Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgical Engineering, Universitat Politècnica de Catalunya (UPC), Av. Eduard Maristany 16, 08019 Barcelona, Spain; Barcelona Research Centre in Multiscale Science and Engineering, Universitat Politècnica de Catalunya (UPC), Av. Eduard Maristany 16, 08019 Barcelona, Spain.
RMS Foundation, Bischmattstrasse 12, 2544 Bettlach, Switzerland.
Acta Biomater. 2018 Aug;76:319-332. doi: 10.1016/j.actbio.2018.06.025. Epub 2018 Jun 20.
Biomaterials can interact with cells directly, that is, by direct contact of the cells with the material surface, or indirectly, through soluble species that can be released to or uptaken from the surrounding fluids. However, it is difficult to characterise the relevance of this fluid-mediated interaction separately from the topography and composition of the substrate, because they are coupled variables. These fluid-mediated interactions are amplified in the case of highly reactive calcium phosphates (CaPs) such as biomimetic calcium deficient hydroxyapatite (CDHA), particularly in static in vitro cultures. The present work proposes a strategy to decouple the effect of ion exchange from topographical features by adjusting the volume ratio between the cell culture medium and biomaterial (V/V). Increasing this ratio allowed mitigating the drastic ionic exchanges associated to the compositional changes experienced by the material exposed to the cell culture medium. This strategy was validated using rat mesenchymal stem cells (rMSCs) cultured on CDHA and beta-tricalcium phosphate (β-TCP) discs using different V/V ratios. Whereas in the case of β-TCP the cell response was not affected by this ratio, a significant effect on cell adhesion and proliferation was found for the more reactive CDHA. The ionic exchange, produced by CDHA at low V/V, altered cell adhesion due to the reduced number of focal adhesions, caused cell shrinkage and further rMCSs apoptosis. This was mitigated when using a high V/V, which attenuated the changes of calcium and phosphate concentrations in the cell culture medium, resulting in rMSCs spreading and a viability over time. Moreover, rMSCs showed an earlier expression of osteogenic genes on CDHA compared to sintered β-TCP when extracellular calcium fluctuations were reduced.
Fluid mediated interactions play a significant role in the bioactivity of calcium phosphates. Ionic exchange is amplified in the case of biomimetic hydroxyapatite, which makes the in vitro characterisation of cell-material interactions especially challenging. The present work proposes a novel and simple strategy to explore the mechanisms of interaction of biomimetic and sintered calcium phosphates with mesenchymal stem cells. The effects of topography and ion exchange are analysed separately by modifying the volume ratio between cell culture medium and biomaterial. High ionic fluctuations interfered in the maturation of focal adhesions, hampering cell adhesion and leading to increased apoptosis and reduced proliferation rate.
生物材料可以直接与细胞相互作用，即细胞直接与材料表面接触，或者间接通过可以从周围流体中释放或吸收的可溶性物质相互作用。然而，由于它们是耦合变量，因此很难单独描述这种流体介导的相互作用与基底的形貌和组成的相关性。这些流体介导的相互作用在高度反应性的钙磷酸盐（CaP）（如仿生钙缺乏羟磷灰石（CDHA））的情况下会被放大，特别是在静态的体外培养中。本工作提出了一种通过调整细胞培养基与生物材料的体积比（V/V）来分离离子交换与形貌特征之间相关性的策略。增加该比例可以减轻与材料暴露于细胞培养基时经历的组成变化相关的剧烈离子交换。该策略使用不同 V/V 比在 CDHA 和β-磷酸三钙（β-TCP）盘上培养大鼠间充质干细胞（rMSCs）进行了验证。对于β-TCP，细胞反应不受该比例影响，但对于更具反应性的 CDHA，细胞粘附和增殖受到显著影响。由于焦点粘连数量减少，CDHA 产生的离子交换改变了细胞粘附，导致细胞收缩和进一步的 rMCSs 凋亡。当使用高 V/V 时，这一点得到了缓解，高 V/V 降低了细胞培养基中钙和磷酸盐浓度的变化，从而使 rMSCs 扩散并随着时间的推移保持活力。此外，当减少细胞外钙波动时，与烧结的β-TCP 相比，rMSCs 在 CDHA 上更早地表达成骨基因。
流体介导的相互作用在钙磷酸盐的生物活性中起着重要作用。在仿生羟磷灰石的情况下，离子交换被放大，这使得钙磷酸盐与间充质干细胞相互作用的体外特性分析具有特别大的挑战性。本工作提出了一种新颖而简单的策略，用于探索仿生和烧结钙磷酸盐与间充质干细胞相互作用的机制。通过改变细胞培养基与生物材料的体积比，可以分别分析形貌和离子交换的影响。高离子波动干扰了焦点粘连的成熟，阻碍了细胞粘附，并导致细胞凋亡增加和增殖率降低。
