School of Biomedical Engineering, Dalhousie University, Halifax, NS, Canada.
Department of Pathology, Dalhousie University, Halifax, NS, Canada.
J Mater Chem B. 2020 Nov 4;8(42):9718-9733. doi: 10.1039/d0tb01570a.
Cells reside in vivo within three dimensional environments in which they interact with extracellular matrices (ECMs) that play an integral role in maintaining tissue homeostasis and preventing tumour growth. Thus, tissue culture approaches that more faithfully reproduce these interactions with the ECM are needed to study cancer development and progression. Many materials exist for modeling tissue environments, and the effects of differing mechanical, physical, and biochemical properties of such materials on cell behaviour are often intricately coupled and difficult to tease apart. Here, an optimized protocol was developed to generate low reaction volume disulfide-crosslinked hyaluronic acid (HA) hydrogels for use in cell culture applications to relate the properties of ECM materials to cell signalling and behaviour. Mechanically, HA hydrogels are comparable to other soft hydrogel materials such as Matrigel and agarose or to tissues lacking type I collagen and other fibrillar ECM components. The diffusion of soluble materials in these hydrogels is affected by unique mass transfer properties. Specifically, HA hydrogel concentration affects the diffusion of anionic particles above 500 kDa, whereas diffusion of smaller particles appears unimpeded by HA content, likely reflecting hydrogel pore size. The HA hydrogels have a strong exclusion effect that limits the movement of proteins into and out of the material once fully formed. Such mass transfer properties have interesting implications for cell culture, as they ultimately affect access to nutrients and the distribution of signalling molecules, affecting nutrient sensing and metabolic activity. The use of disulfide-crosslinked HA hydrogels for the culture of the model prostate cancer cell lines PC3 and LNCaP reveals correlations of protein activation linked to metabolic flux, which parallel and can thus potentially provide insights into cell survival mechanisms in response to starvation that occurs in cancer cell microenvironments.
细胞存在于三维环境中，与细胞外基质（ECM）相互作用，ECM 在维持组织内稳态和防止肿瘤生长方面起着重要作用。因此，需要更真实地模拟这些与 ECM 相互作用的组织培养方法来研究癌症的发展和进展。有许多材料可用于模拟组织环境，而这些材料的机械、物理和生化特性的差异对细胞行为的影响往往错综复杂，难以区分。在这里，开发了一种优化的方案来生成低反应体积的二硫键交联透明质酸（HA）水凝胶，用于细胞培养应用，以将 ECM 材料的特性与细胞信号转导和行为联系起来。在机械性能方面，HA 水凝胶可与其他软质水凝胶材料（如 Matrigel 和琼脂糖）或缺乏 I 型胶原和其他纤维 ECM 成分的组织相媲美。这些水凝胶中可溶性物质的扩散受独特的质量传递特性的影响。具体来说，HA 水凝胶的浓度会影响阴离子颗粒在 500 kDa 以上的扩散，而较小颗粒的扩散似乎不受 HA 含量的阻碍，这可能反映了水凝胶孔径。HA 水凝胶具有很强的排斥效应，一旦完全形成，就会限制蛋白质在材料内外的移动。这种质量传递特性对细胞培养具有有趣的意义，因为它最终会影响营养物质的获取和信号分子的分布，从而影响营养物质的感知和代谢活性。使用二硫键交联的 HA 水凝胶培养模型前列腺癌细胞系 PC3 和 LNCaP 揭示了与代谢通量相关的蛋白质激活的相关性，这与细胞在癌症细胞微环境中发生饥饿时的存活机制平行，因此可能提供深入了解。
