Cell Adhesion to Crystal Surfaces: A Model for Initial Stages in the Attachment of Cells to Solid Substrates

This study addresses the mechanism of the chirally-restricted, ROD-independent adhesion of A6 epithelial cells to the {011} faces of calcium {R,R)-tartrate tetrahydrate crystals. The extensive and rapid adhesion of the cells to these surfaces, in the presence or absence of serum proteins, is distinctly different from the extracellular matrix-mediated adhesion to conventional tissue culture surfaces or to the {101} faces of the same crystals. The differences are manifested by insensitivity to ATP depletion, to disruption of microfilaments and microtubules and even to formaldehyde fixation of the cells. Furthermore, trypsin pretreatment does not affect cell attachment to the {011} faces, nor does trypsin post-treatment cause cell detachment from the crystals. We also noticed that the rapid adhesion to the crystal surface bears several lines of similarity to the early temporal stages in cell adhesion to regular tissue culture surfaces. Based on these observations and additional theoretical considerations, it is proposed that ·the molecular interactions responsible for the cell adhesion to the {011} surfaces may serve as models for an early engagement stage in cell adhesion which precedes, and may be essential for, the formation of stable and long-term contacts

Tuning of Crystal Nucleation and Growth by Proteins: Molecular Interactions at Solid-Liquid Interfaces in Biomineralization

The mineralized tissues of a bivalve mollusk and a sea urchin are both composed of calcium carbonate crystals that are intimately associated with acidic glycoproteins. In vitro studies in which carboxylate-, carbonate- and phosphate-containing crystals are grown in the presence of partially purified acidic glycoproteins from these two tissues show that some of these macro- molecules are able to interact specifically with certain crystal faces. Significantly all the affected crystal faces contain a common stereochemical motif. Interesting differences, however, were observed in the modes of interaction between the mollusk and sea urchin derived acidic glycoproteins. Only the former can induce oriented calcite nucleation in vitro and only the latter can interact from solution with specific calcite crystal faces. These differences are ascribed in part to the fact that the mollusk macromolecules are much more acidic than those from the sea urchin. Some of the acidic glycoproteins are also occluded inside the growing crystals. In the case of the sea urchin, and not of the mollusk, the proteins are preferentially located at specific crystal planes and their presence influences the mechanical properties of the crystal. A detailed study of these composite crystals by X-ray synchrotron radiation shows how the presence of the protein influences the crystal mosaicity. The interactions revealed by these studies follow well defined stereochemical rules, tuned by electrostatic forces. They, in turn, provide new \u27insight into some of the basic underlying processes occurring in biomineralization

On the morphology of ammonium nitrate (III): theory and observation

The aim of this paper is to derive on a theoretical basis the morphology of crystals of ammonium nitrate, phase III, and to compare the results with experimental growth forms. The theory used is based on the concepts of periodic bond chain (PBC), F face and connected net, developed by Hartman and Perdok. Further an Ising model is used to determine roughening temperatures. Based on different criteria theoretical growth forms are predicted and compared with experiments

Forming nacreous layer of the shells of the bivalves Atrina rigida and Pinctada margaritifera: An environmental- and cryo-scanning electron microscopy study

International audienceA key to understanding control over mineral formation in mollusk shells is the microenvironment inside the pre-formed 3-dimensional organic matrix framework where mineral forms. Much of what is known about nacre formation is from observations of the mature tissue. Although these studies have elucidated several important aspects of this process, the structure of the organic matrix and the microenvironment where the crystal nucleates and grows are very difficult to infer from observations of the mature nacre. Here, we use environmental- and cryo-scanning electron microscopy to investigate the organic matrix structure at the onset of mineralization in the nacre of two mollusk species: the bivalves Atrina rigida and Pinctada margaritifera. These two techniques allow the visualization of hydrated biological materials coupled with the preservation of the organic matrix close to physiological conditions. We identified a hydrated gel-like protein phase filling the space between two interlamellar sheets prior to mineral formation. The results are consistent with this phase being the silk-like proteins, and show that mineral formation does not occur in an aqueous solution, but in a hydrated gel-like medium. As the tablets grow, the silk-fibroin is pushed aside and becomes sandwiched between the mineral and the chitin layer

MMP-2, MMP-9 and their inhibitors TIMP-2 and TIMP-1 production by human monocytes in vitro in the presence of different forms of hydroxyapatite particles.

DOI : 10.1016/j.biomaterials.2003.09.034After calcium-phosphates biomaterials based implantation like hydroxyapatite (HA) coating, particles are released in the periprosthetic tissues. Wear-debris induced fibrous membranes contain macrophage subsets that can produce metalloproteinases (MMPs), which are considered to be key enzymes in extra-cellular matrix turnover. Tissue inhibitors of metalloproteinases (TIMPs) are important regulator of MMPs activity. Interleukin-1 mainly produced by monocytes can also regulate MMPs production. In the present work, we have evaluated the effect of HA particles characteristics (size, shape and sintering temperature) on the MMP-2, -9 and their respective inhibitors TIMP-2, -1 production. Our results demonstrate that sintering temperature (that modify crystal size and surface area) have little effect on MMPs and TIMPs production. Non-phagocytable particles induced more MMP-9, although phagocytable particles induced more IL-1β release. The shape of the particles was the most important factor since needle-shaped particles induced the most significant up-regulated expression of MMPs and IL-1β

Effect of Langmuir monolayer of bovine serum albumin protein on the morphology of calcium carbonate

Bovine serum albumin (BSA) Langmuir monolayer, as a model of biomineralization-associated proteins, was used to study its effect on regulated biomineralization of calcium carbonate. The effects of the BSA Langmuir monolayer and the concentration of the subphase solution on the nucleation and growth processes and morphology of the calcium carbonate crystal were investigated. The morphology and polymorphic phase of the resulting calcium carbonate crystals were characterized by scanning electron microscopy (SEM) and X-ray diffraction analysis (XRD). Moreover, the interaction mechanisms of the subphase solution with the BSA Langmuir monolayer were discussed. It was found that BSA Langmuir monolayer could be used as a template to successfully manipulate the polymorphic phase and crystal morphology of calcium carbonate and had obvious influence on the oriented crystallization and growth. The final morphology or aggregation mode of the calcite crystal was closely dependent on the concentration of calcium bicarbonate solution. It is expected that this research would help to better understand the mechanism of biomineralization by revealing the interactions between protein matrices and crystallization of calcium carbonate crystal.Comment: 4 pages, 4 figure

Application of 3D MAPs pipeline identifies the morphological sequence chondrocytes undergo and the regulatory role of GDF5 in this process

The activity of epiphyseal growth plates, which drives long bone elongation, depends on extensive changes in chondrocyte size and shape during differentiation. Here, we develop a pipeline called 3D Morphometric Analysis for Phenotypic significance (3D MAPs), which combines light-sheet microscopy, segmentation algorithms and 3D morphometric analysis to characterize morphogenetic cellular behaviors while maintaining the spatial context of the growth plate. Using 3D MAPs, we create a 3D image database of hundreds of thousands of chondrocytes. Analysis reveals broad repertoire of morphological changes, growth strategies and cell organizations during differentiation. Moreover, identifying a reduction in Smad 1/5/9 activity together with multiple abnormalities in cell growth, shape and organization provides an explanation for the shortening of Gdf5 KO tibias. Overall, our findings provide insight into the morphological sequence that chondrocytes undergo during differentiation and highlight the ability of 3D MAPs to uncover cellular mechanisms that may regulate this process

Enhanced stability and local structure in biologically relevant amorphous materials containing pyrophosphate

There is increasing evidence that amorphous inorganic materials play a key role in biomineralisation in many organisms, however the inherent instability of synthetic analogues in the absence of the complex in vivo matrix limits their study and clinical exploitation. To address this, we report here an approach that enhances long-term stability to >1 year of biologically relevant amorphous metal phosphates, in the absence of any complex stabilisers, by utilising pyrophosphates (P2O7 4-); species themselves ubiquitous in vivo. Ambient temperature precipitation reactions were employed to synthesise amorphous Ca2P2O7.nH2O and Sr2P2O7.nH2O (3.8 < n < 4.2) and their stability and structure were investigated. Pair distribution functions (PDF) derived from synchrotron X-ray data indicated a lack of structural order beyond ~8 A° in both phases, with this local order found to resemble crystalline analogues. Further studies, including 1H and 31P solid state NMR, suggest the unusually high stability of these purely inorganic amorphous phases is partly due to disorder in the P–O–P bond angles within the P2O7 units, which impede crystallization, and to water molecules, which are involved in H-bonds of various strengths within the structures and hamper the formation of an ordered network. In situ high temperature powder X-ray diffraction data indicated that the amorphous nature of both phases surprisingly persisted to ~450° C. Further NMR and TGA studies found that above ambient temperature some water molecules reacted with P2O7 anions, leading to the hydrolysis of some P–O–P linkages and the formation of HPO4 2- anions within the amorphous matrix. The latter anions then recombined into P2O7 ions at higher temperatures prior to crystallization. Together, these findings provide important new materials with unexplored potential for enzyme-assisted resorption and establish factors crucial to isolate further stable amorphous inorganic materials