In Vivo Characteristics of Premixed Calcium Phosphate Cements When Implanted in Subcutaneous Tissues and Periodontal Bone Defects.

Previous studies showed that water-free, premixed calcium phosphate cements (Pre-CPCs) exhibited longer hardening times and lower strengths than conventional CPCs, but were stable in the package. The materials hardened only after being delivered to a wet environment and formed hydroxyapatite as the only product. Pre-CPCs also demonstrated good washout resistance and excellent biocompatibility when implanted in subcutaneous tissues in rats. The present study evaluated characteristics of Pre-CPCs when implanted in subcutaneous tissues (Study I) and used for repairing surgically created two-wall periodontal defects (Study II). Pre-CPC pastes were prepared by combining CPC powders that consisted of CPC-1: Ca(4)(PO(4))(2)O and CaHPO(4), CPC-2: α-Ca(3)(PO(4))(2) and CaCO(3) or CPC-3: DCPA and Ca(OH)(2) with a glycerol at powder-to-liquid mass ratios of 3.5, 2.5, and 2.5, respectively. In each cement mixture, the Ca to P molar ratio was 1.67. The glycerol contained Na(2)HPO(4) (30 mass %) and hydroxypropyl methylcellulose (0.55 %) to accelerate cement hardening and improve washout resistance, respectively. In Study I, the test materials were implanted subcutaneously in rats. Four weeks after the operation, the animals were sacrificed and histopathological observations were performed. The results showed that all of the implanted materials exhibited very slight or negligible inflammatory reactions in tissues contacted with the implants. In Study II, the mandibular premolar teeth of mature beagle dogs were extracted. One month later, two-wall periodontal bone defects were surgically created adjacent to the teeth of the mandibular bone. The defects were filled with the Pre-CPC pastes and the flaps replaced in the preoperative position. The dogs were sacrificed at 1, 3 and 6 months after surgery and sections of filled defects resected. Results showed that one month after surgery, the implanted Pre-CPC-1 paste was partially replaced by bone and was converted to bone at 6 months. The pockets filled with Pre-CPC-2 were completely covered by newly formed bone in 1 month. The Pre-CPC-2 was partially replaced by trabecular bone in 1 month and was completely replaced by bone in 6 months. Examination of 1 month and 3 month samples indicated that Pre-CPC-2 resorbed and was replaced by bone more rapidly than Pre-CPC 1. Both Pre-CPC pastes were highly osteoconductive. When implanted in periodontal defects, Pre-CPC-2 was replaced by bone more rapidly than Pre-CPC-1.

In Vitro and in Vivo Characteristics of Fluorapatite-Forming Calcium Phosphate Cements.

This study reports for the first time in vitro and in vivo properties of fluorapatite (FA)-forming calcium phosphate cements (CPCs). The experimental cements contained from (0 to 3.1) mass % of F, corresponding to presence of FA at levels of approximately (0 to 87) mass %. The crystallinity of the apatitic cement product increased greatly with the FA content. When implanted subcutaneously in rats, the in vivo resorption rate decreased significantly with increasing FA content. The cement with the highest FA content was not resorbed in soft tissue, making it the first known biocompatible and bioinert CPC. These bioinert CPCs might be useful for applications where slow or no resorption of the implant is required to achieve the desired clinical outcome.

Histological analysis of calcium phosphate bone grafts for surgically created periodontal bone defects in dogs.

A calcium phosphate cement (CPC-1), prepared by mixing an equimolar mixture of tetracalcium phosphate and dicalcium phosphate anhydrous with water, has been shown to be highly biocompatible and osteoconductive. A new type of calcium phosphate cement (CPC-2), prepared by mixing a mixture of alpha-tricalcium phosphate and calcium carbonate with pH 7.4 sodium phosphate solution, was also reported to be highly biocompatible. The objective of the present study was to compare the osteoconductivities of CPC-1 and CPC-2 when implanted in surgically created defects in the jaw bones of dogs. At 1 month after surgery, implanted CPC-1 was partially replaced by new bone and converted to bone within 6 months. In comparison, at 1 month after surgery, the defect filled with CPC-2 was mostly replaced by new bone. Therefore, bone formation in CPC-2-filled pocket was more rapid than in CPC-1-filled pocket. These findings supported the hypothesis that CPC-2 converted to bone more rapidly than CPC-1.

Histopathological and cell enzyme studies of calcium phosphate cements.

New types of self-setting calcium phosphate cement (N-CPC), which do not contain tetracalcium phosphate, were recently developed. N-CPCs harden in 10 minutes with phosphate solution as the cement liquid, and form hydroxyapatite as the set product. The objectives of the present study were to evaluate the biocompatibility (Study I) and cell enzyme activity of N-CPCs and a conventional CPC (Study II). Four experimental cements were tested: (1) dicalcium phosphate anhydrous (DCPA) and calcium oxide; (2) DCPA and calcium hydroxide; (3) tricalcium phosphate and calcium carbonate; and (4) DCPA and tetracalcium phosphate. Phosphate solution was used as the cement liquid for cements (1)-(3), and water for cement (4). Sintered hydroxyapatite particles (5) were used as a control. The test materials were implanted subcutaneously in rats. Four weeks after operation, the animals were sacrificed and histopathological observations were performed. Cements (2) and (3) showed no inflammatory reaction, and were surrounded only by very thin fibrous connective tissues. The histopathological reactions of N-CPCs were nearly identical and were similar to (4) and (5). In addition, effects of alkaline phosphatase (ALP-ase) activity--invoked by the presence of cements (3) and (4)--on osteoblast-like cells derived from dog alveolar bone were also examined because ALP-ase activity is closely related to new bone formation. These results indicated that (3) and (4) were highly compatible with subcutaneous tissues and suggested that these cements may enhance new bone formation.

Hepatocyte growth factor in saliva: a possible marker for periodontal disease status.

Hepatocyte growth factor (HGF) acts as a mitogen, motogen, morphogen and anti-apoptotic factor for various kinds of epithelial cells. We previously showed that periodontal fibroblasts secreted an HGF-like chemoattractant for a gingival epithelial cell line and found that HGF content in gingival crevicular fluid was well correlated with probing depth, gingival index, and interleukin-1beta concentration. To examine whether HGF in whole (mixed) saliva would be a useful marker for periodontal disease status, we investigated the relationship between salivary HGF levels and clinical parameters of 65 adults (50 men and 15 women). Unstimulated whole (mixed) saliva was collected from each subject and the HGF level was determined with an ELISA kit. After sample collection, probing depths and bleeding on probing were monitored. Significant correlations were found between salivary HGF levels and the number of probing depths exceeding 4 mm (r = 0.541), the number of probing depths exceeding 6 mm (r = 0.683), the deepest probing depth of each subject (r = 0.558) and the percentage of sites positive for bleeding on probing (r = 0.511). These results suggest that salivary HGF may be a novel marker for periodontal diagnosis in screening tests.

Histopathologic reaction of a calcium phosphate cement for alveolar ridge augmentation.

The objective of the present study was to evaluate the feasibility of using a calcium phosphate cement (CPC) in the reconstruction of a defective alveolar ridge in conjunction with implant placement. The CPC consisted of an equimolar amount of tetracalcium phosphate and dicalcium phosphate anhydrous. At the beginning of the experiment, all mandibular premolar teeth of mature beagle dogs were extracted. After 1 month of healing, alveolar bone was reduced to make a space for a CPC block that was prefabricated from a CPC mixed with water at a powder/liquid ratio of 5 g/mL. After an additional month, 8-mm long hydroxyapatite-coated titanium implants were placed in such a way that the apical half was embedded into alveolar bone and the coronal half in the preformed CPC block. The dogs were sacrificed and biopsies were obtained at 1, 3, and 6 months after surgery. Sections that included implants were evaluated for integration of the CPC block to the alveolar bone and of the implant to the alveolar bone. Additional sections without the implants served as controls. The results obtained from this study show that the CPC ridge augmentation gradually is replaced by natural bone. Six months after surgery, histopathologic features of the augmentation area were quite similar to those of natural alveolar bone. The coronal half of the implants, previously surrounded by the CPC block, was firmly fixed by natural bone. Therefore, this method may be useful for increasing the height of the alveolar ridge.

Fluorescent labeling analysis and electron probe microanalysis for alveolar ridge augmentation using calcium phosphate cement.

Our previous histopathological study showed that the augmentation block, prepared from a calcium phosphate cement (CPC) mixed with H2O at powder to liquid ratio of 5 g/mL, placed on the alveolar bone ridge, was gradually replaced by natural bone. In the present study, fluorescent labeling analysis (FLA) and electron probe microanalysis (EPMA) were performed on the same surgical site of the above histopathological study. Fluorescent labeling agents, that would be incorporated into newly formed mineralized tissues, were injected into dogs intramuscularly twice a week during the 3 week period that ended 1 week before sacrifice. The specimens obtained from the block were subjected to FLA for assessing the extent of new bone formation and to EPMA for measuring the elemental (Ca, P, Mg) distributions. FLA results showed the presence of newly formed bone at 1 month after surgery. EPMA results showed that the elemental distributions in the augmentation site were similar to those of the residual bone area at 6 months after surgery. FLA and EPMA examinations also indicated that the implants were surrounded and fixed by natural bone chronologically. A CPC augmentation block is clearly useful for alveolar ridge augmentation and osteointegrated implant fixation.