Project description:BackgroundEach of the three individual components of the CMG complex (Cdc45, MCM and GINS) is essential for chromosomal DNA replication in eukaryotic cells, both for the initiation of replication at origins and also for normal replication fork progression. The MCM complex is a DNA helicase that most likely functions as the catalytic core of the replicative helicase, unwinding the parental duplex DNA ahead of the moving replication fork, whereas Cdc45 and the GINS complex are believed to act as accessory factors for MCM.ResultsTo investigate interactions between components of the CMG complex, we have used bimolecular fluorescence complementation (BiFC) in the fission yeast Schizosaccharomyces pombe for the first time, to analyse protein-protein interactions between GINS and MCM subunits expressed from their native chromosomal loci. We demonstrate interactions between GINS and MCM in the nuclei of exponentially-growing fission yeast cells and on chromatin in binucleate S-phase cells. In addition we present evidence of MCM-MCM interactions in diploid fission yeast cells. As with GINS-MCM interactions, MCM-MCM interactions also occur on chromatin in S-phase cells.ConclusionBimolecular fluorescence complementation can be used in fission yeast to visualise interactions between two of the three components of the CMG complex, offering the prospect that this technique could in the future be used to allow studies on replication protein dynamics in living S. pombe cells.
Project description:Cementum annuli widths in mammals are is influenced by the nutrition of mammals. Reproductive stress has been is suggested to reduce the width of lead to narrower cementum annuli widths in female Asian black bears (Ursus thibetanus); however, food availability in autumn strongly impacts bear nutrition and likely impacts cementum widths as well. This study aimed to test how cementum annuli widths and the formation of false annuli were influenced by hard mast production. We established two hypotheses: (1) cementum annuli widths become narrower in poor mast years owing to inadequate nutritional conditions and (2) false annuli occur more frequently in poor mast years. We used teeth samples from male bears to avoid reproductive influences and separated width data into "adult" and "subadult" groups. We calculated the proportional width index (PWI) and used linear mixed models to estimate the masting effects on PWI. Generalized linear mixed models estimated the masting effects on false annuli frequency. True annuli widths and false annuli formation showed no significant relationship with mast production in adults. In subadults, poor mast production weak negative influence on false annuli formation. These new data resolve previous questions, allowing us to deduce that cementum annuli widths are a reliable index of reproductive success in female bears.
Project description:Plant-derived natural bioactive molecules are of great therapeutic potential but, so far, their application in nanomedicine has scarcely been studied. This work aimed at comparing two methodologies, i.e., adsorption and in situ incorporation, to prepare hybrid polyphenol/hydroxyapatite nanoparticles. Two flavonoids, baicalin and its aglycone derivative baicalein, and two phenolic acids derived from caffeic acid, rosmarinic and chlorogenic acids, were studied. Adsorption of these polyphenols on pre-formed hydroxyapatite nanoparticles did not modify particle size or shape and loading was less than 10% (w/w). In contrast, presence of polyphenols during the synthesis of nanoparticles significantly impacted and sometimes fully inhibited hydroxyapatite formation but recovered particles could exhibit higher loadings. For most hybrid particles, release profiles consisted of a 24 h burst effect followed by a slow release over 2 weeks. Antioxidant properties of the polyphenols were preserved after adsorption but not when incorporated in situ. These results provide fruitful clues for the valorization of natural bioactive molecules in nanomedicine.
Project description:Spherical MCM-48 mesoporous sieve co-doped with vanadium and iron was successfully synthesized via one-step hydrothermal method. The material was characterized by X-ray diffraction (XRD), nitrogen adsorption-desorption isotherms, inductively coupled plasma (ICP), scanning electron microscopy (SEM), transmission electron microscopy (TEM), diffuse reflectance UV-vis spectra, and X-ray photoelectron spectra (XPS) techniques. Results indicated that the V-Fe-MCM-48 showed an ordered 3D cubic mesostructure with spherical morphology, narrow pore size distribution and high specific surface area. Most of vanadium and iron atoms existing as tetrahedral V4+ and Fe3+ species were co-doped into the silicate framework. The particle sizes of V-Fe-MCM-48 were smaller and the specific area was much higher than those of of V-MCM-48. Additionally, the synthesized V-Fe-MCM-48 exhibited improved hydrothermal stability compared with the pure MCM-48.
Project description:The chemistry between layered MWW zeolite and carbon black pearls (BP 2000) as an inexpensive hard template was investigated to develop a rational one-pot synthesis of MCM-22 microspheres. The characterization results showed that the insertion of BP 2000 in the gel synthesis did not substantially compromise the crystallinity and microporosity, and the microscopic analyses showed that BP 2000 played a key role in controlling the final morphology of the MCM-22 zeolite, creating beautiful dandelion-like microspherical particles. The morphology obtained is due to the tortuous shape of the hard template, the particular MWW particle crystals, the interaction with the external surface of the MWW zeolitic precursor, and the synthesis conditions. The stacking of MWW crystals with edge-to-face orientations generates meso-/macrovoids, allowing access to the interiors of the microspheres. The microspheres were homogeneous with sizes ranging from 6 to 8 μm with an increase of the external surface and a macroporous size distribution centered at 200 nm, which is two times that of the traditional MCM-22 zeolite.
Project description:Cell-based tissue engineering can be used to replace missing or damaged bone, but the optimal methods for delivering therapeutic cells to a bony defect have not yet been established. Using transgenic reporter cells as a donor source, two different collagen-hydroxyapatite (HA) scaffolds, and a critical-size calvarial defect model, we investigated the effect of a cell-attachment period prior to implantation, with or without an extracellular matrix-based seeding suspension, on cell engraftment and osteogenesis. When quantitatively compared, the in-house scaffold implanted immediately had a higher mean radiopacity than in-house scaffolds incubated overnight. Both scaffold types implanted immediately had significantly higher area fractions of donor cells, while the in-house collagen-HA scaffolds implanted immediately had higher area fractions of the mineralization label compared with groups incubated overnight. When the cell loading was compared in vitro for each delivery method using the in-house scaffold, immediate loading led to higher numbers of delivered cells. Immediate loading may be preferable in order to ensure robust bone formation in vivo. The use of a secondary ECM carrier improved the distribution of donor cells only when a pre-attachment period was applied. These results have improved our understanding of cell delivery to bony defects in the context of in vivo outcomes.
Project description:The ordered mesoporous silica MCM-48 with cubic Ia3d structure was synthesized using the cationic surfactant hexadecyltrimethylammonium bromide (CTAB) as a template agent and tetraethylorthosilicate (TEOS) as a silica source. The obtained material was first functionalized with (3-glycidyloxypropyl)trimethoxysilane (KH560); further, two types of amination reagents were used: ethylene diamine (N2) and diethylene triamine (N3). The modified amino-functionalized materials were characterized by powder X-ray diffraction (XRD) at low angles, infrared spectroscopy (FT-IR) and nitrogen adsorption-desorption experiments at 77 K. Characterization from a structural point of view reveals that the ordered MCM-48 mesoporous silica has a highly ordered structure and a large surface area (1466.059 m2/g) and pore volume (0.802 cm3/g). The amino-functionalized MCM-48 molecular sieves were tested for CO2 adsorption-desorption properties at different temperatures using thermal program desorption (TPD). Promising results for CO2 adsorption capacities were achieved for MCM-48 sil KH560-N3 at 30 °C. At 30 °C, the MCM-48 sil KH560-N3 sample has an adsorption capacity of 3.17 mmol CO2/g SiO2, and an efficiency of amino groups of 0.58 mmol CO2/mmolNH2. After nine adsorption-desorption cycles, the results suggest that the performance of the MCM-48 sil KH N2 and MCM-48 sil KH N3 adsorbents is relatively stable, presenting a low decrease in the adsorption capacity. The results reported in this paper for the investigated amino-functionalized molecular sieves as absorbents for CO2 can be considered as promising.
Project description:The elimination of iodide (I-) from water is a tough subject due to its low adsorption tendency and high mobility. In this work, MCM-41/Ag2O nanomaterials were prepared, characterized, and employed to adsorb I- from water. The Ag2O nanoparticles were dispersed homogeneously in the pores or at the surface of the MCM-41 support, and the Ag2O nanoparticles in the pores had small particles sizes due to the confinement of the mesoporous channel. The prepared MCM-41/Ag2O nanomaterials exhibited a higher specific surface area than previously reported Ag2O-based composites. The adsorption of I- by the nanomaterials was able to reach equilibrium at 180 min. The MCM-41/Ag2O nanomaterials showed a better adsorption capacity per unit mass of Ag2O than pure Ag2O nanoparticles and previously reported Ag2O-based composites prepared using other supports. Furthermore, the MCM-41/Ag2O nanomaterials exhibited high selectivity for I- in the presence of high concentrations of competitive anions, such as Cl- or Br-, and could function in a wide range of pH. The chemical reaction between Ag2O and I- and the surface adsorption were the main adsorption mechanisms. These results indicate that MCM-41/Ag2O nanomaterials are a promising and efficient adsorbent material suitable for the removal of I- for practical application.
Project description:In this work, poly (vinyl alcohol) (PVA) was employed to produce a Mesoporous Composition of Matter-48 Modified (MCM-48-M or MCM-48-PVA). After surface modification, MCM-48-M was used to produce nanocomposite (NC) films with polycaprolactone (PCL) as a matrix at room temperature. PCL and MCM-48 nanoparticles (NPs) were chosen due to their great biocompatibility and low toxicity. However, MCM-48-M is more compatible with PCL than MCM-48. NC films were sterilized by gamma radiation with a dose of 25 kGy and characterized by experimental techniques to investigate their chemical, mechanical (tensile) and thermal properties. Scanning electron microscopy (SEM) and transmission electronic microscopy (TEM) results indicated that MCM-48-M exhibited a random distribution in the PCL matrix. The PCL chemical structure was preserved in NC films as described by Fourier transform infrared (FT-IR) spectroscopy as well as the tensile and thermal properties of NC films. FT-IR and thermogravimetric analysis (TGA) results showed surface modification. X-ray diffraction (XRD) and differential scanning calorimetry (DSC) showed that crystalline symmetries were preserved and the crystallinity of NC films had small variations in all samples before and after irradiation, respectively. But, our results did not indicate major changes showing that this method is successful for the sterilization of PCL/MCM-48-PVA NC films.
Project description:Citrate is a critical metabolic substrate and key regulator of energy metabolism in mammalian cells. It has been known for decades that the skeleton contains most (>85%) of the body's citrate, but the question of why and how this metabolite should be partitioned in bone has received singularly little attention. Here, we show that osteoblasts use a specialized metabolic pathway to regulate uptake, endogenous production, and the deposition of citrate into bone. Osteoblasts express high levels of the membranous Na+-dependent citrate transporter solute carrier family 13 member 5 (Slc13a5) gene. Inhibition or genetic disruption of Slc13a5 reduced osteogenic citrate uptake and disrupted mineral nodule formation. Bones from mice lacking Slc13a5 globally, or selectively in osteoblasts, showed equivalent reductions in cortical thickness, with similarly compromised mechanical strength. Surprisingly, citrate content in mineral from Slc13a5-/- osteoblasts was increased fourfold relative to controls, suggesting the engagement of compensatory mechanisms to augment endogenous citrate production. Indeed, through the coordinated functioning of the apical membrane citrate transporter SLC13A5 and a mitochondrial zinc transporter protein (ZIP1; encoded by Slc39a1), a mediator of citrate efflux from the tricarboxylic acid cycle, SLC13A5 mediates citrate entry from blood and its activity exerts homeostatic control of cytoplasmic citrate. Intriguingly, Slc13a5-deficient mice also exhibited defective tooth enamel and dentin formation, a clinical feature, which we show is recapitulated in primary teeth from children with SLC13A5 mutations. Together, our results reveal the components of an osteoblast metabolic pathway, which affects bone strength by regulating citrate deposition into mineral hydroxyapatite.