Institute of Biotechnology, National Taiwan University, Taipei, Taiwan.
Department of Orthopaedic Surgery, Chang Gung Memorial Hospital, Taoyuan, Taiwan.
PLoS One. 2019 Jul 18;14(7):e0215499. doi: 10.1371/journal.pone.0215499. eCollection 2019.
Cells, scaffolds, and factors are the triad of regenerative engineering; however, it is difficult to distinguish whether cells in the regenerative construct are from the seeded cells or host cells via the host blood supply. We performed a novel in vivo study to transplant enhanced green fluorescent pig mesenchymal stem cells (EGFP-pMSCs) into calvarial defect of DsRed pigs. The cell distribution and proportion were distinguished by the different fluorescent colors through the whole regenerative period.
METHOD/RESULTS: Eight adult domestic Ds-Red pigs were treated with five modalities: empty defects without scaffold (group 1); defects filled only with scaffold (group 2); defects filled with osteoinduction medium-loaded scaffold (group 3); defects filled with 5 x 103 cells/scaffold (group 4); and defects filled with 5 x 104 cells/scaffold (group 5). The in vitro cell distribution, morphology, osteogenic differentiation, and fluorescence images of groups 4 and 5 were analyzed. Two animals were sacrificed at 1, 2, 3, and 4 weeks after transplantation. The in vivo fluorescence imaging and quantification data showed that EGFP-pMSCs were represented in the scaffolds in groups 4 and 5 throughout the whole regenerative period. A higher seeded cell density resulted in more sustained seeded cells in bone regeneration compared to a lower seeded cell density. Host cells were recruited by seeded cells if enough space was available in the scaffold. Host cells in groups 1 to 3 did not change from the 1st week to 4th week, which indicates that the scaffold without seeded cells cannot recruit host cells even when enough space is available for cell ingrowth. The histological and immunohistochemical data showed that more cells were involved in osteogenesis in scaffolds with seeded cells.
Our in vivo results showed that more seeded cells recruit more host cells and that both cell types participate in osteogenesis. These results suggest that scaffolds without seeded cells may not be effective in bone transplantation.
细胞、支架和因子是再生工程的三要素；然而，通过宿主血液供应，很难区分再生结构中的细胞是来自种子细胞还是宿主细胞。我们进行了一项新的体内研究，将增强型绿色荧光猪间充质干细胞（EGFP-pMSCs）移植到 DsRed 猪的颅骨缺损中。通过整个再生期不同荧光颜色的区分，可以区分细胞的分布和比例。
方法/结果：对 8 头成年家猪 Ds-Red 进行了 5 种方式的处理：无支架的空缺陷（第 1 组）；仅填充支架的缺陷（第 2 组）；填充负载成骨诱导介质的支架的缺陷（第 3 组）；填充 5×103 个细胞/支架的缺陷（第 4 组）；和填充 5×104 个细胞/支架的缺陷（第 5 组）。分析了第 4 组和第 5 组的细胞体外分布、形态、成骨分化和荧光图像。移植后第 1、2、3 和 4 周处死 2 只动物。体内荧光成像和定量数据显示，整个再生期第 4 组和第 5 组的 EGFP-pMSCs 均存在于支架中。与低细胞密度相比，较高的种子细胞密度可导致骨再生中更多持续的种子细胞。如果支架中有足够的空间，种子细胞可募集宿主细胞。第 1 组至第 3 组的宿主细胞从第 1 周到第 4 周没有变化，这表明即使有足够的细胞生长空间，没有种子细胞的支架也不能募集宿主细胞。组织学和免疫组织化学数据显示，支架中存在种子细胞可促进更多细胞参与成骨。
我们的体内结果表明，更多的种子细胞可募集更多的宿主细胞，并且这两种细胞类型均参与成骨。这些结果表明，没有种子细胞的支架在骨移植中可能无效。
