Porous bovine bone mineral in healing of human extraction sockets: 2. Histochemical observations at 9 months

Background: Porous bovine bone mineral (PBBM) has been used in ridge preservation procedures following tooth extractions. The aim of this study was to investigate histochemically tissue sockets grafted with PBBM at 9 months post-extraction. By using different histochemical stainings, characteristics of the newly formed bone; i.e., lamellar/woven ratio at different socket depths, were investigated and the arrangement of bone around the grafted material, as well as the nature of the amorphous organic material found in all specimens, were examined. Methods: After extraction of 15 single-rooted maxillary teeth from 15 patients, socket sites were grafted with PBBM particles (250 to 1000 mu). Primary soft tissue closure of the grafted site was established using the rotated split palatal pedicle flap technique. At 9 months, a cylindrical tissue specimen, 2.5 mm in diameter, was trephined from each previously grafted site followed by placement of a screw-shaped implant. Horizontal tissue section cuts, 5 mu wide, were prepared for histological examination. Histochemical staining; included alcian blue, periodoic-acid Schiff, Mallory trichrome, reticulin, Van Gieson, and picrosirius red (PSR). PSR stained slides were further evaluated morphometrically, using polarized microscopy to determine the amount of lamellar versus woven bone in superficial, mid and deep specimen section cut areas. Results: All staining methods revealed that newly formed bone encircled and adhered to the grafted material in most specimens. Mallory trichrome staining showed osteoblasts present within an osteoid layer, lining the interface zone of PBBM particles and the new osseous tissue. Morphometric evaluation of the PSR stained slides disclosed a constant pattern of increased osseous tissue in a coronal-apical direction. An average of 17.1% osseous tissue with 1:12.9 lamellar/woven bone ratio was calculated in the superficial area. The average bone tissue fraction was 48.3% with a lamellar/woven ratio of 1:3.8 in the mid section area and in the deep area, it increased to 63.9%, with a lamellar/woven ratio average of 1:1.7. Differences between ratios at these sites were statistically significant (P <0.001). An amorphous organic substance was noted in most grafted particles. This material usually attached cell striae and harbored glycoproteins as revealed by periodic-acid Schiff and alcian blue stainings. Mallory trichrome staining showed denatured protein within the decalcified mineral particles; reticulin, Van Gieson stainings, and polarization of PSR stained sections refuted the existence of collagen in the grafted particles. Conclusions: Cancellous PBBM is a biocompatible filler agent in extraction socket sites and an acceptable graft for edentulous ridge preservation at sites prepared to receive endosseous implants. The osteoconductivity of PBBM was determined based on promoting osseous ingrowth and close integration with the newly generated bone. Grafted particles were not significantly resorbed at 9 months. Further studies are needed to determine the resorbable capability, as well as the nature and significance of the amorphous organic substance of PBBM observed in the grafted particles.