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PubMed1999

Initial anchoring and proliferation of fibroblast L-929 cells on unstable surface of calcium phosphate ceramics.

Calcium phosphate ceramics constructed from beta-tricalcium phosphate (TCP) and hydroxyapatite (HAP) have been successfully used as implant materials. However, there is a possibility that these materials are responsible for an unwanted inflammatory response during wound healing. Since TCP is soluble in the body, the instability of this material may contribute to this inflammatory response. Using composite ceramics of TCP and HAP that possessed Ca/P molar ratios of 1.50, 1.55, 1.60, 1.64, and 1.67, the effect of surface instability on fibroblast L-929 cells was investigated. The time-dependent variation of the initial anchoring ratio, cell density, and cell viability were measured. In general, the cells were severely damaged and ruptured on the highly soluble thin surface layer of the TCP-HAP ceramics. The initial anchoring ratio for TCP-HAP ceramics was as high as that for the polystyrene dish (Lux, control). However, viability at 6 h decreased to less than 50% of the initial cell density on ceramics with a Ca/P molar ratio of 1.64 (20% TCP-80% HAP), while 85% of the cells were viable on Lux. The viability on 100% TCP, whose surface is the most highly soluble among the TCP-HAP ceramics used in this study, was reduced to 20%. Morphological observation showed that the anchored cells were ruptured when grown in culture medium on the 100% TCP. Although the high solubility of the thin surface layer on the TCP-HAP ceramics of the carrier was found to be the dominant factor in the decreasing cell viability, the initial viability was enhanced by the stabilization of the surface of the TCP-HAP ceramics by pre-incubating the scaffolds in a culture medium containing 10% fetal bovine serum for 3 d.

Journal Article↗
PubMed2000

Effect of temperature and immersion on the setting of some calcium phosphate cements.

Calcium phosphate cements based on powders containing alpha-Ca3(PO4)2 and aqueous solutions containing Na2HPO4 as accelerator were used to determine the effects of accelerator concentration, temperature and immersion on the setting time. Increases in accelerator concentration and temperature increased the rate of setting, but immersion had a retarding effect. These results were used to design a method whereby a syringe filled with cement paste can be kept ready for injection of the paste into the implantation site for a long time, whereas setting of the cement paste in the body takes place in a short time.

Journal Article↗
PubMed1978

Biological calcium phosphates and their role in the physiology of bone and dental tissues I. Composition and solubility of calcium phosphates.

Variations in the composition of bone and tooth mineral are consistent with the model that the constituents are a mixed microcrystalline apatite (AP)-octocalcium phosphate (OCP) like phase and an amorphous or submicrocrystalline calcium phosphate (ACP) like phase whereby these phases can occur in different proportions. An appropriate model for a description of the variable composition and the solubility behavior of the apatite phase is given by the formula (formula: see text) in which the compositional parameters x, y, z, and u each account for one type of defect mechanism. Other point defects are formed as well by incorporation of minority amounts of ions such as Cl-, K+, and F-; a number of trace elements can substitute for Ca2+ ions under in vivo conditions. It is suggested that the incorporation of ions in or loss from the crystals in contact with aqueous solutions is reversible. Literature data are used to show the direction in which the solubility product of the apatite phase shifts by incorporation of the different physiologically relevant ions. A quantitative evaluation of the available literature data revealed that Na+ and CO3= incorporation is the main cause for shifts in the solubility product of biological apatites.

Apatites↗
PubMed1999

Cellular mechanisms of calcium phosphate ceramic degradation.

Calcium phosphate (CaP) ceramics are widely used for bone substitution in orthopedic, maxillofacial and dental surgery. Many environmental factors are involved in the gradual degradation of calcium phosphate ceramic after implantation, including physiocochemical processes (dissolution-precipitation) and the effects of various cell types. Several of these cell types degrade ceramics by phagocytotic mechanisms (fibroblasts, osteoblasts, monocytes/macrophages) or by an acidic mechanism with a proton pump to reduce the pH of the microenvironment and resorb these synthetic substrates (osteoclasts). Various mesenchymal cells located at the implantation sites can induce the solubilization of CaP ceramics. Crystal-cell contacts were required to induce such crystal dissolution. Mesenchymal cells such as fibroblastic cells are also actively involved in the ceramic degradation process. In this context, CaP crystals underwent dissolution into the phagosome. If osteoclasts resorb CaP ceramics similarly to the natural bone, they possess a phagocytic capability. This phagocytosis mechanism consisted of three steps: crystal phagocytosis, disappearance of the endophagosome envelope membrane and fragmentation of phagocytosed crystals within the cytoplasm. Similar phenomenons have been observed during the phagocytic mechanism induced by monocytes/macrophages. The cellular mechanisms of CaP ceramic degradation are modulated by various parameters, such as the properties of the ceramic itself, the implantation sites and the presence of various proteins (cytokines, hormones, vitamins, ions, etc.). The cells involved in these mechanisms could intervene directly or indirectly through their cytokine/growth factor secretions and their sensitivity to the same molecules. This article reviews recent knowledge on the cellular mechanisms of calcium phosphate ceramic degradation.

Animals↗
PubMed1975

Precipitation of calcium phosphates from electrolyte solutions. III. Radiometric studies of the kinetics of precipitation and aging of calcium phosphates.

Precipitation and precipitate transformation in the system sodium phosphate (pre-adjusted to pH 7.4)--calcium chloride (25 degrees) was studied by means of radiometric analysis using 45Ca and 32P as tracers. Changes in the pH and the total concentrations of calcium and phosphate were followed during solid phase formation and the data were used to calculate composition changes of the precipitates and their supernatants. In all investigated systems two-step precipitation was observed, the precursor being more basic than the secondary precipitate. The composition of the latter was mostly within the range of the composition of octacalcium phosphate. The course of further chemical changes was dependent on the pH established during secondary precipitation. The heterogeneous exchange of the radionuclides between the solid phase and their supernatant solutions was also followed as a function of time. The results indicate that recrystallization through the mother liquid accompanied by composition changes is the dominant mechanism of equilibrium of the solid phases.

Apatites↗
PubMed2002

A biomechanical analysis of fixation of intra-articular distal radial fractures with calcium-phosphate bone cement.

BACKGROUND: Calcium phosphate cement has been used to treat unstable fractures of the distal end of the radius with the intent of avoiding the stiffness and morbidity associated with prolonged immobilization in a cast or external fixation. The purpose of this study was to compare the stability of the fracture fragments after fixation with augmented calcium phosphate cement with that after alternative methods of percutaneous fracture treatment. METHODS: Both an osteotomy and osteoclasis were used to create a model of an intra-articular fracture of the distal part of the radius (AO type C2) with dorsal bone loss in seven pairs of fresh-frozen upper extremities. One wrist from each pair was fixed with an external fixator and three Kirschner wires, and the contralateral wrist was fixed with calcium phosphate cement (Norian SRS) and three Kirschner wires (augmented calcium phosphate cement). Sequentially increasing loads, up to a total of 100 N, were then applied to the major flexors and extensors of the wrist. Fracture fragment motion was measured by the Optotrak three-dimensional system. RESULTS: Fixation with cement alone failed at the bone-cement interface at <80 N in all specimens. With use of an analysis of variance, augmented external fixation was found to provide significantly increased stability to the radial fragment compared with that provided by augmented calcium phosphate cement in four of the six axes tested (e.g., mean motion [and standard deviation] in flexion-extension was 3.0 degrees +/- 2.93 degrees versus 11.1 degrees +/- 13.08 degrees, respectively; p = 0.001). Augmented calcium phosphate cement was found to provide greater stability for the radial fragment than were Kirschner wires alone in three axes (e.g., mean motion in flexion-extension was 11.1 degrees +/- 13.08 degrees versus 36.5 degrees +/- 13.03 degrees, respectively; p = 0.001). CONCLUSIONS: Calcium phosphate cement alone is insufficient to withstand physiologic flexion-extension motion of the wrist without supplemental wire fixation. When supplemented with Kirschner wires, fixation with bone cement is more stable than are Kirschner wires alone, but it is significantly less stable than augmented external fixation.

Biomechanical Phenomena↗
PubMed2005

Strengthening of calcium phosphate cement by compounding calcium carbonate whiskers.

The purpose of this study was to investigate how aragonite (calcium carbonate) whiskers influenced the strengthening and carbonating of alpha-tricalcium phosphate (alpha-TCP) based calcium phosphate cement. Aragonite whiskers of 0.95 microm width with an aspect ratio of 6.6 were prepared. The cement powder, alpha-TCP containing 0-50 mass% aragonite whisker, was mixed with 0.6 mol/L NaH2PO4 solution and incubated at 37 degrees C and 100% relative humidity. Diametral tensile strength (DTS) value increased significantly when appropriate amount of aragonaite whiskers was added. For example, DTS value of set cement containing 20 mass% aragonite whisker was 5.8 +/- 0.5 MPa, whereas DTS value of set cement containing no whiskers was 1.3 +/- 0.2 MPa after 1-week incubation. SEM observation revealed that the shape of the whiskers and the densification of the structure could have contributed to the strengthening of the set cement. Moreover, FTIR spectra implied that a bone-like carbonated apatite was precipitated in the cement. The results obtained in the present study revealed that the shape as well as any slight dissolution of aragonite whiskers could contribute to improving the properties of a-TCP based calcium phosphate cement.

Apatites↗
PubMed1999

Addition of cohesion promotors to calcium phosphate cements.

Many calcium phosphate cements (CPC) pastes tend to disintegrate upon early contact with blood or other aqueous (body) fluids, which inhibits the use of these materials for clinical use as for bone repair, reconstruction and augmentation. In studies on CPCs based on tetracalcium phosphate and dicalcium hydrogen phosphate others have suggested to use sodium alginate, cellulose derivatives or chitosan derivatives dissolved in the cement liquid for improving the cohesion of CPC pastes. In this study 10 other organic compounds were shown to act as cohesion promotors in the case of CPCs based on alpha-tertiary calcium phosphate as the main active ingredient.

Biocompatible Materials↗
PubMed2005

A novel method for the simultaneous, titrant-free control of pH and calcium phosphate mass yield.

Calcium phosphate (CaP) bioceramics have long been of interest for the unique properties that they exhibit as bone substitute materials. By harnessing the unique bone-bonding capacity of CaP's, biomaterials scientists have made great strides over the past 2 decades to produce novel materials to assist in the treatments of defects caused by trauma, disease, or both. In recent years, however, it has become apparent that the traditional set of techniques used to produce calcium phosphates does not satisfy all of the requirements necessary to meet the challenges of emerging applications. In particular, recent interest in (i) the synthesis of coprecipitated CaP/bioorganic composites and (ii) the investigation of the mechanisms of biomineralization has highlighted the need for new methods to control pH and CaP mass yield.

Biocompatible Materials↗
PubMed2004

Transfection of adherent and suspended cells by calcium phosphate.

DNA-calcium phosphate coprecipitates have been used for 30 years as an efficient method to introduce genetic material into cells. The method involves simple solutions that can be prepared or purchased by the experimentalist. All the numerous variations of the protocol found in the literature are based on the same principle--a spontaneous precipitation that occurs in supersaturated solutions. When DNA is present during this process, it is readily incorporated into the forming calcium phosphate precipitate. Although a wide range of conditions will lead to precipitates, high transfection efficiencies are only obtained within a narrow range of optimized parameters that assure certain properties of the precipitate. This paper describes several physico-chemical parameters that are critical to adapt the method to a particular cell line and/or cultivation condition. Examples of protocols that were established and tested within the authors' laboratory are presented. The article also emphasizes differences between transfections of adherent and suspended cells.

Animals↗
PubMed1979

Effect of human salivary proteins on the precipitation kinetics of calcium phosphate.

Inhibition of calcium phosphate precipitation in saliva, and prevention of the formation of mineral accretions on tooth surfaces, has been ascribed to the existence of inhibiting salivary macromolecules. Marked reductions in the crystal growth rate of hydroxyapatite (HA) seeds were measured in supersaturated solutions containing either of two proline-rich proteins, PRP1 or PRP3, or statherin; the three macromolecules were isolated from human parotid saliva. The reductions were also observed when the HA seeds were pretreated with solutions of the macromolecules before adding them to the supersaturated calcium phosphate solution. This effect was very similar in the case of the two PRPs and it was directly related to the extent of adsorption site coverage of these proteins on the HA seeds. The effect of statherin was larger than anticipated from its adsorption behavior. However, comparison on the basis of number of moles adsorbed per unit area of HA shows that the PRP are more effective inhibitors than statherin. The macromolecule concentrations used were considerably lower than those in the salivary secretions, therefore these macromolecules could readily prevent mineral accretion on tooth surfaces through their adsorption onto the enamel surface.

Adsorption↗
PubMed1990

Intracellular distribution of DNA internalized through calcium phosphate precipitation.

Although calcium phosphate precipitation is the most commonly used method for DNA-mediated gene transfer, the mechanism for its action is unknown. We showed recently that both transient and stable expression of exogenous genes in the transfected cells are entirely dependent on DNA internalized through active endocytosis. We now report on the subcellular distribution of the endocytosed DNA. After exposure to calcium phosphate-precipitated DNA, cultured fibroblasts internalized less than 10% of the DNA into the nuclei fraction. About 20% was recovered in each of the putative plasma membrane and vesicular organelle fractions. Although over 50% was recovered in the cytosolic fraction, it was completely degraded to oligonucleotides of smaller than 100 bp. In contrast, intact DNA molecules were recovered in all the other subcellular fractions. Similar patterns of DNA distribution were observed not only in the easily transformed mouse cells (Ltk-) but also in the transformation-resistant human primary fibroblasts. In conclusion, DNA-mediated gene transfer by calcium phosphate precipitation is an inefficient procedure because over 50% of the DNA is almost immediately degraded and released into the cytosol. Contrary to accepted views, DNA macromolecules do not seem to pass through the cytosol before reaching the nuclei. A novel transport pathway is proposed in which exogenous DNA molecules may be transferred directly by intermediary vesicles from the endocytic-lysosomal compartment to the nucleus.

Animals↗
PubMed1996

Comparison of calcium phosphate cement mixture and pure calcium hydroxide as direct pulp-capping agents.

Calcium phosphate cement (CPC) and pure calcium hydroxide were used as direct pulp-capping agents on the deliberately exposed pulp tissue of 60 teeth in five monkeys. Their effects on the pulp tissue of individual teeth were observed and histologically compared after 12, 20 and 24 weeks. The results showed that both materials produced similar responses with regard to their biocompatibility and induction of hard tissue barrier formation. Vasodilatation, chronic inflammation and calcification nidi scattered within the pulp tissue of the 12-week group were observed. Twenty weeks after application of capping materials, a crude reparative dentinal bridge with inclusion of soft tissue or bulky capping agents appeared. A more mature, hard tissue barrier with a better degree of mineralization and formation of dentinal tubules was demonstrated in the 24-week group. The above findings associated with the abilities of self-setting and fair compressive strength suggest that CPC appears superior to pure calcium hydroxide and may have potential for clinical application, although many issues remain to be further investigated.

Animals↗
PubMed2001

Effect on composition of dry mechanical grinding of calcium phosphate mixtures.

For diverse reasons, calcium phosphates used to prepare hydraulic calcium phosphate cements can be ground mixed. The grinding with a rotating micromill of monocalcium phosphate monohydrate or anhydrous, dicalcium phosphate dihydrate or anhydrous with calcium oxide, calcium hydroxide, calcium carbonate, tetracalcium phosphate, or alpha- or beta-tricalcium phosphate was studied for different calcium to phosphate (Ca/P) ratios, rotating rates, masses of balls, and environmental conditions. During dry grinding by ball milling, anhydrous or hydrated acid calcium phosphates can mechanochemically react with anhydrous or hydrated basic calcium salts to form dicalcium phosphate dihydrate or anhydrous, noncrystalline calcium phosphate, and/or calcium deficient or stoichiometric hydroxyapatite, depending on the Ca/P ratio in the mixture and the time of grinding. The reaction rate is a function of the rotation rate and the mass of the balls. Water is not necessary to initiate the reaction but facilitates it because hydrated salts react faster than the corresponding anhydrous salts. Neither carbon dioxide nor carbonate ions seem to have any influence on the transformation kinetics. The transformations that occur during grinding influence the final mechanical properties of hydraulic calcium phosphate cements prepared from these materials. Thus, if a grinding step of the starting materials is planed, the grinding conditions will have to be particularly well defined to obtain reproducible results.

Biocompatible Materials↗