Whole Genome Gene Expression: Histologic and Immunohistologic Study of Grafted Bone Using Allograft and Xenograft

The aim of this study was to explore the influence of different bone grafts, demineralized freeze-dried bone allograft (DFDBA, OraGraft), and deproteinized bovine bone mineral (DBBM, Bio-Oss) implanted in mouse calvaria defects on gene expression.

Male C57BL/6MLac mice were separated into three groups as follows: group 1-defect without graft as control, group 2-DFDBA, and group 3-DBBM. Affymetrix DNA microarrays were used to characterize gene expression in bone after 3 months of graft healing. Differential expression of designated genes discovered by microarray analysis was confirmed using real-time polymerase chain reaction (PCR) and immunohistochemistry.

Compared with normal bone healing, 355 and 1,108 coding genes of bone grafted with DFDBA were upregulated and downregulated, respectively. The upregulated genes were mainly involved in chemokine signaling, macrophage activity, osteoclast activity, cytokine expression, T-cell receptor signaling, apoptosis, and MAPK signaling. The downregulated genes were predominantly involved in calcium regulation in cardiac cells, chemokine signaling, MAPK signaling, and adipogenesis. A total of 306 and 817 coding genes of bone grafted with DBBM were upregulated and downregulated, respectively. The upregulated genes were mainly involved in osteoclast activity, chemokine signaling, B cell receptor signaling, macrophage activity, and signaling of T-cell receptor, MAPK, IL-5, and IL-1. The downregulated genes were predominantly involved in calcium regulation in the cardiac cell and osteoclast activity. Real-time PCR revealed that the DFDBA and DBBM groups showed a higher mRNA level of MMP12, Bcl2A1, S100A4, and Postn compared with the control (P < .05). Histology showed that, compared with the control, the volume of new bone was higher in both types of bone grafts. Immunohistochemistry using an MMP12 antibody confirmed the microarray results because the MMP12 immunoreactivity intensified, and a positive expression of MMP12 increased significantly in the DFDBA and DBBM groups.

Both DFDBA and DBBM had a gene expression network involved in new bone formation, which coincided with an increased expression of MMP-12 and osteoclast activity. Both types of graft materials appeared to connect with genes that stimulate bone remodeling at 3 months of bone grafting.