Heparin hydrogel sandwich cultures of primary hepatocytes
Heparin hydrogel sandwich cultures of primary hepatocytes
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DOI：
10.1016/j.eurpolymj.2014.12.033
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发表时间：
2015-11-01
影响因子：
6
通讯作者：
Revzin, Alexander
中科院分区：
文献类型：
--
作者：
Foster, Elena;You, Jungmok;Revzin, Alexander
原代肝细胞的长期体外维持在肝组织工程中一直是一个挑战，因为这些细胞在标准培养条件下去分化并失去其表型。一些最成功的培养系统采用胶原蛋白或基质。作为一种替代基质，我们的实验室一直在探索使用含肝素的水凝胶作为培养功能性肝细胞的支架。本研究的特点是肝素凝胶夹层培养系统维持原代大鼠肝细胞。采用紫外光诱导硫代肝素（Hep-SH）与二丙烯酸聚乙二醇（PEGDA）前体进行巯基偶联反应制备了肝素凝胶层。肝功能分析显示，在胶原蛋白和肝素凝胶夹层培养系统中，细胞以相似的高水平维持白蛋白分泌至少三周。此外，肝素凝胶的肝细胞色素P450活性比胶原蛋白凝胶高1.6倍。肝素的另一个明显优势是它能够以可控的方式结合和释放生长因子。我们观察到，在含有肝细胞生长因子（HGF）的肝素凝胶三明治中培养的肝细胞在24天后产生的白蛋白量比在胶原三明治中培养的细胞高1.8倍。总的来说，我们证明肝素凝胶三明治是一个很好的系统，维持分化，极化原代肝细胞超过三周。本研究中使用的生物材料肝素-聚乙二醇水凝胶在调节机械性能、调节肝素含量或引入光敏化学物质以促进凝胶降解方面具有显著的灵活性。我们看到这种新系统在肝细胞维持和干细胞分化研究中的多种应用。(C) 2014 Elsevier Ltd.版权所有。
Long-term in vitro maintenance of primary hepatocytes has been a challenge in hepatic tissue engineering, since these cells dedifferentiate and lose their phenotype in standard culture conditions. Some of the most successful culture systems employ collagen or Matrigel matrices. As an alternative matrix, our laboratory has been exploring the use of heparin-containing hydrogels as scaffolds for cultivation of functional hepatocytes. The present study characterized a heparin gel sandwich culture system for maintenance of primary rat hepatocytes. The heparin gel layers were fabricated via UV light induced thiol-ene coupling reaction of thiolated heparin (Hep-SH) and diacrylated poly(ethylene glycol) (PEGDA) precursors. Analysis of hepatic function revealed that cells sustained albumin secretion for at least three weeks at similarly high levels for collagen and heparin gel sandwich culture systems. Furthermore, cytochrome P450 activity of hepatocytes was similar to 1.6 times higher in heparin gels compared to collagen gels. Another distinct advantage of heparin is its ability to bind and release growth factors in a controlled manner. We observed that after 24 days hepatocytes cultured in heparin gel sandwich containing hepatocyte growth factor (HGF) produced similar to 1.8 times higher amounts of albumin compared to cells in collagen sandwiches. Overall, we demonstrate heparin gel sandwich to be an excellent system for maintaining differentiated, polarized primary hepatocytes for over three weeks. The biomaterial used in this study, heparin-PEG hydrogel, offers significant flexibility in terms of tuning mechanical properties, adjusting heparin content or introducing photolabile chemistries to promote gel degradation. We see multiple applications for this novel system in hepatocyte maintenance and stem cell differentiation studies. (C) 2014 Elsevier Ltd. All rights reserved.