人胚胎干细胞来源的肝细胞的体外成熟及机制研究

人工肝技术有望成为肝移植的有效替代疗法，其应用的主要瓶颈在于如何短时间大量获得功能成熟的肝细胞。人多能干细胞是再生医学的重要细胞资源，然而目前人多能干细胞来源的肝细胞仅具有有限的白蛋白合成量、尿素生成量及药物代谢能力，其更类似于胎儿或新生儿时期的肝细胞，而非成人肝细胞。如何促进其进一步成熟达到与成人肝细胞功能相当的状态仍是一个悬而未决的问题。申请人前期研究中实现了二维培养条件下维持原代肝细胞的体外长期培养并揭示了机械张力导致的去分化机制。该研究提示了二维培养条件下获得成熟肝细胞需要抑制“机械张力导致的去分化”。以此为基础，申请人拟将肌动蛋白聚合抑制剂Latrunculin B和非肌肉肌球蛋白II ATPase抑制剂Blebbistatin作为基础，通过叠加筛选策略发现活性小分子促进人胚胎干细胞来源的肝细胞的成熟，从而建立促进肝细胞功能成熟的培养体系，并对肝细胞成熟的分子机制进行研究。

Artificial liver technology is expected to become a promising alternative therapy for liver transplantation, and the main bottleneck of its application lies in how to obtain a large number of mature hepatocytes in a short time. Human pluripotent stem cells (hPSCs) are important cell resources in regenerative medicine. However, the hepatocytes derived from hPSCs now only have limited albumin synthesis, urea production and drug metabolism capacity, which are more similar to fetal or neonatal hepatocytes than adult hepatocytes. How to promote the further maturation of the hepatocytes derived from hPSCs to achieve the same function as adult hepatocytes is still an open question. In the previous study, we achieved the long-term culture of primary hepatocytes in vitro under two-dimensional culture condition, and revealed the mechanism of dedifferentiation caused by mechanical tension. This study suggests that it is necessary to inhibit the dedifferentiation induced by mechanical tension for obtaining mature hepatocytes under two-dimensional culture condition. Based on it, we intend to use actin polymerization inhibitor latrunculin B and non-muscle myosin II ATPase inhibitor Blebbistatin as the basis, and find active small molecules that can promote the maturation of hepatocytes derived from human embryonic stem cells by iterative screening strategy, so as to establish a culture system which can promote the functional maturation of hepatocytes, and study the molecular mechanism of hepatocytes maturation.