The Effect of Cyclosporine a On Xenogeneic Moose (Alces Alces) Bone Morphogenetic Protein-Induced Immune Inhibition of Heterotopic Osteoinduction in the Rat Guided Muscle Island Flap Bioassay Model

Abstract

Experimental heterotopic osteoinduction can be induced to take place in a pre-determined intramuscular space using a standard-sized membrane cylinder around a vascular muscle island flap, and implanting an osteoinductive agent such as bone morphogenetic protein (BMP) and demineralized bone matrix (DBM) inside the flap. This kind of newly introduced guided muscle assay method, analyzed by radiomorphometry and histology, was used to study quantitative differences in the osteoinductive activities of xenogenic moose BMP (mBMP) and allogeneic DBM in the Wistar rat, as well as the effect of cyclosporine A (CyA). Calcified tissue in the flaps was quantified by computerized radiomorphometry at 21 days from implantation in five experimental groups: 1) partially purified mBMP; 2)mBMP with CyA treatment; 3) mBMP Plus active allogeneic DBM with CyA; 4) active allogeneic DBM alone; and 5) inactive DBM (control). The results were presented as radiopacity percentages of normal rat femoral bone values (=100). New bone formation was confirmed by microscopy of hematoxylin-eosin and azure II-stained samples. The mean radiopacity, compared to mean rat femoral bone values (=100%), in the mBMP group was 20%. This did not significantly exceed the result in the control flaps with inactive DBM. When CyA was administered perorally to mBMP-implanted animals, the radiopacity increased from 20% to 35% ( p =0.026). When active allogeneic DBM was added to this combination, the increase was up to 53% ( p =0.002). Implantation of active allogeneic DBM alone gave 60%. In microscopy, cartilage was the main sign of tissue transformation in the mBMP-implanted groups without DBM, and new bone with marrow in the active allogeneic DBM-implanted groups. In the control flaps with inactive DBM, no bone or cartilage developed. The quantity of new bone formation with mBMP could be significantly improved with CyA, but not to the superior level induced by active allogeneic DBM. The findings were explained by the marked immunogenicity of xenogeneic partially purified BMP. The advantage of the new model, compared to the conventional intramuscular assays, is a reduction of variability in soft tissue intensities of the measured objects. The standard volume reserved for osteoinduction to take place will make the model also eminently suitable for histomorphometry.