Control of Vertebrate Skeletal Mineralization by Polyphosphates

BACKGROUND:Skeletons are formed in a wide variety of shapes, sizes, and compositions of organic and mineral components. Many invertebrate skeletons are constructed from carbonate or silicate minerals, whereas vertebrate skeletons are instead composed of a calcium phosphate mineral known as apatite. No one yet knows why the dynamic vertebrate skeleton, which is continually rebuilt, repaired, and resorbed during growth and normal remodeling, is composed of apatite. Nor is the control of bone and calcifying cartilage mineralization well understood, though it is thought to be associated with phosphate-cleaving proteins. Researchers have assumed that skeletal mineralization is also associated with non-crystalline, calcium- and phosphate-containing electron-dense granules that have been detected in vertebrate skeletal tissue prepared under non-aqueous conditions. Again, however, the role of these granules remains poorly understood. Here, we review bone and growth plate mineralization before showing that polymers of phosphate ions (polyphosphates: (PO(3)(-))(n)) are co-located with mineralizing cartilage and resorbing bone. We propose that the electron-dense granules contain polyphosphates, and explain how these polyphosphates may play an important role in apatite biomineralization. PRINCIPAL FINDINGS/METHODOLOGY:The enzymatic formation (condensation) and destruction (hydrolytic degradation) of polyphosphates offers a simple mechanism for enzymatic control of phosphate accumulation and the relative saturation of apatite. Under circumstances in which apatite mineral formation is undesirable, such as within cartilage tissue or during bone resorption, the production of polyphosphates reduces the free orthophosphate (PO(4)(3-)) concentration while permitting the accumulation of a high total PO(4)(3-) concentration. Sequestering calcium into amorphous calcium polyphosphate complexes can reduce the concentration of free calcium. The resulting reduction of both free PO(4)(3-) and free calcium lowers the relative apatite saturation, preventing formation of apatite crystals. Identified in situ within resorbing bone and mineralizing cartilage by the fluorescent reporter DAPI (4',6-diamidino-2-phenylindole), polyphosphate formation prevents apatite crystal precipitation while accumulating high local concentrations of total calcium and phosphate. When mineralization is required, tissue non-specific alkaline phosphatase, an enzyme associated with skeletal and cartilage mineralization, cleaves orthophosphates from polyphosphates. The hydrolytic degradation of polyphosphates in the calcium-polyphosphate complex increases orthophosphate and calcium concentrations and thereby favors apatite mineral formation. The correlation of alkaline phosphatase with this process may be explained by the destruction of polyphosphates in calcifying cartilage and areas of bone formation. CONCLUSIONS/SIGNIFICANCE:We hypothesize that polyphosphate formation and hydrolytic degradation constitute a simple mechanism for phosphate accumulation and enzymatic control of biological apatite saturation. This enzymatic control of calcified tissue mineralization may have permitted the development of a phosphate-based, mineralized endoskeleton that can be continually remodeled

Identification of genes expressed by immune cells of the colon that are regulated by colorectal cancer-associated variants.

A locus on human chromosome 11q23 tagged by marker rs3802842 was associated with colorectal cancer (CRC) in a genome-wide association study; this finding has been replicated in case-control studies worldwide. In order to identify biologic factors at this locus that are related to the etiopathology of CRC, we used microarray-based target selection methods, coupled to next-generation sequencing, to study 103 kb at the 11q23 locus. We genotyped 369 putative variants from 1,030 patients with CRC (cases) and 1,061 individuals without CRC (controls) from the Ontario Familial Colorectal Cancer Registry. Two previously uncharacterized genes, COLCA1 and COLCA2, were found to be co-regulated genes that are transcribed from opposite strands. Expression levels of COLCA1 and COLCA2 transcripts correlate with rs3802842 genotypes. In colon tissues, COLCA1 co-localizes with crystalloid granules of eosinophils and granular organelles of mast cells, neutrophils, macrophages, dendritic cells and differentiated myeloid-derived cell lines. COLCA2 is present in the cytoplasm of normal epithelial, immune and other cell lineages, as well as tumor cells. Tissue microarray analysis demonstrates the association of rs3802842 with lymphocyte density in the lamina propria (p = 0.014) and levels of COLCA1 in the lamina propria (p = 0.00016) and COLCA2 (tumor cells, p = 0.0041 and lamina propria, p = 6 × 10(-5)). In conclusion, genetic, expression and immunohistochemical data implicate COLCA1 and COLCA2 in the pathogenesis of colon cancer. Histologic analyses indicate the involvement of immune pathways

Mechanical thrombectomy in acute stroke – Five years of experience in Poland

Objectives Mechanical thrombectomy (MT) is not reimbursed by the Polish public health system. We present a description of 5 years of experience with MT in acute stroke in Comprehensive Stroke Centers (CSCs) in Poland. Methods and results We retrospectively analyzed the results of a structured questionnaire from 23 out of 25 identified CSCs and 22 data sets that include 61 clinical, radiological and outcome measures. Results Most of the CSCs (74%) were founded at University Hospitals and most (65.2%) work round the clock. In 78.3% of them, the working teams are composed of neurologists and neuro-radiologists. All CSCs perform CT and angio-CT before MT. In total 586 patients were subjected to MT and data from 531 of them were analyzed. Mean time laps from stroke onset to groin puncture was 250±99min. 90.3% of the studied patients had MT within 6h from stroke onset; 59.3% of them were treated with IV rt-PA prior to MT; 15.1% had IA rt-PA during MT and 4.7% – emergent stenting of a large vessel. M1 of MCA was occluded in 47.8% of cases. The Solitaire device was used in 53% of cases. Successful recanalization (TICI2b–TICI3) was achieved in 64.6% of cases and 53.4% of patients did not experience hemorrhagic transformation. Clinical improvement on discharge was noticed in 53.7% of cases, futile recanalization – in 30.7%, mRS of 0–2 – in 31.4% and mRS of 6 in 22% of cases. Conclusion Our results can help harmonize standards for MT in Poland according to international guidelines

Cognitive aspects of participation: Evidence from two studies

The trouble of proving the effects of participation lies in the mismatch between three aspects of ownership: physical (possession), legal (ownership proper) and psychological (participation). In our interdisciplinary systemic model of ownership, we propose 10 relationships related to ownership/participation from: „A is a part of B” - greatest involvement to „A does not know about B” - the least involvement. „A” and „B” may take different values of: a person, an institution, a community, a group, an object (material, energetic, informational, purchasing). Once formalized we can view the studies in participation from one, system theory point of view, and formulate hypotheses related to many aspects of ownership. A multilevel analysis with multiple measures of both participation and effectiveness from two data sets has supported the proposed model

RESTORATION OF THE PAINTED CEILING IN THE SO-CALLED „PRINCES’ HALL” OF THE MONASTERY AT LUBIĄŻ

The authors presented the results of restoration carried out on the painted ceiling adorning the „Princes’Hall” in the palace of abbots within the monastery site at Lubiąż, Wroclaw voivodship. The abbot’s palace is at present adapted for the use of the National Museum, Wroclaw. The painting under discussion was executed in 1737 by a Dutch artist, Christian F. Bentuma with oil paints laid on the bolus ground layer brought on to the canvas. The whole comprises ten sections the total area of which amounts to 296.3 sq. metres and its artistic composition is formed of numerous human figures representing the Apotheosis of the Holy Faith containing a highly complicated illustrative programme. Prior to strating the restoration it has prove that the convas constituting support is in extremely poor condition and as a result of high amplitudes of temperatures prevailing within the room it exhibited large slackening areas and undulations and, in addition, was seriously decayed by the rainfall water which was formerly penetrating through the roof. Numerous holes and cuts were found in the canvas and in the painted layer several swellings as well as crackings and whitenings of the original varnish. Before the basic restoration the painting was dismantled and its individual sections lowered by the simultaneous use of four ropes. Within the next step of restoration they were lined with linen canvas having the structure similar to that of original canvas, their front and rear surfaces mechanically cleaned whereas the losses filled. Also all the weakened areas were covered with a mass prepared from bee-wax and gum turpentine. Then the cross stretching was applied, the sections fastened to stretchers and the losses retouched by stippling. As the ground for retouching the varnish with 5 % addition of linseed oil was used and the retouches were carried out with the „Rembrandt” oil paints laid on the oil-resin binder

Structure and dynamics of a DNA duplex containing single a-anomeric deoxyadenosine residue.

Structure and dynamics of an undecamer DNA duplex containing a single α-anomeric deoxyadenosine residue placed in opposition to a thymidine unit have been studied using simulation of molecular dynamics in aqueous solution. Despite several noticeable deviations from the B-DNA duplex structure caused by the anomerisation, such as: West type puckering of the α-anomeric sugar, disrupted base stacking pattern and unstable duplex bending, the formation of a non-classical α-dA-T pair was observed. A novel way of visual presentation of trajectory data allowing high throughput screening of the conformational parameters is presented

Nicotine-induced Disturbances of Meiotic Maturation in Cultured Mouse Oocytes: Alterations of Spindle Integrity and Chromosome Alignment

Abstract We investigated whether nicotine exposure in vitro of mouse oocytes affects spindle and chromosome function during meiotic maturation (M-I and M-II). Oocytes in germinal vesicle (GV) stage were cultured in nicotine for 8 h or for 16 h, to assess effects in M-I and in metaphase II (M-II). The latter culture setting used the three protocols: 8 h nicotine then 8 h medium (8N + 8M); 16 h nicotine (16N); 8 h medium then 8 h nicotine (8M + 8N). Non-toxic concentrations of nicotine at 1.0, 2.5, 5.0 and 10.0 mmol/L were used. Spindle-chromosome configurations were analyzed with wide-field optical sectioning microscopy. In 8 h cultures, nicotine exposure resulted in dose-related increased proportions of M-I oocytes with defective spindle-chromosome configurations. A dose-related delayed entry into anaphase I was also detected. In 16 h cultures, nicotine exposure for the first 8 h (8N + 8M), or for 16 h (16N), resulted in dose- and time-related increased proportions of oocytes arrested in M-I (10 mmol/L; 8 h: 53.2%, controls 9.6%; 16 h: 87.6%, controls 8.5%). Defects in M-I spindles and chromosomes caused M-I arrest leading to dose-related decreased proportions of oocytes that reached metaphase-II (10 mmol/L 8 h: 46.8%, controls 90.4%;16 h: 12.4%, controls 91.5%). A delayed anaphase-I affected the normal timing of M-II, leading to abnormal oocytes with dispersed chromosomes, or with double spindles and no polar body. Nicotine exposure during the second 8 h (8M + 8N) resulted in dose-related, increased proportions of M-II oocytes with defective spindles and chromosomes (10 mmol/L: 42.9%, controls 2.0%). Nicotine has no adverse effects on GV break down, but induces spindle and chromosome defects compromising oocyte meiotic maturation and development

The bulge region of HIV-1 TAR RNA binds metal ions in solution

Binding of Mg(2+), Ca(2+) and Co(NH(3))(6)(3+) ions to the HIV-1 TAR RNA in solution was analysed by (19)F NMR spectroscopy, metal ion-induced RNA cleavages and Brownian dynamics (BD) simulations. Chemically synthesised 29mer oligoribonucleotides of the TAR sequence labelled with 5-fluorouridine (FU) were used for (19)F NMR-monitored metal ion titration. The chemical shift changes of fluorine resonances FU-23, FU-25 and FU-40 upon titration with Mg(2+) and Ca(2+) ions indicated specific, although weak, binding at the bulge region with the dissociation constants (K(d)) of 0.9 ± 0.6 and 2.7 ± 1.7 mM, respectively. Argininamide, inducing largest (19)F chemical shifts changes at FU-23, was used as a reference ligand (K(d) = 0.3 ± 0.1 mM). In the Pb(2+)-induced TAR RNA cleavage experiment, strong and selective cleavage of the C24-U25 phosphodiester bond was observed, while Mg(2+) and Ca(2+) induced cuts at all 3-nt residues of the bulge. The inhibition of Pb(2+)-specific TAR cleavage by di- and trivalent metal ions revealed a binding specificity [in the order Co(NH(3))(6)(3+) > Mg(2+) > Ca(2+)] at the bulge site. A BD simulation search of potential magnesium ion sites within the NMR structure of HIV-1 TAR RNA was conducted on a set of 20 conformers (PDB code 1ANR). For most cases, the bulge region was targeted by magnesium cations