Gradually softening hydrogels for modeling hepatic stellate cell behavior during fibrosis regression.
Gradually softening hydrogels for modeling hepatic stellate cell behavior during fibrosis regression.
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DOI：
10.1039/c6ib00027d
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发表时间：
2016-06-13
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影响因子：
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通讯作者：
Burdick JA
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作者：
Caliari SR;Perepelyuk M;Soulas EM;Lee GY;Wells RG;Burdick JA
细胞外基质（ECM）向驻留细胞呈现一组不断演变的机械提示。我们开发了甲基丙烯酸化透明质酸（MeHA）水凝胶，其含有稳定的和水解可降解的交联，以向细胞提供逐渐软化（但不完全可降解）的环境，模仿生理事件，如纤维化消退。为了证明这种细胞培养系统的实用性，我们研究了大鼠肝星状细胞的表型，肝纤维化肌成纤维细胞的主要肝脏前体，在这种软化的环境。在组织培养塑料上机械引发的星状细胞获得肌成纤维细胞表型，其在接种到硬（~20 kPa）水凝胶上时持续存在。然而，在硬-软（~20至~3 kPa）水凝胶上机械引发的星状细胞显示出14天内肌成纤维细胞表型的逆转，与硬水凝胶上的星状细胞相比，细胞面积、肌成纤维细胞标志物α-平滑肌肌动蛋白（α-SMA）表达和具有PDZ结合基序的Yes相关蛋白/转录辅激活因子（雅普/TAZ）核定位减少。然而，硬到软的水凝胶上的细胞没有完全恢复。它们表现出胶质细胞酸性蛋白（GFAP）表达减少，并通过引入额外的交联反应水凝胶的再硬化，经历了异常快速的肌成纤维细胞再活化。这些特征是具有中间表型的星状细胞的典型特征，据报道在体内发生纤维化消退和再损伤。总之，这些数据表明，力学在纤维化消退中起着重要作用，并且将动态力学线索整合到模型系统中有助于捕获体内观察到的细胞行为。
The extracellular matrix (ECM) presents an evolving set of mechanical cues to resident cells. We developed methacrylated hyaluronic acid (MeHA) hydrogels containing both stable and hydrolytically degradable crosslinks to provide cells with a gradually softening (but not fully degradable) milieu, mimicking physiological events such as fibrosis regression. To demonstrate the utility of this cell culture system, we studied the phenotype of rat hepatic stellate cells, the major liver precursors of fibrogenic myofibroblasts, within this softening environment. Stellate cells that were mechanically primed on tissue culture plastic attained a myofibroblast phenotype, which persisted when seeded onto stiff (~ 20 kPa) hydrogels. However, mechanically primed stellate cells on stiff-to-soft (~ 20 to ~ 3 kPa) hydrogels showed reversion of the myofibroblast phenotype over 14 days, with reductions in cell area, expression of the myofibroblast marker alpha-smooth muscle actin (α-SMA), and Yes-associated protein/Transcriptional coactivator with PDZ-binding motif (YAP/TAZ) nuclear localization when compared to stellate cells on stiff hydrogels. Cells on stiff-to-soft hydrogels did not fully revert, however. They displayed reduced expression of glial fibrillary acidic protein (GFAP), and underwent abnormally rapid re-activation to myofibroblasts in response to re-stiffening of the hydrogels through introduction of additional crosslinks. These features are typical of stellate cells with an intermediate phenotype, reported to occur in vivo with fibrosis regression and re-injury. Together, these data suggest that mechanics play an important role in fibrosis regression and that integrating dynamic mechanical cues into model systems helps capture cell behaviors observed in vivo.