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首页> 外文期刊>Acta biomaterialia >Bilayered chitosan-based scaffolds for osteochondral tissue engineering: influence of hydroxyapatite on in vitro cytotoxicity and dynamic bioactivity studies in a specific double-chamber bioreactor.
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Bilayered chitosan-based scaffolds for osteochondral tissue engineering: influence of hydroxyapatite on in vitro cytotoxicity and dynamic bioactivity studies in a specific double-chamber bioreactor.

机译：用于骨软骨组织工程的基于壳聚糖的双层支架：羟基磷灰石对特定双腔生物反应器中体外细胞毒性和动态生物活性研究的影响。

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Osteochondral tissue engineering presents a current research challenge due to the necessity of combining both bone and cartilage tissue engineering principles. In the present study, bilayered chitosan-based scaffolds are developed based on the optimization of both polymeric and composite scaffolds. A particle aggregation methodology is proposed in order to achieve an improved integrative bone-cartilage interface needed for this application, since any discontinuity is likely to cause long-term device failure. Cytotoxicity was evaluated by the MTS assay with the L929 fibroblast cell line for different conditions. Surprisingly, in composite scaffolds using unsintered hydroxyapatite, cytotoxicity was observed in vitro. This work reports the investigation that was conducted to overcome and explain this behaviour. It is suggest that the uptake of divalent cations may induce the cytotoxic behaviour. Sintered hydroxyapatite was consequently used and showed no cytotoxicity when compared to the controls. Microcomputed tomography (micro-CT) was carried out to accurately quantify porosity, interconnectivity, ceramic content, particle and pore sizes. The results showed that the developed scaffolds are highly interconnected and present the ideal pore size range to be morphometrically suitable for the proposed applications. Dynamical mechanical analysis (DMA) demonstrated that the scaffolds are mechanically stable in the wet state even under dynamic compression. The obtained elastic modulus was, respectively, 4.21+/-1.04, 7.98+/-1.77 and 6.26+/-1.04 MPa at 1 Hz frequency for polymeric, composite and bilayered scaffolds. Bioactivity studies using both a simulated body fluid (SBF) and a simulated synovial fluid (SSF) were conducted in order to assure that the polymeric component for chondrogenic part would not mineralize, as confirmed by scanning electron microscopy (SEM), inductively coupled plasma-optical emission spectroscopy (ICP) and energy-dispersive spectroscopy (EDS) for different immersion periods. The assays were carried out also under dynamic conditions using, for this purpose, a specifically designed double-chamber bioreactor, aiming at a future osteochondral application. It was concluded that chitosan-based bilayered scaffolds produced by particle aggregation overcome any risk of delamination of both polymeric and composite parts designed, respectively, for chondrogenic and osteogenic components that are mechanically stable. Moreover, the proposed bilayered scaffolds could serve as alternative, biocompatible and safe biodegradable scaffolds for osteochondral tissue engineering applications.
机译：骨软骨组织工程提出了当前的研究挑战，这是因为必须将骨骼和软骨组织工程原理结合起来。在本研究中，基于聚合物和复合材料支架的优化开发了基于壳聚糖的双层支架。提出了一种粒子聚集方法，以实现此应用所需的改进的集成式骨-软骨界面，因为任何不连续性都可能导致长期的设备故障。在不同条件下，用L929成纤维细胞系通过MTS试验评估细胞毒性。令人惊讶地，在使用未烧结的羟基磷灰石的复合支架中，在体外观察到细胞毒性。这项工作报告了为克服和解释此行为而进行的调查。提示摄取二价阳离子可能诱导细胞毒性行为。因此，使用烧结的羟基磷灰石，并且与对照相比没有细胞毒性。进行了微型计算机断层扫描（micro-CT），以准确定量孔隙率，互连性，陶瓷含量，颗粒和孔径。结果表明，开发的支架高度互连，并呈现理想的孔径范围，以形态上适合所提出的应用。动态力学分析（DMA）表明，即使在动态压缩下，支架在潮湿状态下也具有机械稳定性。对于聚合物支架，复合支架和双层支架，在1Hz频率下获得的弹性模量分别为4.21 +/- 1.04、7.98 +/- 1.77和6.26 +/- 1.04MPa。为了确保软骨形成部分的聚合物成分不会矿化，使用了模拟体液（SBF）和模拟滑液（SSF）进行了生物活性研究，这一点已通过扫描电子显微镜（SEM）进行了感应耦合等离子体处理。发射光谱（ICP）和能量色散光谱（EDS）用于不同的浸没时间。为此，在动态条件下也使用专门设计的双腔生物反应器进行了测定，目的是将来的骨软骨应用。结论是，通过颗粒聚集生产的基于壳聚糖的双层支架克服了分别设计用于机械稳定的软骨形成和成骨组分的聚合物和复合部件分层的任何风险。而且，提出的双层支架可以用作骨软骨组织工程应用的替代的，生物相容的和安全的可生物降解的支架。

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