Tissue segregation restores the induction of bone formation by the mammalian transforming growth factor-β3 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-β 3 (hTGF-β 3 ) 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-β 3 (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-β 3 , a molecular signalling pathway inhibiting the induction of membranous bone formation. Non-segregated hTGF-β 3 /treated constructs also showed non-osteogenic expression profiles when compared to non-segregated untreated specimens. Segregated hTGF-β 3 /treated 7% HA/CC constructs showed significantly greater induction of bone formation across the macroporous spaces and, compared to non-segregated hTGF-β 3 /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-β 3 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-β 3 .