14 research outputs found
Guanine crystal formation by the unicellular organism Phacotus lenticularis is part of a cellular stress response.
No full textOrganic crystals, and in particular guanine crystals, are widely used by multicellular organisms for manipulating light and producing structural colors. Many single celled eukaryotic organisms also produce organic crystals, and guanine is the most abundant type produced. Their functions are thought to be related to the fact that guanine is nitrogen rich. Here we studied a freshwater unicellular eukaryotic alga, Phacotus lenticularis, and found that when the growth medium is depleted in phosphorus, the alga stops reproducing and produces intracellular birefringent particles inside vesicles. Cryo-SEM showed that these particles are faceted and are located within membranes inside the cell. Using Raman spectroscopy, we showed that these particles are β-guanine crystals. 3D tomograms produced using cryo-soft-X-ray-microscopy quantitatively documented the increase in cell volume and distribution of guanine crystals within the cells with increasing time of phosphorous deprivation. The tomograms also showed additional morphological changes in other cellular organelles, namely starch granules, chloroplasts, nuclear DNA and membranes. The combined observations all indicate that under phosphorous depletion, the algal cells undergo a massive stress response. As guanine crystal formation is part of this response, we conclude that guanine crystals are formed in response to stress, and this is not related to nitrogen availability. Upon addition of phosphate to the P-depleted media, the algal cells, with their guanine crystals, resume reproduction. From this we conclude that the guanine crystals somehow contribute to the recovery from stress
Study of Osteoclast Adhesion to Cortical Bone Surfaces: A Correlative Microscopy Approach for Concomitant Imaging of Cellular Dynamics and Surface Modifications
No full textBone remodeling relies on the coordinated
functioning of osteoblasts,
bone-forming cells, and osteoclasts, bone-resorbing cells. The effects
of specific chemical and physical bone features on the osteoclast
adhesive apparatus, the sealing zone ring, and their relation to resorption
functionality are still not well-understood. We designed and implemented
a correlative imaging method that enables monitoring of the same area
of bone surface by time-lapse light microscopy, electron microscopy,
and atomic force microscopy before, during, and after exposure to
osteoclasts. We show that sealing zone rings preferentially develop
around surface protrusions, with lateral dimensions of several micrometers,
and ∼1 μm height. Direct overlay of sealing zone rings
onto resorption pits on the bone surface shows that the rings adapt
to pit morphology. The correlative procedure presented here is noninvasive
and performed under ambient conditions, without the need for sample
labeling. It can potentially be applied to study various aspects of
cell-matrix interactions
Introduction of correlative light and airSEMTM microscopy imaging for tissue research under ambient conditions
No full textNATO Advanced Study Institute on Engineering of Crystalline Materials Properties : State of the Art in Modeling Design and Applications. New Materials for better Defence and Security
No full textThis volume collects the lecture notes (ordered alphabetically according to the first author surname) of the talks delivered by the main speakers at the Erice 2007 International School of Crystallography, generously selected by NATO as an Advanced Study Institute (# 982582). The aim of the school was to discuss the state-of-the-art in molecular materials design, that is, the rational analysis and fabrication of crystalline solids showing a predefined structural organization of their component molecules and ions, which results in the manifestation of a specific collective property of technological interest. The School was held on June 7–17, 2007, in Erice (an old town, over 3000 years, located on the top of a Sicilian hill that oversees the sea near Trapani). The school developed following two parallel lines. First we established “where we are” in terms of modelling, design, synthesis and applications of crystalline solids with predefined properties. Second, we attempted to define current and possible futuristic lines of development in the quest for novel molecule-based materials with potential applications in magnetism, conductivity and superconductivity, non-lineal optics (NLO), drug delivery, and nanotechnology. In recent years, solid state chemistry and crystal engineering have evolved at the intersection between the top-down and bottom-up approaches towards materials design and fabrication. An ever-increasing number of scientists are learning how to control self-assembly, molecular recognition, and other fundamental processes on the way to achieving ‘tailor-made’ materials, such as crystal nucleation, crystal growth, and polymorphism
Orchestrated regulation of iron trafficking proteins in the kidney during iron overload facilitates systemic iron retention.
No full textThe exact route of iron through the kidney and its regulation during iron overload are not completely elucidated. Under physiologic conditions, non-transferrin and transferrin bound iron passes the glomerular filter and is reabsorbed through kidney epithelial cells, so that hardly any iron is found in the urine. To study the route of iron reabsorption through the kidney, we analyzed the location and regulation of iron metabolism related proteins in kidneys of mice with iron overload, elicited by iron dextran injections. Transferrin Receptor 1 was decreased as expected, following iron overload. In contrast, the multi-ligand hetero-dimeric receptor-complex megalin/cubilin, which also mediates the internalization of transferrin, was highly up-regulated. Moreover, with increasing iron, intracellular ferritin distribution shifted in renal epithelium from an apical location to a punctate distribution throughout the epithelial cells. In addition, in contrast to many other tissues, the iron exporter ferroportin was not reduced by iron overload in the kidney. Iron accumulated mainly in interstitial macrophages, and more prominently in the medulla than in the cortex. This suggests that despite the reduction of Transferrin Receptor 1, alternative pathways may effectively mediate re-absorption of iron that cycles through the kidney during parenterally induced iron-overload. The most iron consuming process of the body, erythropoiesis, is regulated by the renal erythropoietin producing cells in kidney interstitium. We propose, that the efficient re-absorption of iron by the kidney, also during iron overload enables these cells to sense systemic iron and regulate its usage based on the systemic iron state
Cubilin expression in iron loaded kidneys is elevated both in the cortex and the medulla.
No full textThe fixed kidney sections were incubated with cubilin or TfR1 antibodies and stained by IF or IHC. (A-F) Cubilin was observed apically in the cortex, as expected, with increased staining in kidneys from iron overloaded mice (compare A to D). Interestingly, in PIO, significant cubilin expression was also observed in the medulla (compare B to E, and C to F). Inserts are negative controls (N.C) for cubilin staining; (G-I) Visualization of TfR1 by IHC (G-H) and by IF (I). (G) TfR1 staining was seen mainly in the medulla, (the dashed line separates the cortex from the medulla. In the medulla, TfR1 was seen apically (H-I) (indicated by arrows). Inserts are negative controls (N.C) for TfR1 staining; scale bar of 50μm, n = 3 for control and iron-loaded mice each.</p
Ferritin distribution in renal epithelial cells is regulated by the iron status: Kidney sections from control, iron-loaded and Irp2-/- mice were stained with H-ferritin antibody.
No full textFerritin was apically polarized in kidneys from Irp2-/- mice and to some extent also in wild-type control mice (arrow heads). In contrast, in iron overloaded mice ferritin was distributed throughout the cells and also found in basolateral regions (arrows). Scale bar represents 50μm.</p
PIO did not cause morphologic kidney damage, but affected the spleen.
No full textFixed kidney and spleen sections from control and iron-loaded mice were histologically stained (H&E) and imaged. No damage was observed in kidney sections of iron-loaded mice (B and D) compared to the control sections (A and C). However, iron accumulation caused damage to the spleen as can be evaluated by moderate degree of hemosiderosis (seen as brownish pigment in the red pulp of section F compared to E), and by a mild depletion of lymphocytes in the white pulp (indicated by red arrowheads); the inserts in the left corner of E and F are higher magnifications of these sections. Scale bar 100 μm, n = 3 for control and iron-loaded mice each.</p
Cubilin and Tfr1 are regulated by iron in opposite directions.
No full textTfR1 (A) and cubilin (B) protein levels were tested by Western-blot analysis of kidney lysates. Membranes were probed with either cubilin, TfR1 or actin antibodies. TfR1 levels in iron over-loaded mice were decreased. Cubilin levels were increased in kidneys from iron–loaded mice. (C, D and E) Kidney mRNA levels of TfR1, cubilin and megalin were evaluated by qPCR. (C) TfR1 mRNA levels correlated with protein levels (n = 5, *P(D) no significant difference in cubilin mRNA levels was observed (n = 6). (E) Megalin mRNA expression increased in iron overloaded kidneys (n = 7, **p<0.005).</p
Iron accumulates in the kidney-interstitium of PIO mice.
No full textFixed kidney sections from PIO mice (A-B and D-F) and from dietary iron overloaded mice (C) were stained and imaged. (A-C) Ferric iron was visualized with Prussian blue staining in cortex and medulla, respectively. In PIO mice, most iron accumulated in and around glomeruli and in the interstitium, and markedly little iron was detected in renal epithelial cells. In contrast, following dietary iron overload, most iron accumulated in proximal tubule epithelium of the cortex. (D) Light microscope; the black square indicates a sub-region of the medulla, which is enlarged in (E and F). (E) AirSEM analysis of the sub-region followed by EDX [Fe] mapping (F) indicated iron accumulation (marked with arrows) in the interstitium of the medulla. (G) Regions of interest in tubules and interstitium of cortex and medulla were selected and iron levels were quantified using airSEM. At least 2-fold increase of interstitial iron in the medulla compared to cortex was measured. ** P< 0.0001.</p
