Harnessing endogenous signals from hepatocytes using a low volume multi-well plate.
Harnessing endogenous signals from hepatocytes using a low volume multi-well plate.
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DOI：
10.1039/c7ib00010c
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发表时间：
2017-05-22
期刊：
影响因子：
--
通讯作者：
Revzin A
中科院分区：
文献类型：
--
作者：
Gheibi P;Son KJ;Stybayeva G;Revzin A
肝细胞是高度分化的上皮细胞，一旦离开体内环境，就会失去其表型和功能。鉴于肝脏培养在药物毒性、生物人工肝辅助装置和基础生物学研究中的重要性，在体外维持肝功能方面已经付出了相当大的努力。迄今为止使用的方法包括肝细胞与基质细胞的共同培养，将这些细胞组织成球状，并将它们嵌入生物活性凝胶中。我们的团队最近证明，在没有对流的情况下，被限制在微流体通道中的原代大鼠肝细胞通过上调包括肝细胞生长因子（HGF）在内的内源性信号来维持上皮表型。本研究的目的是从微流控装置过渡到生物实验室中普遍存在的小体积多孔板，微流控装置在某种程度上是专业化和具有挑战性的。使用3D打印和微成型的结合，我们已经构建了插入物，可以放置在标准的12孔板中，可以用来创建低容量培养条件下，原代肝细胞保持分化表型。包括白蛋白合成和表达在内的检测证实了这种表型增强。重要的是，我们证实了小体积培养板中HGF的上调，并证明了在我们的细胞培养平台中抑制HGF信号传导会降低肝脏表型。总体而言，本研究概述了一种新的细胞培养系统，该系统利用多孔板格式的微流体通道的低体积效应。除肝细胞外，该系统还可用于维持其他难以培养的细胞，包括干细胞和原发性癌细胞。
Hepatocytes are highly differentiated epithelial cells that lose their phenotype and function when removed from the in vivo environment. Given the importance of hepatic cultures for drug toxicity, bioartificial liver assist devices and basic biology studies, considerable efforts have been focused on maintenance of hepatic function in vitro. The methods used to date include co-cultivation of hepatocytes with stromal cells, organizing these cells into spheroids and imbedding them into bioactive gels. Our team has recently demonstrated that primary rat hepatocytes confined to microfluidic channels in the absence of convection maintained epithelial phenotype through upregulation of endogenous signals including hepatocyte growth factor (HGF). The objective of the present study was to transition from microfluidic devices, which are somewhat specialized and challenging to use and towards low volume multiwell plates ubiquitous in the biology laboratories. Using a combination of 3D printing and micromolding we have constructed inserts that could be placed into standard 12-well plates and could be used to create low volume culture conditions under where primary hepatocytes maintained differentiated phenotype. This phenotype enhancement was confirmed by assays including albumin synthesis and expression. Importantly we confirmed upregulation of HGF inside the low volume culture plates and demonstrated that inhibition of HGF signaling degraded hepatic phenotype in our cell culture platform. Overall, this study outlines a new cell culture system that leverages low volume effects of microfluidic channels in a multiwell plate format. Beyond hepatocytes, such system may be of use in maintenance of other difficult-to-culture cells including stem cells and primary cancer cells.
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DOI：
10.1073/pnas.0906820106
发表时间：
2009-09-15
影响因子：
11.1
作者：
Kidambi, Srivatsan;Yarmush, Rubin S.;Nahmias, Yaakov
通讯作者：
Nahmias, Yaakov
DOI：
10.1073/pnas.1103100109
发表时间：
2012-01-17
影响因子：
11.1
作者：
Przybyla, Laralynne M.;Voldman, Joel
通讯作者：
Voldman, Joel
影响因子：
2.9
作者：
DUNN, JCY;TOMPKINS, RG;YARMUSH, ML
通讯作者：
YARMUSH, ML
影响因子：
5.1
作者：
Thulé, PM;Liu, J;Phillips, LS
通讯作者：
Phillips, LS
DOI：
10.1039/c5ib00105f
发表时间：
2015-07
期刊：
Integrative biology : quantitative biosciences from nano to macro
影响因子：
--
作者：
Patel D;Haque A;Gao Y;Revzin A
通讯作者：
Revzin A