首页> 美国卫生研究院文献>International Journal of Molecular Sciences >A Growth Factor-Free Co-Culture System of Osteoblasts and Peripheral Blood Mononuclear Cells for the Evaluation of the Osteogenesis Potential of Melt-Electrowritten Polycaprolactone Scaffolds
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A Growth Factor-Free Co-Culture System of Osteoblasts and Peripheral Blood Mononuclear Cells for the Evaluation of the Osteogenesis Potential of Melt-Electrowritten Polycaprolactone Scaffolds

机译:成骨细胞和外周血单个核细胞的无生长因子共培养系统用于评估熔体电写聚己内酯支架的成骨潜能

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摘要

Scaffolds made of biodegradable biomaterials are widely used to guide bone regeneration. Commonly, in vitro assessment of scaffolds’ osteogenesis potential has been performed predominantly in monoculture settings. Hence, this study evaluated the potential of an unstimulated, growth factor-free co-culture system comprised of osteoblasts (OB) and peripheral blood mononuclear cells (PBMC) over monoculture of OB as an in vitro platform for screening of bone regeneration potential of scaffolds. Particularly, this study focuses on the osteogenic differentiation and mineralized matrix formation aspects of cells. The study was performed using scaffolds fabricated by means of a melt electrowriting (MEW) technique made of medical-grade polycaprolactone (PCL), with or without a surface coating of calcium phosphate (CaP). Qualitative results, i.e., cell morphology by fluorescence imaging and matrix mineralization by von Kossa staining, indicated the differences in cell behaviours in response to scaffolds’ biomaterial. However, no obvious differences were noted between OB and OB+PBMC groups. Hence, quantitative investigation, i.e., alkaline phosphatase (ALP), tartrate-resistant acid phosphatase (TRAP) activities, and gene expression were quantitatively evaluated by reverse transcription-polymerase chain reaction (RT-qPCR), were evaluated only of PCL/CaP scaffolds cultured with OB+PBMC, while PCL/CaP scaffolds cultured with OB or PBMC acted as a control. Although this study showed no differences in terms of osteogenic differentiation and ECM mineralization, preliminary qualitative results indicate an obvious difference in the cellon-mineralized ECM density between scaffolds cultured with OB or OB+PBMC that could be worth further investigation. Collectively, the unstimulated, growth factor-free co-culture (OB+PBMC) system presented in this study could be beneficial for the pre-screening of scaffolds’ in vitro bone regeneration potential prior to validation in vivo.
机译:由可生物降解的生物材料制成的支架被广泛用于引导骨骼再生。通常,体外评估支架的成骨潜能主要是在单培养条件下进行的。因此,本研究评估了由成骨细胞(OB)和外周血单核细胞(PBMC)组成的无刺激,无生长因子的共培养系统相对于OB的单培养的潜力,以此作为筛选支架骨再生潜力的体外平台。特别地,该研究集中于细胞的成骨分化和矿化基质形成方面。这项研究是通过使用由医用级聚己内酯(PCL)制成的熔融电写(MEW)技术制造的支架进行的,该支架有或没有磷酸钙(CaP)的表面涂层。定性结果,即荧光成像的细胞形态和von Kossa染色的基质矿化,表明了响应支架生物材料的细胞行为的差异。但是,OB组和OB + PBMC组之间没有明显差异。因此,通过逆转录-聚合酶链反应(RT-qPCR)定量评估了碱性磷酸酶(ALP),抗酒石酸酸性磷酸酶(TRAP)活性和基因表达的定量研究,仅评估了PCL / CaP支架。用OB + PBMC培养,而用OB或PBMC培养的PCL / CaP支架充当对照。尽管这项研究没有显示出成骨分化和ECM矿化方面的差异,但初步的定性结果表明,用OB或OB + PBMC培养的支架之间的细胞/非矿化ECM密度存在明显差异,值得进一步研究。总体而言,本研究中提出的无刺激,无生长因子的共培养(OB + PBMC)系统可能有利于在进行体内验证之前对支架的体外骨再生潜力进行预筛选。

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