Center for Engineering in Medicine/Surgical Services, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
PLoS One. 2011;6(5):e20137. doi: 10.1371/journal.pone.0020137. Epub 2011 May 16.
The elucidation of the effect of extracellular matrices on hepatocellular metabolism is critical to understand the mechanism of functional upregulation. We have developed a system using natural extracellular matrices [Adipogel] for enhanced albumin synthesis of rat hepatocyte cultures for a period of 10 days as compared to collagen sandwich cultures. Primary rat hepatocytes isolated from livers of female Lewis rats recover within 4 days of culture from isolation induced injury while function is stabilized at 7 days post-isolation. Thus, the culture period can be classified into three distinct stages viz. recovery stage [day 0-4], pre-stable stage [day 5-7] and the stable stage [day 8-10]. A Metabolic Flux Analysis of primary rat hepatocytes cultured in Adipogel was performed to identify the key metabolic pathways modulated as compared to collagen sandwich cultures. In the recovery stage [day 4], the collagen-soluble Adipogel cultures shows an increase in TriCarboxylic Acid [TCA] cycle fluxes; in the pre-stable stage [day 7], there is an increase in PPP and TCA cycle fluxes while in the stable stage [day 10], there is a significant increase in TCA cycle, urea cycle fluxes and amino acid uptake rates concomitant with increased albumin synthesis rate as compared to collagen sandwich cultures throughout the culture period. Metabolic analysis of the collagen-soluble Adipogel condition reveals significantly higher transamination reaction fluxes, amino acid uptake and albumin synthesis rates for the stable vs. recovery stages of culture. The identification of metabolic pathways modulated for hepatocyte cultures in presence of Adipogel will be a useful step to develop an optimization algorithm to further improve hepatocyte function for Bioartificial Liver Devices. The development of this framework for upregulating hepatocyte function in Bioartificial Liver Devices will facilitate the utilization of an integrated experimental and computational approach for broader applications of Adipogel in tissue e engineering and regenerative medicine.
阐明细胞外基质对肝细胞代谢的影响对于理解功能上调的机制至关重要。我们开发了一种使用天然细胞外基质[Adipogel]的系统，用于增强大鼠肝细胞培养物在 10 天内的白蛋白合成，与胶原夹层培养物相比。从雌性 Lewis 大鼠肝脏中分离的原代大鼠肝细胞在分离诱导损伤后 4 天内恢复，而在分离后 7 天内功能稳定。因此，培养期可分为三个不同阶段，即恢复阶段[第 0-4 天]、预稳定阶段[第 5-7 天]和稳定阶段[第 8-10 天]。对在 Adipogel 中培养的原代大鼠肝细胞进行代谢通量分析，以鉴定与胶原夹层培养物相比被调节的关键代谢途径。在恢复阶段[第 4 天]，胶原可溶性 Adipogel 培养物中三羧酸[TCA]循环通量增加；在预稳定阶段[第 7 天]，PPP 和 TCA 循环通量增加，而在稳定阶段[第 10 天]，TCA 循环、尿素循环通量和氨基酸摄取率显著增加，同时与胶原夹层培养物相比，整个培养期的白蛋白合成速率增加。胶原可溶性 Adipogel 条件下的代谢分析表明，稳定期相对于恢复期的转氨基反应通量、氨基酸摄取和白蛋白合成率显著升高。在 Adipogel 存在下培养肝细胞的代谢途径的鉴定将是开发优化算法以进一步提高生物人工肝脏设备中肝细胞功能的有用步骤。在生物人工肝脏设备中上调肝细胞功能的这一框架的发展将促进综合实验和计算方法的利用，以更广泛地应用于组织工程和再生医学中的 Adipogel。
