Cell-Laden Agarose-Collagen Composite Hydrogels for Mechanotransduction Studies
Cell-Laden Agarose-Collagen Composite Hydrogels for Mechanotransduction Studies
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DOI：
10.3389/fbioe.2020.00346
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发表时间：
2020-04-21
影响因子：
5.7
通讯作者：
Wuertz-Kozak, Karin
中科院分区：
文献类型：
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作者：
Cambria, Elena;Brunner, Silvio;Wuertz-Kozak, Karin
随着对细胞力学传导机制研究的不断深入，对生物材料的要求越来越高。琼脂糖是软骨和椎间盘机械生物学研究的标准生物材料，但缺乏粘附基序和机械转导所必需的细胞-基质相互作用。在这里，胶原蛋白I型共混在两个浓度（2和4.5毫克/毫升）与琼脂糖2%重量/体积。复合水凝胶的结构均匀性，流变性能和尺寸稳定性方面进行了表征。进一步测试髓核（NP）细胞活力、增殖、形态、基因表达、GAG产生、粘附和机械转导能力。共混水凝胶呈现两种聚合物的均匀网络。虽然加入4.5 mg/mL胶原蛋白显着降低了储能模量，并增加了凝胶的损耗模量，含有2 mg/mL胶原蛋白的混合凝胶显示出类似的机械性能琼脂糖。对于所有基于琼脂糖的凝胶，水凝胶尺寸保持超过21天。包埋的细胞是活的（>80%），并且呈现出减少的增殖和体内NP细胞典型的圆形形态。从第1天到第7天，混合水凝胶中I型和II型胶原蛋白以及聚集蛋白聚糖的基因表达显著增加，进一步导致第7天与琼脂糖凝胶相比显著上级GAG/DNA比率。琼脂糖-胶原水凝胶不仅促进细胞粘附，相反，琼脂糖凝胶，但也表现出5.36倍高的粘着斑激酶磷酸化（pFAK/β-微管蛋白）时，不压缩，并增加pFAK/FAK值压缩后10分钟。琼脂糖-胶原因此优于琼脂糖，模拟由非纤维基质和胶原组成的天然组织，并允许在高度可重复的系统中探索复杂的加载。
The increasing investigation of cellular mechanotransduction mechanisms requires biomaterials combining biofunctionality and suitable mechanical properties. Agarose is a standard biomaterial for cartilage and intervertebral disc mechanobiology studies, but lacks adhesion motifs and the necessary cell-matrix interaction for mechanotransduction. Here, collagen type I was blended at two concentrations (2 and 4.5 mg/mL) with agarose 2% wt/vol. The composite hydrogels were characterized in terms of structural homogeneity, rheological properties and size stability. Nucleus pulposus (NP) cell viability, proliferation, morphology, gene expression, GAG production, adhesion and mechanotransduction ability were further tested. Blended hydrogels presented a homogenous network of the two polymers. While the addition of 4.5 mg/mL collagen significantly decreased the storage modulus and increased the loss modulus of the gels, blended gels containing 2 mg/mL collagen displayed similar mechanical properties to agarose. Hydrogel size was conserved over 21 days for all agarose-based gels. Embedded cells were viable (>80%) and presented reduced proliferation and a round morphology typical of NP cells in vivo. Gene expression of collagen types I and II and aggrecan significantly increased in blended hydrogels from day 1 to 7, further resulting in a significantly superior GAG/DNA ratio compared to agarose gels at day 7. Agarose-collagen hydrogels not only promoted cell adhesion, contrary to agarose gels, but also showed a 5.36-fold higher focal adhesion kinase phosphorylation (pFAK/beta-tubulin) when not compressed, and increased pFAK/FAK values 10 min after compression. Agarose-collagen thus outperforms agarose, mimics native tissues constituted of non-fibrillar matrix and collagens, and allows exploring complex loading in a highly reproducible system.