Culturing cells in human serum: differentiation and metabolic reprogramming in human hepatoma cells. Homo sapiens

Huh7.5 cells, which are cultured in human serum containing media instead of FBS containing media, undergo growth arrest and become differentiated. In this study we further characterized human serum cultured Huh7.5 cells, particularly with regards to morphology, metabolic profile and hepatocyte specific functions. Similar to primary hepatocytes, HS-cultured Huh7.5 cells become columnar, are polarized, and develop bile canalicular structures on the apical side of the cells. Further analysis by microarray showed that significant transcriptional changes occur in 22-32% of the genes. HS cultured Huh7.5 cells undergo extensive metabolic re-programming. We showed a reversal of a Warburg-like metabolic profile that is typical for proliferating cells. In HS cultured cells large lipid droplets can be found in addition to large glycogen stores. Metabolic modeling shows an increased reliance on beta-oxidation, which was confirmed by an increase in ketone body production, the end products of β-oxidation. Additionally we showed increased conversion of glucose to glycogen. Bile secretion is increased, as are the enzymes involved in the degradation of xenobiotics. Overall these data show that only changing the serum in cell culture media can result in the establishment of a cell type that morphologically, metabolically and functionally resemble primary hepatocytes. Overall design: Samples were analyzed for Huh7.5 cells cultured in fetal bovine serum as well as at different time points – 8 days, 15 days and 23 days – after being transferred to human serum. Biological triplicates were used in each case.

Huh7.5细胞在含人血清而非胎牛血清（Fetal Bovine Serum, FBS）的培养基中培养时，会发生生长停滞并分化。本研究对人血清培养的Huh7.5细胞展开了深入表征，重点考察其形态学特征、代谢谱及肝细胞特异性功能。
与原代肝细胞（primary hepatocytes）相似，经人血清培养的Huh7.5细胞会转变为柱状形态，出现细胞极化，并在细胞顶侧形成胆小管结构。基因芯片（microarray）分析结果显示，22%~32%的基因发生了显著转录变化。
人血清培养的Huh7.5细胞发生了广泛的代谢重编程：研究发现，增殖细胞典型的沃伯格（Warburg）样代谢谱发生了逆转。该培养条件下的细胞中，除存在大量糖原储存外，还可观察到大型脂滴。代谢模型分析表明，细胞对β氧化（beta-oxidation）的依赖程度增强，这一结论可通过作为β氧化终产物的酮体生成量增加得到验证。此外，葡萄糖向糖原的转化水平亦有所提升。胆汁分泌能力增强，外源性物质（xenobiotics）降解相关酶的活性也显著升高。
综上，上述数据表明，仅通过更换细胞培养基中的血清类型，即可构建出在形态学、代谢特征及功能层面均与原代肝细胞高度相似的细胞类型。
实验设计：本研究对两类样本进行了检测，一类为胎牛血清培养的Huh7.5细胞，另一类为转入人血清培养基后分别于第8天、15天、23天收获的Huh7.5细胞。所有实验组均设置3次生物学重复。