Simultaneous gene transfer of bone morphogenetic protein (BMP)-2 and BMP-7 by in vivo electroporation induces rapid bone formation and BMP-4 expression
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概要
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<p><b>Background:</b> Transcutaneous in vivo electroporation is expected to be an effective gene-transfer method for promoting bone regeneration using the BMP-2 plasmid vector. To promote enhanced osteoinduction using this method, we simultaneously transferred cDNAs for BMP-2 and BMP-7, as inserts in the non-viral vector pCAGGS.<br /><b>Methods:</b> First, an in vitro study was carried out to confirm the expression of BMP-2 and BMP-7 following the double-gene transfer. Next, the individual BMP-2 and BMP-7 plasmids or bothtogether were injected into rat calf muscles, and transcutaneous electroporation was applied 8 times at 100 V, 50 msec.<br /><b>Results:</b> In the culture system, the simultaneous transfer of the BMP-2 and BMP-7 genes led to a much higher ALP activity in C2C12 cells than did the transfer of either gene alone. In vivo, ten days after the treatment, soft X-ray analysis showed that muscles that received both pCAGGS-BMP-2 and pCAGGS-BMP-7 had better-defined opacities than those receiving a single gene. Histological examination showed advanced ossification in calf muscles that received the double-gene transfer.BMP-4 mRNA was also expressed, and RT-PCR showed that its level increased for 3 days in a timedependent manner in the double-gene transfer group. Immunohistochemistry confirmed that BMP-4-expressing cells resided in the matrix between muscle fibers.<br /><b>Conclusion:</b> The simultaneous transfer of BMP-2 and BMP-7 genes using in vivo electroporation induces more rapid bone formation than the transfer of either gene alone, and the increasedexpression of endogenous BMP-4 suggests that the rapid ossification is related to the induction ofBMP-4.</p>
- 2006-08-03