Laboratory of Biology of Stem and Neoplastic Cells, Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, R. Weigla 12, 53-114 Wroclaw, Poland.
Department of Surgery, Faculty of Veterinary Medicine, Wroclaw University of Environmental and Life Sciences, pl. Grunwaldzki 51, 50-366 Wroclaw, Poland.
Int J Mol Sci. 2020 Dec 20;21(24):9726. doi: 10.3390/ijms21249726.
Cell-based therapies using mesenchymal stem cells (MSCs) are a promising tool in bone tissue engineering. Bone regeneration with MSCs involves a series of molecular processes leading to the activation of the osteoinductive cascade supported by bioactive factors, including fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 (BMP-2). In this study, we examined the biological characteristics and osteogenic differentiation potential of sheep bone marrow MSCs (BM-MSCs) treated with 20 ng/mL of FGF-2 and 100 ng/mL BMP-2 in vitro. The biological properties of osteogenic-induced BM-MSCs were investigated by assessing their morphology, proliferation, phenotype, and cytokine secretory profile. The osteogenic differentiation was characterized by Alizarin Red S staining, immunofluorescent staining of osteocalcin and collagen type I, and expression levels of genetic markers of osteogenesis. The results demonstrated that BM-MSCs treated with FGF-2 and BMP-2 maintained their primary MSC properties and improved their osteogenic differentiation capacity, as confirmed by increased expression of osteocalcin and collagen type I and upregulation of osteogenic-related gene markers , , , , , and . Furthermore, sheep BM-MSCs produced a variety of bioactive factors involved in osteogenesis, and supplementation of the culture medium with FGF-2 and BMP-2 affected the secretome profile of the cells. The results suggest that sheep osteogenic-induced BM-MSCs may be used as a cellular therapy to study bone repair in the preclinical large animal model.
基于间充质干细胞（MSCs）的细胞疗法是骨组织工程中一种很有前途的工具。MSC 骨再生涉及一系列分子过程，这些过程导致支持生物活性因子（包括成纤维细胞生长因子-2（FGF-2）和骨形态发生蛋白-2（BMP-2））的成骨诱导级联的激活。在这项研究中，我们研究了用 20ng/ml 的 FGF-2 和 100ng/ml 的 BMP-2 体外处理的绵羊骨髓间充质干细胞（BM-MSCs）的生物学特性和成骨分化潜能。通过评估其形态、增殖、表型和细胞因子分泌谱来研究成骨诱导 BM-MSCs 的生物学特性。通过茜素红 S 染色、骨钙素和 I 型胶原的免疫荧光染色以及成骨相关基因标志物的表达水平来鉴定成骨分化。结果表明，用 FGF-2 和 BMP-2 处理的 BM-MSCs 保持了其原代 MSC 特性并提高了其成骨分化能力，这通过骨钙素和 I 型胶原的表达增加以及成骨相关基因标志物的上调得到证实， ， ， ， ， 。此外，绵羊 BM-MSCs 产生了多种参与成骨的生物活性因子，并且 FGF-2 和 BMP-2 对细胞培养物的补充影响了细胞的分泌组谱。结果表明，绵羊成骨诱导 BM-MSCs 可作为细胞疗法用于研究临床前大动物模型中的骨修复。
