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Bioresorption of ceramic strontium-85-labeled calcium phosphate implants in dog femora. A pilot study to quantitate bioresorption of ceramic implants of hydroxyapatite and tricalcium orthophosphate in vivo.

Ceramic hydroxyapatite and tricalcium orthophosphate were radioactively labeled with their strontium-85 analogs. Porous blocks of these materials (dimensions 2.5 X 1.25 X 0.5 cm) were implanted into surgically created defects in dog femora. The fate of the implants was studied by roentgenography, radioactivity measurements, histology, and microradiography. The radioactivity over the implant site was followed for 22 weeks. Implants were retrieved after 20-25 and 50-55 weeks. Hydroxyapatite implants were not affected by biodegradation processes, while tricalcium orthophosphate implants were subject to extensive bioresorption (25%-30% in 22 weeks). Resorption debris from tricalcium orthophosphate implants was found in mononuclear phagocytes and multinuclear osteoclastlike cells. The supposition is that tricalcium orthophosphate is transformed into hydroxyapatite in a physiologic environment. Labeling of ceramic calcium phosphate implants with strontium-85 analogs offers an adequate technique to quantitate bioresorption in vivo.

Animals↗

Augmentation of the edentulous mandible using bone and hydroxyapatite: a comparative study in dogs.

This study compared autogenous bone, freeze-dried homografts, and hydroxyapatite in interpositional grafting of the mandible in beagle dogs. Healing was assessed radiologically and histologically. Results showed that the rate of incorporation of the autografts was greater than that of the homografts, and at both 4 and 12 weeks healing was more advanced. There was also rapid healing around the hydroxyapatite grafts, which became firmly bonded to the surrounding bone. The apatite was not resorbed, and new trabeculae of bone had been deposited onto much of its surface. This investigation confirms that hydroxyapatite is well tolerated and may be a suitable substitute for autogenous bone for augmentation of the atrophic mandible.

Alveolar Ridge Augmentation↗

Fractionation and characterization of chromosomal proteins by the hydroxyapatite dissociation method.

A method was developed which enables the characterization and fractionation of chromosomal proteins according to their chromatin binding properties. The method is based on the ability of hydroxyapatite to bind native chromatin in solutions which do not dissocciate chromosomal proteins from the DNA. The proteins are then selectively dissociated from the immobilized chromatin by treatment with NaCl, urea, or guanidine HCl. The hydroxyapatite dissociation method represents a rapid one-step fractionation procedure which results in the quantitative recovery of chromosomal proteins devoid of nucleic acids and is suitable for large preparations. The hydroxyapatite dissociation method provides a versatile procedure for the study and preparation of chromosomal proteins. The patterns of dissociation of both histones and nonhistone chromosomal proteins by NaCl and urea from chicken oviduct chromatin were characterized by this method. In addition, this technique enabled the purification of the major histone species in a single operation. Partial purification of specific nonhistone proteins, including the estrogen receptor, was also achieved. We suggest that this method will be a useful tool in elucidation of the chemical and biological properties of the proteins from chromatin.

Animals↗

[Determination of Ca/P atomic ratio in calcium-hydroxyapatite by neutron activation analysis].

The Ca/P atomic ratios of synthesized calcium-hydroxyapatite (Ca10(PO4)6(OH2) were determined by neutron activation analysis. In order to obtain calibration curves, the standard samples, such as calcium monohydrogenphosphate, calcium dihydrogenphosphate, etc., were prepared. gamma and beta ray spectra, and beta ray intensities of calcium-hydroxyapatite and standard samples irradiated with reactor neutrons were measured by using a Ge(Li) detector, a low background beta ray spectrometer and GM detector. Based on these results, deviation coefficient of the calibration curve, accuracy of the Ca/P atomic ratio, minimum determination limit, etc. were compared in detail. From the present analysis, the Ca/P atomic ratio of calcium-hydroxyapatite was able to obtain with the standard deviation of about 2% for 1 mg sample.

Activation Analysis↗

Sintered porous DP-bioactive glass and hydroxyapatite as bone substitute.

There is extensive experimental and surgical experience with the use of bone tissue to fill defects in the skeleton, to bridge non-union sites, and to pack defects in bone created from cyst curettage. DP-bioactive glass with a chemical composition of Na2O 8.4%, SiO2 39.6%, P2O5 12% and CaO 40% has been reported as an alternative bone substitute of high mechanical strength, good biocompatibility. and which has a tight bond with living tissue. The bonding layer between DP-bioactive glass and bone tissue was considered to be formed by dissolution of calcium and phosphate ions from the DP-bioactive glass into the surrounding body fluids. The biological hydroxyapatite was suspected to deposit directly onto the bonding layer. In order to confirm the interaction between the DP-bioactive glass and bone tissue, the developed bioactive glass was implanted into rabbit femur condyle for 2-32 weeks. The histological evaluation of DP-bioactive glass as a bone substitute was also investigated in the study. Porous hydroxyapatite bioceramic was used in the control group and the results were compared with those of DP-bioactive glass. The interface between the DP-bioactive glass and bone tissue examined with SEM-EPMA showed that the bioactive glass formed a reaction layer on the surface within 2 weeks after operation and formed a direct bond with natural bone. The elements contained in the bioactive glass apparently interdiffuse with the living bone and biological hydroxyapatite deposited onto the diffusion area, which was proved by EPMA and TEM. After implantation for over 8 weeks, the DP-bioactive glass was gradually biodegraded and absorbed by the living bone. Histological examination using the optical microscope showed that osteocytes grow into the inside of the DP-bioactive glass and the bioactive glass would be expected to be a part of bone.

Animals↗

Heterogeneous fluoridated hydroxyapatites partially substituted with fluoride ions.

The partially substituted fluoridated apatites H-FHAp0.5 and FH-HAp0.5 were synthesized heterngeneously by supplying fluoride at a concentration equivalent to half of the maximum value of fluorapatite during the initial or final half of the experimental period. Although X-ray diffraction patterns and SEM photos of H-FHAp0.5 and FH-HAp0.5 apatites were not significantly different, high-resolution electron microscopy showed quite different features; H-FHAp0.5 type apatite was an elongated hexagon with electron beam damage in the core, and FH-HAp0.5 type apatite was a rather wide hexagon analogous to fluorapatite. Energy dispersive spectroscopy attached to the SEM showed H-FHAp0.5 apatite to have a higher fluoride peak intensity than FH-HAp0.5. These results supported the previous speculations on the two different types of heterogeneous fluoridated hydroxyapatites: hydroxyapatite covered with fluorapatite and fluorapatite coverted with hydroxyapatite.

Crystallization↗

Frontotemporal reconstruction with hydroxyapatite cement and the radial forearm free flap.

BACKGROUND: Contemporary surgical reconstruction of extensive frontal and temporal bony and soft tissue defects requires the use of myocutaneous free flaps. These flaps are associated with donor site morbidity, lengthy operative time, and lack of rigid protection for the brain. To circumvent these problems, we introduce a new surgical technique for reconstruction of frontotemporal defects. OBJECTIVES: To evaluate and discuss hydroxyapatite and radial forearm free flap (RFFF) reconstructive options for patients with soft tissue and bony defects in the frontal and temporal regions after ablative cancer surgery. METHODS: Eight consecutive patients with extensive soft tissue and bony defects in the frontal and temporal regions underwent primary reconstruction with hydroxyapatite cement for replacement of bone, and the RFFF for soft tissue coverage. Patient follow-up ranged from 6 to 48 months. Outcome was determined in terms of immediate and delayed postoperative complications, donor site morbidity, and long-term aesthetic results. RESULTS: Excellent bony and soft tissue contour restoration was achieved in all patients. The bony and soft tissue volume was maintained throughout the follow-up period. One patient had a stroke. No other postoperative complications were encountered. Morbidity from RFFF harvest was minimum. CONCLUSIONS: The combination of hydroxyapatite cement and the RFFF is a viable alternative to the conventional myocutaneous free flap reconstruction of extensive frontotemporal defects. This technique provides excellent aesthetic results, provides rigid protection for the brain, produces minimal and well-tolerated donor site morbidity, and reduces operative time by avoiding intraoperative changes in patient position.

Aged↗

Use of the hydroxyapatite ocular implant in the pediatric population.

OBJECTIVES: There is always a hesitancy to use an ocular implant after enucleation in children, especially in cases of eyes enucleated for retinoblastoma. DESIGN AND SETTING: In this prospective study, we report our experience of 60 consecutive cases in which we used the hydroxyapatite ocular implant after enucleation in children younger than age 10 years. RESULTS: The mean age at the time of enucleation and hydroxyapatite placement was 27 months (range, 1 to 108 months). The histopathologic diagnoses after enucleation included retinoblastoma in 51 patients, endophthalmitis in two patients, persistent hyperplastic primary vitreous in two patients, neovascular glaucoma resulting from Coats' disease in one patient, uveal melanoma in two patients, undifferentiated intraocular neoplasm in one patient, and blind painful traumatized eye in one patient. After a mean follow-up of 19 months (range, 6 to 36 months), two cases of conjunctival erosion and five cases of conjunctival thinning were recorded. These problems were associated with a flat posterior prosthesis vault in all seven cases. There were no cases of orbital cellulitis, implant extrusion, or implant migration. All children showed good small-degree prosthesis motility despite the lack of peg placement. Only one older child underwent drilling and peg placement and she had excellent small- and large-degree prosthesis motility. We generally postpone the drilling and peg placement stage and tedious prosthesis revision in children until they are able to fully cooperate with the ocularist. CONCLUSIONS: The hydroxyapatite ocular implant is well tolerated and provides good prosthesis motility in children with few complications. It is critical for the ocularist to fit the prosthesis comfortably with a high posterior vault.

Child↗

Clinicopathological correlation of technetium bone scan in vascularization of hydroxyapatite implants. A primate model.

OBJECTIVES: To report the histopathological and bone scan characteristics of the stages of hydroxyapatite fibrovascular integration and to consider the implications for the timing of peg drilling in a primate model. DESIGN: Three monkeys received hydroxyapatite implants covered only anteriorly with a fascia lata button to which the rectus muscles were sutured. Weekly bone scans were evaluated quantitatively and qualitatively. The orbits were harvested at 2, 4, and 8 weeks and examined histopathologically. RESULTS: Quantitatively, the implant's technetium uptake increased, then reached a plateau by 4 weeks. Peripheral uptake was present on the images and histologically at 2 weeks. When bone scan images suggested complete vascularization by the fourth week, the implant was 99% vascularized histologically. Completion of vascularization was ascertained at 8 weeks, without further discernible changes in the bone scans. CONCLUSIONS: The technetium bone scan is sensitive to the vascularization of the hydroxyapatite implant and discerns when complete vascularization is approached. This primate study models closely the clinical findings we have recently reported. We advocate at least a 4-week interval between the time the bone scan suggests full vascularization and peg drilling.

Animals↗

Porous bioactive glass and hydroxyapatite ceramic affect bone cell function in vitro along different time lines.

We describe the effects on cell function of treating porous bioactive glass (BG) such that its surface is a composite of carbonated hydroxyapatite and serum protein. The effects on bone cell function of porous hydroxyapatite (HA) ceramic and porous glass treated to become amorphous calcium phosphate only also were studied subsequent to their having adsorbed a serum protein layer. Substrates treated for different durations were seeded with MC3T3-E1 cells and cultured for 3-17 days. Whereas cells seeded on any substrates, BG and HA produced collagen types I and III, bone sialoprotein, and osteopontin, there were significant differences between HA and BG, and among the various surface conditions created on BG. Covering the glass surface with hydroxyapatite and serum protein enhanced expression of high alkaline phosphatase activity, high rates of cell proliferation, and production of mineralized extracellular matrix. The enhancement may be due to the adsorption of a high quantity of fibronectin from the serum onto the reacted bioactive glass surface.

Biocompatible Materials↗

Fatigue properties of hydroxyapatite-coated dental implants after exposure to a periodontal pathogen.

We studied the fatigue properties of rods (4 mm diameter) of hydroxyapatite-coated, titanium alloy implant material after it was exposed to a periodontal pathogen, Actinobacillus actinomycetemcomitans (Aa). We varied the crystallinity of the hydroxyapatite (HA) coating in these rods to the levels of, 60.5%, 52.8%, and 47.8%. Each rod was first inoculated with Aa in the log phase of its growth cycle. After 48 h, we counted the adhered cells. We measured the dissolution of HA coating due to bacterial exposure alone by determining the calcium and phosphate concentrations in the bacterial growth media. Once the adherent bacteria were removed from these rods, we subjected them to 5 million cycles of fatigue testing after immersion in Lactated Ringer's solution. We then determined the calcium and phosphate concentrations in the fatigue media. We found additional coating loss after fatiguing of the samples. This coating loss was a cumulative effect of bacterial exposure and fatigue loading of the hydroxyapatite-coated dental implant alloy. The lower crystallinity sample showed a higher loss of coating within the range of crystallinity studied here. The HA coating in implants during clinical use may undergo such changes, because they are exposed to the same bacteria.

Aggregatibacter actinomycetemcomitans↗

The effect of partial vs. full hydroxyapatite coating on periprosthetic bone quality around the canine madreporic femoral stem.

While hydroxyapatite coatings have been shown to enhance osseointegration of metal prostheses, the optimal extent of the coating over the femoral stem component remains unclear. In this study, we investigated the effect of the application of hydroxyapatite to the proximal half of the femoral stem vs. full hydroxyapatite coating on the madreporic-surfaced femoral stems in a canine model over time. Total hip arthroplasties were performed on 22 dogs. Fifteen had half-HA coated stems, and seven had fully HA coated stems. The dogs were followed for 3 weeks, 3 months, or 1 year postoperatively. Micromotion near the femoral component was assessed at three levels along the medial aspect of the stems. Percent bone apposition, trabecular strut width, connectivity index, and cortical porosity measurements were performed at five levels and in four quadrants about the femoral stems. Micromotion was greater adjacent to the half-coated stems at 3 weeks postoperatively only. There was a trend to decreased micromotion from proximal to distal in both stem types. Both designs fixed proximally and tended to load share laterally more than in the other three quadrants. Fully HA coated stems induced greater percent bone apposition, wider trabecular struts, and more connectivity compared to half-HA coated stems.

Animals↗

Bone bonding in sintered hydroxyapatite combined with a new synthesized agent, TAK-778.

We studied the stimulatory effects of TAK-778, a new synthetic 3-benzothiepin derivative that promotes osteoblast differentiation, in bone bonding to sintered hydroxyapatite implants in rabbit tibiae. Smooth-surfaced rectangular plates (15 x 10 x 2 mm) made of sintered hydroxyapatite were implanted into the proximal metaphyses of bilateral rabbit tibiae, with TAK-778-containing sustained-release microcapsules packed into the medullary cavity in one limb and untreated microcapsules packed in the contralateral limb to serve as a paired control. At 4, 8, and 16 weeks after implantation, bone bonding at the bone-implant interfaces was evaluated by a detaching test and undecalcified histological examination. The tensile failure load increased from 4 to 16 weeks for both groups; however, the tensile failure load of the TAK-778-treated group was significantly greater than that of the control group at each interval after implantation. Histologically, the TAK-778-treated specimens showed greater active new bone formation mainly in the medullary cavity and more extensive bonding between the implant and bone than the untreated specimens. The results of this study suggest that adding osteoinductive TAK-778 to hydroxyapatite implants may significantly accelerate bone apposition to the implants and improve the bonding process at the interface. This would help to establish an earlier and stronger bonding of orthopedic ceramic implants between the surrounding bone tissue.

Absorbable Implants↗

Manipulation of selective cell adhesion and growth by surface charges of electrically polarized hydroxyapatite.

Adherence of cells to a surface, such as a biomaterial surface, can be significantly influenced by the surface charge on that material. The applicability of electrically charged hydroxyapatite ceramics to selective cell adhesion was examined, and we show that polarized hydroxyapatite has significant effects on cell growth and adhesion. The surface charge applied to polarized hydroxyapatite promotes (i) enhanced colony formation of osteoblast-like cells, (ii) activation of gap junctions, and (iii) specific orienting of neuroblastoma cells. These findings will be of great utility and have significance in applications of tissue engineering, for example, in potential treatments for osteoporosis.

Cell Adhesion↗

Purification of a functional gene therapy vector derived from Moloney murine leukaemia virus using membrane filtration and ceramic hydroxyapatite chromatography.

The ability of membrane ultra- and diafiltration and two chromatography media, Matrex Cellufine Sulfate (Millipore) and Macro-Prep ceramic hydroxyapatite (Bio-Rad), to adsorb, elute, and purify gene therapy vectors based on Moloney murine leukaemia virus (MoMuLV) carrying the 4070A amphotropic envelope protein was studied. Membrane ultra- and diafiltration provided virus concentration up to 160-fold with an average recovery of infectious viruses of 77 +/- 14%. In batch experiments, Macro-Prep ceramic hydroxyapatite (type 2, particle size 40 microm) proved superior to Matrex Cellufine Sulfate for MoMuLV vector particle adsorption. Furthermore, functional vector particles could be eluted using phosphate buffer pH 6.8 (highest titres from >or=300 mM phosphate) from the Macro-Prep adsorbent, with higher specific titres (cfu/mg protein) than the starting material. Similar results were obtained when this ceramic hydroxyapatite was packed into a column and used in a liquid chromatography system. Recovery of transduction-competent virus was between 18 and 31% for column experiments and 32 and 46% for batch experiments.

Ceramics↗

Surface and biological evaluation of hydroxyapatite-based coatings on titanium deposited by different techniques.

Hydroxyapatite coatings have been deposited on titanium cp by plasma spray, sol-gel, and sputtering techniques for dental implant applications. The latter two techniques are of current interest, as they allow coatings of micrometer dimensions to be deposited. Coating morphology, composition, and structure have been investigated by scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS) and X-ray diffraction (XRD). All coatings were homogeneous and exhibited a rough morphology suitable for implant applications. The sputtered (after annealing), plasma spray, and sol-gel coatings all showed diffraction peaks corresponding to hydroxyapatite. The surface contaminants were observed to be different for the different coating types. The sputtered coatings were found to have a composition most similar to hydroxyapatite; the sol-gel deposits also showed a high concentration of hydroxyl ions. A discrepancy in the Ca/P ratio was observed for the plasma spray coatings, and a small concentration of carbonate ions was found in the sputter-deposited coatings. The in vitro cell-culture studies using MG63 osteoblast-like cells demonstrated the ability of cells to proliferate on the materials tested. The sol-gel coating promotes higher cell growth, greater alkaline phosphatase activity, and greater osteocalcin production compared to the sputtered and plasma-sprayed coatings.

Alkaline Phosphatase↗

Interaction of chlorhexidine digluconate with and adsorption of chlorhexidine on hydroxyapatite.

It is well known that chlorhexidine digluconate provides an effective microbicidal activity during oral rinsing, and therefore, it was considered worthwhile to investigate its interaction with hydroxyapatite on a fundamental level. The kinetics of uptake (or reaction) of the compound from aqueous solutions by synthetic hydroxyapatite was studied at 23 degrees C for four time periods by monitoring its concentration. There was no uptake at low concentrations for any time period. The uptake curves for higher concentrations shifted towards the lower concentrations as the period increased and became more and more vertically oriented to the concentration axis. The concentrations of calcium ions increased, phosphate ions decreased and hydrogen ions decreased a little for a given period as the concentration of the compound was increased. All of these experimental facts can be qualitatively explained on the basis of the solubility considerations of hydroxyapatite and of chlorhexidine phosphate, the reaction product that slowly precipitates out of the solution. The needle-shaped birefringent crystals of the phosphate salt are clearly visible in the apatite matrix under a microscope, and its refractive index and differential Fourier transform infrared spectra match almost exactly with those of a well-characterized, synthesized phosphate salt. To explore the nature of interaction, the uptake of chlorhexidine base was studied from p-dioxane and it is irreversible. The uptake is total below a threshold equilibrium concentration and constant above it.

Adsorption↗

Formation of hydroxyapatite in water, Hank's solution, and serum at physiological temperature.

The influence of de-ionized water, Hank's saline solution, and bovine calf serum on formation of stoichiometric (Ca/P = 1.67) hydroxyapatite (SHAp) at physiological temperature was studied. SHAp formed in aqueous solution by acid-base reaction of particulate Ca(H(2)PO(4))(2).H(2)O and Ca(4)(PO(4))(2)O. Hydroxyapatite formation is accompanied by an initial period of surface hydration of the precursors, an induction period, and a period during which the bulk of the conversion to hydroxyapatites occurs. The formation of SHAp occurred more rapidly in Hank's solution and distilled water than in serum. The formation of SHAp from these precursors is strongly inhibited by serum. There were two primary exothermal events associated with SHAp formation: initial heat evolution peak, which was associated with reactant dissolution, and the major heat evolution peak, which was associated with SHAp formation. The presence of the constitutents in serum depresses both. This is a result of serum macromolecules adsorbing onto the available surfaces regardless of whether they are reactants or products. Variations in heat evolution behavior, pH, and the times of disappearance of the reactants and appearance of SHAp correlate with one another.

Blood Physiological Phenomena↗