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The induction of endochondral bone formation by transforming growth factor-beta(3): experimental studies in the non-human primate Papio ursinus.

Ripamonti U, Ramoshebi LN, Teare J, Renton L, Ferretti C - J. Cell. Mol. Med. (2008)

Bottom Line: Strikingly and in marked contrast to the rodent bioassay, recombinant human (h)TGF-beta(3), when implanted in the rectus abdominis muscle of adult non-human primates Papio ursinus at doses of 5, 25 and 125 mug per 100 mg of insoluble collagenous matrix as carrier, induces rapid endochondral bone formation resulting in large corticalized ossicles by day 30 and 90.In the same animals, the delivery of identical or higher doses of theTGF-beta(3) protein results in minimal repair of calvarial defects on day 30 with limited bone regeneration across the pericranial aspect of the defects on day 90.RT-PCR, Western and Northern blot analyses of tissue specimens generated by the TGF-beta(3) isoform demonstrate robust expression of Smad-6 and Smad-7 in orthotopic calvarial sites with limited expression in heterotopic rectus abdominis sites.

View Article: PubMed Central - PubMed

Affiliation: Bone Research Unit, Medical Research Council/University of the Witwatersrand, Johannesburg, South Africa. ugo.ripamonti@wits.ac.za

ABSTRACT
Transforming growth factor-beta(3) (TGF-beta(3)), a multi-functional growth modulator of embryonic development, tissue repair and morphogenesis, immunoregulation, fibrosis, angiogenesis and carcinogenesis, is the third mammalian isoform of the TGF-beta subfamily of proteins. The pleiotropism of the signalling proteins of the TGF-beta superfamily, including the TGF-beta proteins per se, are highlighted by the apparent redundancy of soluble molecular signals initiating de novo endochondral bone induction in the primate only. In the heterotopic bioassay for bone induction in the subcutaneous site of rodents, the TGF-beta(3) isoform does not initiate endochondral bone formation. Strikingly and in marked contrast to the rodent bioassay, recombinant human (h)TGF-beta(3), when implanted in the rectus abdominis muscle of adult non-human primates Papio ursinus at doses of 5, 25 and 125 mug per 100 mg of insoluble collagenous matrix as carrier, induces rapid endochondral bone formation resulting in large corticalized ossicles by day 30 and 90. In the same animals, the delivery of identical or higher doses of theTGF-beta(3) protein results in minimal repair of calvarial defects on day 30 with limited bone regeneration across the pericranial aspect of the defects on day 90. Partial restoration of the bone induction cascade by the hTGF-beta(3) protein is obtained by mixing the hTGF-beta(3) device with minced fragments of autogenous rectus abdominis muscle thus adding responding stem cells for further bone induction by the hTGF-beta(3) protein. The observed limited bone induction in hTGF-beta(3)/treated and untreated calvarial defects in Papio ursinus and therefore by extension to Homo sapiens, is due to the influence of Smad-6 and Smad-7 down-stream antagonists of the TGF-beta signalling pathway. RT-PCR, Western and Northern blot analyses of tissue specimens generated by the TGF-beta(3) isoform demonstrate robust expression of Smad-6 and Smad-7 in orthotopic calvarial sites with limited expression in heterotopic rectus abdominis sites. Smad-6 and -7 overexpression in hTGF-beta(3)/treated and untreated calvarial defects may be due to the vascular endothelial tissue of the arachnoids expressing signalling proteins modulating the expression of the inhibitory Smads in pre-osteoblastic and osteoblastic calvarial cell lines controlling the induction of bone in the primate calvarium.

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High power view of a section prepared from a specimen of the 125 μg hTGF-β3 osteogenic device additionally pre-treated with autogenous fragments of rectus abdominis muscle showing the induction of chondrogenesis (lilac arrow) between mineralized bone in blue (light blue arrows) and residual collagenous matrix as carrier (white arrow). The addition of morcellized fragments of autogenous rectus abdominis muscle engineers endochondral bone formation with large islands of chondrogenesis as a recapitulation of embryonic development 90 days after implantation. Undecalcified section cut at 5 μm stained free-floating with Goldner's trichrome. Original magnification ×125.
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fig05: High power view of a section prepared from a specimen of the 125 μg hTGF-β3 osteogenic device additionally pre-treated with autogenous fragments of rectus abdominis muscle showing the induction of chondrogenesis (lilac arrow) between mineralized bone in blue (light blue arrows) and residual collagenous matrix as carrier (white arrow). The addition of morcellized fragments of autogenous rectus abdominis muscle engineers endochondral bone formation with large islands of chondrogenesis as a recapitulation of embryonic development 90 days after implantation. Undecalcified section cut at 5 μm stained free-floating with Goldner's trichrome. Original magnification ×125.

Mentions: On day 90, the addition of minced fragments of autogenous rectus abdominis muscle to the TGF-β3 osteogenic device induced solid blocks of mineralized bone across the defects with newly formed trabeculae covered by osteoid seams facing highly cellular diploic lacunae (Fig. 4). Doses of 25 and 125 μg of the TGF-β3 protein induced mineralized trabeculae of newly formed bone with haematopoietic bone marrow (Figs. 4E, F and G). The addition of autogenous rectus abdominis responding cells also induced large islands of chondrogenic differentiation in treated calvarial defects which were evident on day 90 post-implantation (Fig. 5).


The induction of endochondral bone formation by transforming growth factor-beta(3): experimental studies in the non-human primate Papio ursinus.

Ripamonti U, Ramoshebi LN, Teare J, Renton L, Ferretti C - J. Cell. Mol. Med. (2008)

High power view of a section prepared from a specimen of the 125 μg hTGF-β3 osteogenic device additionally pre-treated with autogenous fragments of rectus abdominis muscle showing the induction of chondrogenesis (lilac arrow) between mineralized bone in blue (light blue arrows) and residual collagenous matrix as carrier (white arrow). The addition of morcellized fragments of autogenous rectus abdominis muscle engineers endochondral bone formation with large islands of chondrogenesis as a recapitulation of embryonic development 90 days after implantation. Undecalcified section cut at 5 μm stained free-floating with Goldner's trichrome. Original magnification ×125.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401141&req=5

fig05: High power view of a section prepared from a specimen of the 125 μg hTGF-β3 osteogenic device additionally pre-treated with autogenous fragments of rectus abdominis muscle showing the induction of chondrogenesis (lilac arrow) between mineralized bone in blue (light blue arrows) and residual collagenous matrix as carrier (white arrow). The addition of morcellized fragments of autogenous rectus abdominis muscle engineers endochondral bone formation with large islands of chondrogenesis as a recapitulation of embryonic development 90 days after implantation. Undecalcified section cut at 5 μm stained free-floating with Goldner's trichrome. Original magnification ×125.
Mentions: On day 90, the addition of minced fragments of autogenous rectus abdominis muscle to the TGF-β3 osteogenic device induced solid blocks of mineralized bone across the defects with newly formed trabeculae covered by osteoid seams facing highly cellular diploic lacunae (Fig. 4). Doses of 25 and 125 μg of the TGF-β3 protein induced mineralized trabeculae of newly formed bone with haematopoietic bone marrow (Figs. 4E, F and G). The addition of autogenous rectus abdominis responding cells also induced large islands of chondrogenic differentiation in treated calvarial defects which were evident on day 90 post-implantation (Fig. 5).

Bottom Line: Strikingly and in marked contrast to the rodent bioassay, recombinant human (h)TGF-beta(3), when implanted in the rectus abdominis muscle of adult non-human primates Papio ursinus at doses of 5, 25 and 125 mug per 100 mg of insoluble collagenous matrix as carrier, induces rapid endochondral bone formation resulting in large corticalized ossicles by day 30 and 90.In the same animals, the delivery of identical or higher doses of theTGF-beta(3) protein results in minimal repair of calvarial defects on day 30 with limited bone regeneration across the pericranial aspect of the defects on day 90.RT-PCR, Western and Northern blot analyses of tissue specimens generated by the TGF-beta(3) isoform demonstrate robust expression of Smad-6 and Smad-7 in orthotopic calvarial sites with limited expression in heterotopic rectus abdominis sites.

View Article: PubMed Central - PubMed

Affiliation: Bone Research Unit, Medical Research Council/University of the Witwatersrand, Johannesburg, South Africa. ugo.ripamonti@wits.ac.za

ABSTRACT
Transforming growth factor-beta(3) (TGF-beta(3)), a multi-functional growth modulator of embryonic development, tissue repair and morphogenesis, immunoregulation, fibrosis, angiogenesis and carcinogenesis, is the third mammalian isoform of the TGF-beta subfamily of proteins. The pleiotropism of the signalling proteins of the TGF-beta superfamily, including the TGF-beta proteins per se, are highlighted by the apparent redundancy of soluble molecular signals initiating de novo endochondral bone induction in the primate only. In the heterotopic bioassay for bone induction in the subcutaneous site of rodents, the TGF-beta(3) isoform does not initiate endochondral bone formation. Strikingly and in marked contrast to the rodent bioassay, recombinant human (h)TGF-beta(3), when implanted in the rectus abdominis muscle of adult non-human primates Papio ursinus at doses of 5, 25 and 125 mug per 100 mg of insoluble collagenous matrix as carrier, induces rapid endochondral bone formation resulting in large corticalized ossicles by day 30 and 90. In the same animals, the delivery of identical or higher doses of theTGF-beta(3) protein results in minimal repair of calvarial defects on day 30 with limited bone regeneration across the pericranial aspect of the defects on day 90. Partial restoration of the bone induction cascade by the hTGF-beta(3) protein is obtained by mixing the hTGF-beta(3) device with minced fragments of autogenous rectus abdominis muscle thus adding responding stem cells for further bone induction by the hTGF-beta(3) protein. The observed limited bone induction in hTGF-beta(3)/treated and untreated calvarial defects in Papio ursinus and therefore by extension to Homo sapiens, is due to the influence of Smad-6 and Smad-7 down-stream antagonists of the TGF-beta signalling pathway. RT-PCR, Western and Northern blot analyses of tissue specimens generated by the TGF-beta(3) isoform demonstrate robust expression of Smad-6 and Smad-7 in orthotopic calvarial sites with limited expression in heterotopic rectus abdominis sites. Smad-6 and -7 overexpression in hTGF-beta(3)/treated and untreated calvarial defects may be due to the vascular endothelial tissue of the arachnoids expressing signalling proteins modulating the expression of the inhibitory Smads in pre-osteoblastic and osteoblastic calvarial cell lines controlling the induction of bone in the primate calvarium.

Show MeSH
Related in: MedlinePlus