Research ArticleBone Regeneration

In situ bone tissue engineering via ultrasound-mediated gene delivery to endogenous progenitor cells in mini-pigs

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Science Translational Medicine  17 May 2017:
Vol. 9, Issue 390, eaal3128
DOI: 10.1126/scitranslmed.aal3128

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Bubbles and BMP-6 for bone repair

Treatments for bone nonunions (fractures that fail to heal) include surgery and bone grafting. As an alternative to viral gene delivery, Bez et al. developed a two-step therapy. First, endogenous mesenchymal stem/progenitor cells were recruited to the bone nonunion by implanting a collagen sponge in the defect site. Two weeks later, bone morphogenetic protein-6 (BMP-6) plasmid DNA and microbubbles were injected into nonunions, and ultrasound was applied to oscillate the microbubbles, which helped the recruited progenitors take up the BMP-6. This therapy led to transient BMP-6 secretion, bone regeneration, and fracture healing over 6 weeks in critical-sized tibial nonunions in mini-pigs.

Abstract

More than 2 million bone-grafting procedures are performed each year using autografts or allografts. However, both options carry disadvantages, and there remains a clear medical need for the development of new therapies for massive bone loss and fracture nonunions. We hypothesized that localized ultrasound-mediated, microbubble-enhanced therapeutic gene delivery to endogenous stem cells would induce efficient bone regeneration and fracture repair. To test this hypothesis, we surgically created a critical-sized bone fracture in the tibiae of Yucatán mini-pigs, a clinically relevant large animal model. A collagen scaffold was implanted in the fracture to facilitate recruitment of endogenous mesenchymal stem/progenitor cells (MSCs) into the fracture site. Two weeks later, transcutaneous ultrasound-mediated reporter gene delivery successfully transfected 40% of cells at the fracture site, and flow cytometry showed that 80% of the transfected cells expressed MSC markers. Human bone morphogenetic protein-6 (BMP-6) plasmid DNA was delivered using ultrasound in the same animal model, leading to transient expression and secretion of BMP-6 localized to the fracture area. Micro–computed tomography and biomechanical analyses showed that ultrasound-mediated BMP-6 gene delivery led to complete radiographic and functional fracture healing in all animals 6 weeks after treatment, whereas nonunion was evident in control animals. Collectively, these findings demonstrate that ultrasound-mediated gene delivery to endogenous mesenchymal progenitor cells can effectively treat nonhealing bone fractures in large animals, thereby addressing a major orthopedic unmet need and offering new possibilities for clinical translation.

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