Transmembrane bone matrix gelatin-induced differentiation of bone

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Abstract
In response to chemically-defined bone matrix gelatin (BMG) inside a diffusion chamber implanted in a muscle pouch, mesenchymal cells migrate directionally, aggregate and differentiate into new bone, on theoutside of the chamber. BMG diffuses through double membranes 275 to 300 μm in thickness. The inner membrane of pore size is 0.025 μm and the outer membrane of pore size is 0.45 μm. The inner membrane is 1/20 the pore size and the combination is twice the thickness of membranes previously reported to transfer osteoinductive activity of living cells. Autoradiographs show35S-cysteine-labelled BMG produces very high transmembrane grain counts while3H-proline labelled BMG produces very low transmembrane grain counts. Electron micrographs demonstrate that gelatin-derived, uranyl-acetate-stained fine granules interspersed with ruthenium red-staining coarse granules, diffuse through the membrane of 0.025 μm pore size from the inside out. Solitary pale-staining collagen fibrils, possibly formed in interstitial fluid by renaturation of BMG are found in the interior of the chamber and in the interior of the outer 0.45 μm but not the inner 0.025 μm pore size membrane. Densely-stained new bone collagen fiber bundles cover the outer membrane, fill the 0.45 μm subsurface pores for a depth of 0.20 to 30 μm, and thereby attach the new cartilage and bone deposits to the outer surface of the chamber. BMG powders solubilize rapidly in diffusion chambers and produce high yields of new bone. The relationship between denatured collagen and renatured gelatin fibrils in the process of transfer of the bone morphogen from BMG to mesenchymal cell receptors is an intriguing subject for further investigation.