Tytgat Institute for Liver and Intestinal Research, Academic Medical Center (AMC), University of Amsterdam, Postbus 22660, 1100 DD Amsterdam, The Netherlands.
Surgical Laboratory, Academic Medical Center (AMC), University of Amsterdam, Postbus 22660, 1100 DD Amsterdam, The Netherlands.
Mitochondrion. 2018 Mar;39:30-42. doi: 10.1016/j.mito.2017.08.011. Epub 2017 Aug 24.
Human liver cell lines, like HepaRG and C3A, acquire higher functionality when cultured in the AMC-Bio-Artificial Liver (AMC-BAL). The three main differences between BAL and monolayer culture are the oxygenation (40% vs 20%O), dynamic vs absent medium perfusion and 3D vs 2D configuration. Here, we investigated the background of the differences between BAL-cultures and monolayers.
We performed whole-genome microarray analysis on HepaRG monolayer and BAL-cultures. Next, mitochondrial biogenesis was studied in monolayer and BAL-cultures of HepaRG and C3A. The driving forces for mitochondrial biogenesis by BAL-culturing were investigated in representative culture models differing in oxygenation level, medium flow or 2D vs 3D configuration.
Gene-sets related to mitochondrial energy metabolism were most prominently up-regulated in HepaRG-BAL vs monolayer cultures. This was confirmed by a 2.4-fold higher mitochondrial abundance with increased expression of mitochondrial OxPhos complexes. Moreover, the transcript levels of mitochondria-encoded genes were up to 3.6-fold induced and mitochondrial membrane potential activity was 8.3-fold increased in BAL vs monolayers. Culturing with 40% O, dynamic medium flow and/or in 3D increased the mitochondrial abundance and expression of mitochondrial complexes vs standard monolayer culturing. The stimulatory effect of the BAL culture on mitochondrial biogenesis was confirmed in C3A cells in which mitochondrial abundance increased 2.2-fold with induction of mitochondria-encoded genes.
The increased functionality of liver cell lines upon AMC-BAL culturing is associated with increased mitochondrial biogenesis. High oxygenation, medium perfusion and 3D configuration contribute to the up-regulation of the mitochondrial biogenesis.
人源肝细胞系，如 HepaRG 和 C3A，在 AMC-Bio-Artificial Liver（AMC-BAL）中培养时会获得更高的功能。BAL 与单层培养的三个主要区别是氧合作用（40%对 20%O）、动态与无培养基灌注以及 3D 与 2D 结构。在这里，我们研究了 BAL 培养与单层培养之间差异的背景。
我们对 HepaRG 单层和 BAL 培养进行了全基因组微阵列分析。接下来，我们研究了 HepaRG 和 C3A 的单层和 BAL 培养中的线粒体生物发生。在不同氧合水平、培养基流动或 2D 与 3D 结构的代表性培养模型中，研究了 BAL 培养促进线粒体生物发生的驱动力。
与 HepaRG 单层培养相比，BAL 培养中与线粒体能量代谢相关的基因集显著上调。这通过线粒体丰度增加 2.4 倍得到证实，同时还增加了线粒体 OxPhos 复合物的表达。此外，BAL 与单层培养相比，线粒体编码基因的转录水平提高了 3.6 倍，线粒体膜电位活性提高了 8.3 倍。与标准单层培养相比，40%O、动态培养基流动和/或 3D 培养可增加线粒体丰度和线粒体复合物的表达。在 C3A 细胞中，BAL 培养对线粒体生物发生的刺激作用得到了证实，其中线粒体丰度增加了 2.2 倍，同时诱导了线粒体编码基因。
AMC-BAL 培养可增加人源肝细胞系的功能，与线粒体生物发生增加有关。高氧合、培养基灌注和 3D 结构有助于上调线粒体生物发生。
