Jean-Claude Petit

Induction of cementogenesis and periodontal ligament regeneration by recombinant human transforming growth factor-β3 in Matrigel with rectus abdominis responding cells

BACKGROUND AND OBJECTIVE In primates and in primates only, the transforming growth factor-b proteins induce endochondral bone formation. Transforming growth factor-b3 also induces periodontal tissue regeneration. Two regenerative treatments using human recombinant transforming growth factor-b3 were examined after implantation in mandibular furcation defects of the nonhuman primate, Papio ursinus. MATERIAL AND METHODS Class III furcation defects were surgically created bilaterally in the mandibular first and second molars of two adult Chacma baboons (P. ursinus). Different doses of recombinant transforming growth factor-beta3 reconstituted with Matrigel matrix were implanted in the rectus abdominis muscle to induce heterotopic ossicles for subsequent transplantation to selected furcation defects. Twenty days after heterotopic implantation, periodontal defects were re-exposed, further debrided and implanted with minced fragments of induced heterotopic ossicles. Contralateral class III furcation defects were implanted directly with recombinant transforming growth factor-beta3 in Matrigel matrix with the addition of minced fragments of autogenous rectus abdominis muscle. Treated quadrants were not subjected to oral hygiene procedures so as to study the effect of the direct application of the recombinant morphogen in Matrigel on periodontal healing. Histomorphometric analyses on undecalcified sections cut from specimen blocks harvested on day 60 measured the area of newly formed alveolar bone and the coronal extension of the newly formed cementum along the exposed root surfaces. RESULTS Morphometric analyses showed greater alveolar bone regeneration and cementogenesis in furcation defects implanted directly with 75 microg of transforming growth factor-beta3 in Matrigel matrix with the addition of minced muscle tissue. CONCLUSION Matrigel matrix is an optimal delivery system for the osteogenic proteins of the transforming growth factor-beta superfamily, including the mammalian transforming growth factor-beta3 isoform. The addition of minced fragments of rectus abdominis muscle provides responding stem cells for further tissue induction and morphogenesis by the transforming growth factor-beta3 protein.

Cementogenesis and the induction of periodontal tissue regeneration by the osteogenic proteins of the transforming growth factor‐β superfamily

The antiquity and severity of periodontal diseases are demonstrated by the hard evidence of alveolar bone loss in gnathic remains of the Pliocene/Pleistocene deposits of the Bloubank Valley at Sterkfontein, Swartkrans and Kromdrai in South Africa. Extant Homo has characterized and cloned a superfamily of proteins which include the bone morphogenetic proteins that regulate tooth morphogenesis at different stages of development as temporally and spatially connected events. The induction of cementogenesis, periodontal ligament and alveolar bone regeneration are regulated by the co-ordinated expression of bone morphogenetic proteins. Naturally derived and recombinant human bone morphogenetic proteins induce periodontal tissue regeneration in mammals. Morphological analyses on undecalcified sections cut at 3-6 mum on a series of mandibular molar Class II and III furcation defects induced in the non-human primate Papio ursinus show the induction of cementogenesis. Sharpeys fibers nucleate as a series of composite collagen bundles within the cementoid matrix in close relation to embedded cementocytes. Osteogenic protein-1 and bone morphogenetic protein-2 possess a structure-activity profile, as shown by the morphology of tissue regeneration, preferentially cementogenic and osteogenic, respectively. In Papio ursinus, transforming growth factor-beta(3) also induces cementogenesis, with Sharpeys fibers inserting into newly formed alveolar bone. Capillary sprouting and invasion determine the sequential insertion and alignment of individual collagenic bundles. The addition of responding stem cells prepared by finely mincing fragments of autogenous rectus abdominis muscle significantly enhances the induction of periodontal tissue regeneration when combined with transforming growth factor-beta(3) implanted in Class II and III furcation defects of Papio ursinus.

Immediate reconstruction of massive cranio-orbito-facial defects with allogeneic and alloplastic matrices in baboons

40 cranio-orbito-facial osseous defects were created in 20 adult male baboons (Papio ursinus) to test the effectiveness of an allogeneic and an alloplastic matrix implant for the functional and morphological repair of the disassembled craniofacial complex. In each animal, one defect was reconstructed with a craniofacial bone segment harvested from donor adult baboons, and processed so as to obtain autolysed antigen-extracted allogeneic (AAA) bone matrix, preserving the bone morphogenetic protein (BMP) activity essential for bone induction. The contralateral defect was implanted with spherical macrobeads of polymethylmethacrylate (PMMA) coated with poly-2-hydroxyethylmethacrylate (PHEMA), and sintered into a porous molded implant, replicating the structural anatomy of the avulsed osseous segment. Histological analysis was carried out on undecalcified and decalcified bone sections prepared from specimens harvested at 3, 6 and 12 months after surgery. In AAA bone, the morphogenetic response was characterized by vascular invasion and mesenchymal cell aggregation after partial resorption of the implanted matrix. This was followed by bone deposition at the osteotomy interfaces and within the medullary spaces of the implanted AAA bone. Although bone ingrowth did occur in some PMMA/PHEMA specimens, the majority of implants showed fibrous union at the recipient interfaces. The limited bone ingrowth may be related to narrow interconnections between larger porous spaces after chemical synthesis of the two polymeric components. Osteogenesis in AAA bone appeared consistent with osteoconductive invasion from the viable bone at the recipient interfaces. In addition, the finding of a delicate trabecular-like bone, appositional to the central areas of the implanted matrix, suggests bone formation by induction.

Tissue segregation enhances calvarial osteogenesis in adult primates.

Calvarial defects of adult mammals have a limited potential for regeneration. Osteogenesis may be impaired by the intrusion within the defect of nonosteogenic tissues during healing, which inhibits centripetal bone deposition from the margins of the defects. Using a primate model, we evaluated bone regeneration in calvarial defects treated with demineralized bone matrix versus defects treated according to the strategy of guided tissue regeneration. Sixty-four defects, 25 mm in diameter, were prepared in 16 adult male baboons (Papio ursinus). In each animal, 2 contralateral defects were covered with a template of porous polymer, 35 mm in diameter. Templates were positioned over the calvarial margins to prevent penetration of the pericranium and the temporalis muscle within the defects, thus maintaining tissue segregation during healing. The third defect was implanted with allogeneic demineralized bone matrix, as positive control. The fourth defect was left untreated and was used as negative control. Undecalcified bone sections (7 microns thick) were prepared from the harvested specimens 3 and 6 months after surgery. Although untreated defects showed limited osteogenesis after fusion of the pericranium with the dura, defects covered with the polymeric template often showed extensive bone deposition extending centripetally from the margins of the craniotomies. Histomorphometry demonstrated that defects treated with demineralized bone matrix and defects covered with the polymeric template had greater amounts of bone and osteoid when compared with untreated defects (p < 0.01). At 6 months, greater amounts of bone formed in demineralized bone matrix-treated defects when compared with defects covered with the polymeric template (p < 0.01). The results of this study show that calvarial membranous bones of adult primates retain the potential to regenerate when segregation between the different tissue components participating in the healing of the wound is maintained. In the same model, however, implantation of allogeneic demineralized bone matrix, used as a positive control provided the most effective treatment for the defects.

Synergistic induction of periodontal tissue regeneration by binary application of human osteogenic protein-1 and human transforming growth factor-β3 in Class II furcation defects of Papio ursinus

BACKGROUND AND OBJECTIVE Binary applications of recombinant human osteogenic protein-1 (hOP-1) and transforming growth factor-β3 (hTGF-β3) synergize to induce pronounced bone formation. To induce periodontal tissue regeneration, binary applications of hOP-1 and hTGF-β(3) were implanted in Class II furcation defects of the Chacma baboon, Papio ursinus. MATERIAL AND METHODS Defects were created bilaterally in the furcation of the first and second mandibular molars of three adult baboons. Single applications of 25 μg hOP-1 and 75 μg hTGF-β(3) in Matrigel(®) matrix were compared with 20:1 binary applications, i.e. 25 μg hOP-1 and 1.25 μg hTGF-β(3). Morcellated fragments of autogenous rectus abdominis striated muscle were added to binary applications. Sixty days after implantation, the animals were killed and the operated tissues harvested en bloc. Undecalcified sections were studied by light microscopy, and regenerated tissue was assessed by measuring volume and height of newly formed alveolar bone and cementum. RESULTS The hOP-1 and hTGF-β(3) induced periodontal tissue regeneration and cementogenesis. Qualitative morphological analysis of binary applications showed clear evidence for considerable periodontal tissue regeneration. Quantitatively, the differences in the histomorphometric values did not reach statistical significance for the group size chosen for this primate study. The addition of morcellated muscle fragments did not enhance tissue regeneration. Binary applications showed rapid expansion of the newly formed bone against the root surfaces following fibrovascular tissue induction in the centre of the treated defects. CONCLUSION Binary applications of hOP-1 and hTGF-β(3) in Matrigel(®) matrix in Class II furcation defects of P. ursinus induced substantial periodontal tissue regeneration, which was tempered, however, by the anatomy of the furcation defect model, which does not allow for the rapid growth and expansion of the synergistic induction of bone formation, particularly when additionally treated with responding myoblastic stem cells.

Induction of cementogenesis and periodontal ligament regeneration by the bone morphogenetic proteins

The complex tissue morphologies of the periodontal tissues, the locking of the teeth into the alveolar bone with the overlying gingival tissues, and the temporo-mandibular joint with the associated powerful masticatory muscles are a superb example of design architecture and engineering.