title: Tissue segregation restores the induction of bone formation by the mammalian transforming growth factor-β(3) in calvarial defects of the non-human primate Papio ursinus.
abstract: A diffusion molecular hypothesis from the dura and/or the leptomeninges below that would control the induction of calvarial membranous bone formation by the recombinant human transforming growth factor-β3 (hTGF-β3) was investigated. Coral-derived calcium carbonate-based macroporous constructs (25 mm diameter; 3.5/4 mm thickness) with limited hydrothermal conversion to hydroxyapatite (7% HA/CC) were inserted into forty calvarial defects created in 10 adult Chacma baboons Papio ursinus. In 20 defects, an impermeable nylon foil membrane (SupraFOIL(®)) was inserted between the cut endocranial bone and the underlying dura mater. Twenty of the macroporous constructs were preloaded with hTGF-β3 (125 μg in 1000 μl 20 mM sodium succinate, 4% mannitol pH4.0), 10 of which were implanted into defects segregated by the SupraFOIL(®) membrane, and 10 into non-segregated defects. Tissues were harvested on day 90, processed for decalcified and undecalcified histology and quantitative real-time polymerase chain reaction (qRT-PCR). Segregated untreated macroporous specimens showed a reduction of bone formation across the macroporous spaces compared to non-segregated constructs. qRT-PCR of segregated untreated specimens showed down regulation of osteogenic protein-1 (OP-1), osteocalcin (OC), bone morphogenetic protein-2 (BMP-2), RUNX-2 and inhibitor of DNA binding-2 and -3 (ID2,ID3) and up regulation of TGF-β3, a molecular signalling pathway inhibiting the induction of membranous bone formation. Non-segregated hTGF-β3/treated constructs also showed non-osteogenic expression profiles when compared to non-segregated untreated specimens. Segregated hTGF-β3/treated 7% HA/CC constructs showed significantly greater induction of bone formation across the macroporous spaces and, compared to non-segregated hTGF-β3/treated constructs, showed up regulation of OP-1, OC, BMP-2, RUNX-2, ID2 and ID3. Similar up-regulated expression profiles were seen for untreated non-segregated constructs. TGF-β signalling via ID genes creates permissive or refractory micro-environments that regulate the induction of calvarial bone formation which is controlled by the exogenous hTGF-β3 upon segregation of the calvarial defects. The dura is the common regulator of the induction of calvarial bone formation modulated by the presence or absence of the SupraFOIL(®) membrane with or without hTGF-β3.
authors.affiliation: Bone Research Laboratory, School of Oral Health Sciences, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa. Electronic address: ugo.ripamonti@wits.ac.za.
authors.forename: U
authors.initials: U
authors.lastname: Ripamonti
authors.affiliation: Bone Research Laboratory, School of Oral Health Sciences, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Molecular and Cellular Biology Laboratories, School of Clinical Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Roland Manfred
authors.initials: RM
authors.lastname: Klar
authors.affiliation: Bone Research Laboratory, School of Oral Health Sciences, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Department of Oral Biological Sciences, School of Oral Health Sciences, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Ruqayya
authors.initials: R
authors.lastname: Parak
authors.affiliation: Molecular and Cellular Biology Laboratories, School of Clinical Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Caroline
authors.initials: C
authors.lastname: Dickens
authors.affiliation: Molecular and Cellular Biology Laboratories, School of Clinical Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Therese
authors.initials: T
authors.lastname: Dix-Peek
authors.affiliation: Molecular and Cellular Biology Laboratories, School of Clinical Medicine, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Raquel
authors.initials: R
authors.lastname: Duarte
mesh_terms: D000818:Animals; D037421:Anthozoa / Q000737:chemistry; D018786:Bone Substitutes / Q000737:chemistry*; D002119:Calcium Carbonate / Q000737:chemistry; D017886:Durapatite / Q000737:chemistry; D006801:Humans; D010012:Osteogenesis / Q000187:drug effects*; D048549:Papio ursinus; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000627:therapeutic use; D012886:Skull / Q000187:drug effects* / Q000293:injuries* / Q000473:pathology / Q000502:physiology; D053782:Transforming Growth Factor beta3 / Q000008:administration & dosage / Q000627:therapeutic use*
chemical_list: D018786:Bone Substitutes; D011994:Recombinant Proteins; D053782:Transforming Growth Factor beta3; D017886:Durapatite; D002119:Calcium Carbonate
keywords: Calvarial osteogenesis; Osteogenic proteins; Primates; Redundancy; Tissue segregation; Transforming growth factor-β(3); qRT-PCR