gallate (ECG), and (-)-epicatechin (EC), are believed to be responsible for the beneficial effects of tea. ‘Benifuuki’, a tea (Camellia sinensis L.) cultivar grown in Japan, is rich in the anti-allergic molecule epigallocatechin-3-O-(3-O-methyl) gallate (EGCG3”Me). Pulverized Benifuuki green tea powder (BGP) is more widely distributed than leaf tea in Japan. Japanese people mix their pulverized tea with water directly, whereas it is common to drink leaf tea after extraction. However, few studies of the effects of BGP particle size on polyphenol bioavailability have been performed. This study was conducted to investigate the absorption of catechins in rats after the intragastric administration of Benifuuki green tea. Therefore, we assessed the plasma concentrations of catechins following the ingestion of BGP with different mean particle sizes (2.86, 18.6, and 76.1 μm) or Benifuuki green tea infusion (BGI) as a control in rats. The bioavailabilities of EGCG3”Me, EGCG, ECG, EGC, and EC were analyzed after the oral administration of a single dose of Benifuuki green tea (125 mg/rat) to rats. The plasma concentrations of tea catechins were determined by HPLC analysis combined with of electrochemical detection (ECD) using a coulometric array. The AUC (area under the drug concentration versus time curve; min μg/mL) of ester-type catechins (EGCG3”Me, EGCG, and ECG) for the BGP 2.86 μm were significantly higher than those in the infusion and 18.6 and 76.1 μm BGP groups, but the AUC of free-type catechins (EGC and EC) showed no differences between these groups. Regarding the peak plasma level of EGCG3”Me adjusted for intake, BGP 2.86 μm and BGI showed higher values than the BGP 18.6 and 76.1 μm groups, and the peak plasma levels of the other catechins displayed the same tendency. The present study demonstrates that the bioavailability of ester-type catechins (EGCG and ECG) can be improved by reducing the particle size of green tea, but the plasma level of EGCG3”Me in the BGI group was similar to that in the BGP 2.86 μm group. This result suggests that drinking Benifuuki green tea with a particle size of around 2 μm would deliver the anti-allergic EGCG3”Me and the anti-oxidant EGCG efficiently.     

Pigmented Nanoflagellates Grazing on <Emphasis Type="Italic">Synechococcus</Emphasis>: Seasonal Variations and Effect of Flagellate Size in the Coastal Ecosystem of Subtropical Western Pacific

We investigated seasonal variation of grazing impact of the pigmented nanoflagellates (PNF) with different sizes upon Synechococcus in the subtropical western Pacific coastal waters using grazing experiments with fluorescently labeled Synechococcus (FLS). For total PNF, conspicuous seasonal variations of ingestion rates on Synechococcus were found, and a functional response was observed. To further investigate the impact of different size groups, we separated the PNF into four categories (<3, 3–5, 5–10, and >10 μm). Our results indicated that the smallest PNF (<3 μm PNF) did not ingest FLS and was considered autotrophic. PNF of 3–5 μm in size made up most of the PNF community; however, their ingestion on Synechococcus was too low (0.1–1.9 Syn PNF⁻¹ h⁻¹) to support their growth, and they had to depend on other prey or photosynthesis to survive. The ingestion rate of the 3–5 μm group exhibited no significant seasonal variation; by contrast, the ingestion rates of 5–10 and >10 μm PNFs showed significant seasonal variation. During the warm season, 3–5 μm PNF were responsible for the grazing of 12% of Synechococcus production, 5–10 μm PNF for 48%, and >10 μm PNF for 2%. Taken together, our results demonstrate that the PNF of 3–10 μm consumed most Synechococcus during the warm season and exhibited a significant functional response to the increase in prey concentration.