CHANGES IN CELL COMPOSITION AND LIPID METABOLISM MEDIATED BY SODIUM AND NITROGEN AVAILABILITY IN THE MARINE DIATOM PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE)1

The effects of nitrogen starvation in the presence or absence of sodium in the culture medium were monitored in batch cultures of the marine diatom Phaeodactylum tricornutum Bohlin. During nitrogen starvation in the presence of sodium, cell nitrogen and chlorophyll a decreased, mainly as a consequence of continued cell division. These decreases were accompanied by decreases in the rates of photosynthesis and respiration. There was no change in either cell volume or carbohydrate, but both carbon and lipid increased. During nitrogen starvation in the absence of sodium, cell division ceased. Cell nitrogen and chlorophyll a remained constant, and respiration did not decrease, but the changes in the photosynthetic rate and the lipid content per cell were similar to cultures that were nitrogen-starved in the presence of sodium. The carbon-to-nitrogen ratio increased in both cultures. Nitrogen, in the form of nitrate, and sodium were resupplied to cultures that had been preconditioned in nitrogen- and sodium-deficient medium for 5 d. Control cultures to which neither nitrate or sodium were added remained in a static state with respect to cell number, volume, and carbohydrate but showed slight increases in lipid. Cells in cultures to which 10 mM nitrate alone was added showed a similar response to cultures where no additions were made. Cells in cultures to which 50 mM sodium alone was added divided for 2 d, with concomitant small decreases in all measured constituents. Cell division resumed in cultures to which both sodium and nitrate were added. The lipid content fell dramatically in these cells and was correlated to metabolic oxidation via measured increases in the activity of the glyoxylate cycle enzyme, isocitrate lyase. We conclude that lipids are stored as a function of decreased growth rate and are metabolized to a small extent when cell division resumes. However, much higher rates of metabolism occur if cell division resumes in the presence of a nitrogen source.     

Secretory function of autophagy in innate immune cells

Eukaryotic cells utilize two main secretory pathways to transport proteins to the extracellular space. Proteins with a leader signal sequence often undergo co‐translational transport into the endoplasmic reticulum (ER), and then to the Golgi apparatus before they reach their destination. This pathway is called the conventional secretory pathway. Proteins without signal peptides can bypass this ER‐Golgi system and are secreted by a variety of mechanisms collectively called the unconventional secretory pathway. The molecular mechanisms of unconventional secretion are emerging. Autophagy is a conserved bulk degradation mechanism that regulates many intracellular functions. Recent evidence implicates autophagy in the secretory pathway. This review focuses on potential secretory roles of autophagy and how they could modulate the functions of innate immune cells that secrete a wide range of mediators in response to environmental and biological stimuli. We provide a brief overview of the secretory pathways, enumerate the potential mechanistic themes by which autophagy interacts with these pathways and describe their relevance in the context of innate immune cell function.     

Quantitative computed tomography-based finite element models of the human lumbar vertebral body: effect of element size on stiffness,damage, and fracture strength predictions

This study investigated the numerical convergence characteristics of specimen-specific "voxel-based" finite element models of 14 excised human cadaveric lumbar vertebral bodies (age: 37-87; M = 6, F = 8) that were generated automatically from clinical-type CT scans. With eventual clinical applications in mind, the ability of the model stiffness to predict the experimentally measured compressive fracture strength of the vertebral bodies was also assessed. The stiffness of "low"-resolution models (3 x 3 x 3 mm element size) was on average only 4% greater (p = 0.03) than for "high"-resolution models (1 x 1 x 1.5 mm) despite interspecimen variations that varied over four-fold. Damage predictions using low- vs high-resolution models were significantly different (p = 0.01) at loads corresponding to an overall strain of 0.5%. Both the high (r2 = 0.94) and low (r2 = 0.92) resolution model stiffness values were highly correlated with the experimentally measured ultimate strength values. Because vertebral stiffness variations in the population are much greater than those that arise from differences in voxel size, these results indicate that imaging resolution is not critical in cross-sectional studies of this parameter. However, longitudinal studies that seek to track more subtle changes in stiffness over time should account for the small but highly significant effects of voxel size. These results also demonstrate that an automated voxel-based finite element modeling technique may provide an excellent noninvasive assessment of vertebral strength.     

Adipose Vascular Endothelial Growth Factor Regulates Metabolic Homeostasis through Angiogenesis

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Highlights? Two-way modulations of adipose VEGF were generated with aP2-Cre transgene ? Adipose VEGF KO reduces vasculature, increases hypoxia and inflammation in fat ? Adipose VEGF KO accelerates the development of metabolic disease in high-fat diet ? Induced adipose VEGF has opposite effect on fat and restores metabolic homeostasis     

The last dicynodont: an Australian Cretaceous relict

Some long-forgotten fossil evidence reveals that a dicynodont (mammal-like reptile of the infraorder Dicynodontia) inhabited Australia as recently as the Early Cretaceous, ca. 110 Myr after the supposed extinction of dicynodonts in the Late Triassic. This remarkably late occurrence more than doubles the known duration of dicynodont history (from ca. 63 Myr to ca. 170 Myr) and betrays the profound impact of geographical isolation on Australian terrestrial faunas through the Mesozoic. Australia's late-surviving dicynodont may be envisaged as a counterpart of the ceratopians (horned dinosaurs) in Cretaceous tetrapod faunas of Asia and North America.     

Development and validation of an HPLC method for the determination of dibenzoylmethane in rat plasma and its application to the pharmacokinetic study

A highly sensitive and simple high-performance liquid chromatographic (HPLC) assay has been developed and validated for the quantification of dibenzoylmethane (DBM) in rat plasma. DBM and internal standard (I.S.) 1-(5-chloro-2-hydroxy-4-methylphenyl)-3-phenyl-1,3-propanedione (CHMPP) were extracted from rat plasma by ethyl acetate/methanol (95:5, v/v) and analyzed using reverse-phase gradient elution with a Phenomenex Gemini C18 5-mum column. A gradient of mobile phase (mobile phase A: water/methanol (80:20, v/v) with 0.1% TFA and mobile phase B: acetonitrile with 0.1% TFA) at a flow rate of 0.2 mL/min, and ultraviolet (UV) detection at 335 nm were utilized. The lower limit of quantification (LLOQ) using 50 microL rat plasma was 0.05 microg/mL. The calibration curve was linear over a concentration range of 0.05-20 microg/mL. The mean recoveries were 80.6+/-5.7, 83.4+/-1.6 and 77.1+/-3.4% with quality control (QC) level of 0.05, 1 and 20 microg/mL, respectively. Intra- and inter-day assay accuracy and precision fulfilled US FDA guidance for industry bioanalytical method validation. Stability studies showed that DBM was stable in rat plasma after 4h incubation at room temperature, one month storage at -80 degrees C and three freeze/thaw cycles, as well as in reconstitute buffer for 48 h at 4 degrees C. The utility of the assay was confirmed by the successful analysis of plasma samples from DBM pharmacokinetics studies in the rats after oral and intravenous administrations.     

RhoA and RhoC have distinct roles in migration and invasion by acting through different targets

Several studies suggest that RhoA and RhoC, despite their sequence similarity, have different roles in cell migration and invasion, but the molecular basis for this is not known. Using RNAi, we show that RhoA-depleted cells became elongated and extended multiple Rac1-driven narrow protrusions in 2D and 3D environments, leading to increased invasion. These phenotypes were caused by combined but distinct effects of the Rho-regulated kinases ROCK1 and ROCK2. Depletion of ROCK2 induced multiple delocalized protrusions and reduced migratory polarity, whereas ROCK1 depletion selectively led to cell elongation and defective tail retraction. In contrast, RhoC depletion increased cell spreading and induced Rac1 activation around the periphery in broad lamellipodia, thereby inhibiting directed migration and invasion. These effects of RhoC depletion are mediated by the formin FMNL3, which we identify as a new target of RhoC but not RhoA. We propose that RhoA contributes to migratory cell polarity through ROCK2-mediated suppression of Rac1 activity in lamellipodia, whereas RhoC promotes polarized migration through FMNL3 by restricting lamellipodial broadening.     

Transgenic lines of melon (Cucumis melo L. var. makuwa cv. ‘Silver Light’) expressing antifungal protein and chitinase genes exhibit enhanced resistance to fungal pathogens

The oriental melon (Cucumis melo L. var. makuwa cv. ‘Silver Light’) is an important fruit crop in the tropical and subtropical regions. However, oriental melon production is severely decreased by fungal diseases. In this study, antifungal protein (AFP) and chitinase (CHI) fusion genes were introduced into oriental melons to control fungal diseases caused by Rhizoctonia solani and Fusarium oxysporum. Transformation of oriental melon (Cucumis melo L. var. makuwa cv. ‘Silver Light’) with Agrobacterium tumefaciens strain LBA4404 containing antifungal protein (AFP) and chitinase (CHI) fusion genes under the control of the cauliflower mosaic virus (CaMV) 35S promoter and neomycin phosphotransferase (nptII) gene as a selectable marker was performed. Cotyledon explants of oriental melon were inoculated by Agrobacterium suspensions with pBI121–AFP–CHI and cultured in a regeneration medium. After regeneration, genomic DNA polymerase chain reaction (PCR) was conducted to confirm the presence of putative transgenic shoots. Southern blot analysis confirmed that the AFP–CHI fusion gene was incorporated into the genomic DNA of the PCR-positive lines. RT-PCR analysis showed that the AFP–CHI fusion gene was expressed in the individual transgenic lines. Western blot analysis revealed the accumulation of CHI protein in leaves. A segregation analysis of the T₁ generation confirmed the inheritance of the transgene. Our results demonstrated that the AFP–CHI fusion gene was effective in protecting the transgenic melon plants against fungal disease caused by Rhizoctonia solani and Fusarium oxysporum.