Department of Orthopedic Surgery, Physical Medicine and Rehabilitation, University Hospital Grosshadern, Ludwig-Maximilians-University Munich, Marchioninistr. 15, 81377, Munich, Germany.
Sirion Biotech GmbH, Am Klopferspitz 19, 82152, Martinsried, Germany.
Gene Ther. 2019 Feb;26(1-2):16-28. doi: 10.1038/s41434-018-0047-2. Epub 2018 Oct 27.
Bone can be engineered in vivo by implantation of gene-activated muscle tissue fragments. This expedited approach may be further improved by use of muscle tissue with attached fascia. The aim of this in vitro study was to provide an in depth comparison of the osteogenic differentiation capacity of muscle alone and muscle with fascia after BMP-2 transduction. Skeletal muscle tissue from rats was cut into pieces with and without a fascia layer on the surface. Adenoviral BMP-2 or GFP vectors were used for transduction. Osteogenic differentiation within the tissue fragments was evaluated and compared by qRT-PCR, alizarin red S staining, histomorphometry and immunohistology. Transduction efficiency and level of transgene expression were higher for muscle with fascia than muscle alone. Transduction with BMP-2 led to a significant upregulation of bone marker genes, proteins, and calcium deposition in both groups. Interestingly, histological evaluation revealed that osteoinduction did not occur within the fascia layer itself. The upregulation of bone marker genes in muscle with fascia was significantly lower after 2 weeks but similar after 4 weeks of in vitro culture in comparison to muscle alone. The fascia layer led to higher transduction efficiency and enhanced BMP-2 expression. Despite fascia's lower capacity for osteogenic differentiation, muscle implants may benefit from the fascia layer by the improved ability to deliver BMP-2. The presented data may contribute to the development of a novel, cost-effective, single-surgery bone engineering technology and encourage the evaluation of the osteoregenerative potential of muscle with fascia in an animal model.
通过植入基因激活的肌肉组织片段，可以在体内构建骨骼。通过使用附有筋膜的肌肉组织，这种加速方法可能会进一步得到改善。本体外研究的目的是深入比较 BMP-2 转导后单独的肌肉和附有筋膜的肌肉的成骨分化能力。将大鼠的骨骼肌组织切成带有和不带有表面筋膜层的小块。使用腺病毒 BMP-2 或 GFP 载体进行转导。通过 qRT-PCR、茜素红 S 染色、组织形态计量学和免疫组织化学评估和比较组织碎片内的成骨分化。与单独的肌肉相比，附有筋膜的肌肉的转导效率和转基因表达水平更高。转导 BMP-2 导致两组的骨标记基因、蛋白质和钙沉积水平均显著上调。有趣的是，组织学评估显示，成骨诱导并未发生在筋膜层本身。与单独的肌肉相比，在体外培养 2 周后，附有筋膜的肌肉中的骨标记基因的上调显著降低，但在 4 周后相似。筋膜层导致更高的转导效率和增强的 BMP-2 表达。尽管筋膜层的成骨分化能力较低，但肌肉植入物可能受益于筋膜层，因为其能够更好地递送 BMP-2。所提供的数据可能有助于开发一种新型、经济高效、单次手术的骨工程技术，并鼓励在动物模型中评估附有筋膜的肌肉的成骨再生潜力。
