State Key Laboratory of Biotherapy and Cancer Center/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, 610041, People's Republic of China.
Center for Cell Lineage and Atlas (CCLA), Bioland Laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory), Guangzhou, 510005, China.
Stem Cell Res Ther. 2021 Jul 2;12(1):375. doi: 10.1186/s13287-021-02419-1.
Direct reprogramming of human fibroblasts to hepatocyte-like cells was proposed to generate large-scale functional hepatocytes demanded by liver tissue engineering. However, the difficulty in obtaining large quantities of human fibroblasts greatly restricted the extensive implementation of this approach. Meanwhile, human umbilical cord mesenchymal stem cells (HUMSCs) are the preferred cell source for HLCs with the advantages of limited ethical concerns, easy accessibility, and propagation in vitro. However, no direct reprogramming protocol for converting HUMSCs to hepatoblast-like cells (HLCs) has been reported.
HLCs were successfully generated from HUMSCs by forced expression of FOXA3, HNF1A, and HNF4A (collectively as 3TFs) and c-Myc. In vitro and in vivo functional experiments were conducted to demonstrate the hepatic phenotype, characterization, and function of HUMSC-derived HLCs (HUMSC-iHeps). ChIP-seq and RNA-seq were integrated to reveal the potential molecular mechanisms underlying c-Myc-mediated reprogramming.
We showed that c-Myc greatly improved the trans-differentiation efficiency for HLCs from HUMSCs, which remained highly efficient in reprogramming fibroblasts into HLCs, suggesting c-Myc could promote direct reprogramming and its potentially widespread applicability for generating large amounts of HLCs in vitro. Mice transplantation experiments further confirmed the therapeutic potential of HUMSC-iHeps by liver function restoration and survival prolongation. Besides, in vivo safety assessment demonstrated the low risk of the tumorigenic potential of HUMSC-iHeps. We found that c-Myc functioned predominantly at an early phase of reprogramming, and we further unraveled the regulatory network altered by c-Myc.
c-Myc enhanced reprogramming efficiency of HLCs from HUMSCs. A large scale of functional HLCs generated more conveniently from HUMSCs could benefit biomedical studies and applications of liver diseases.
通过直接重编程将人成纤维细胞转化为肝细胞样细胞，以生成大量用于肝组织工程的功能性肝细胞，这一方法被提出。然而，大量获得人成纤维细胞的困难极大地限制了这种方法的广泛实施。同时，人脐带间充质干细胞（HUMSCs）因其伦理问题有限、易于获得和体外增殖等优势，成为肝细胞样细胞（HLCs）的首选细胞来源。然而，目前尚无将 HUMSCs 直接重编程为肝前体细胞（HLCs）的报道。
通过强制表达 FOXA3、HNF1A 和 HNF4A（统称为 3TFs）和 c-Myc，我们成功地将 HLCs 从 HUMSCs 中诱导生成。通过体外和体内功能实验，证明了 HUMSC 来源的 HLC（HUMSC-iHeps）的肝表型、特征和功能。整合 ChIP-seq 和 RNA-seq 揭示了 c-Myc 介导的重编程的潜在分子机制。
我们发现 c-Myc 极大地提高了 HUMSCs 向 HLCs 的转分化效率，并且在将成纤维细胞重编程为 HLCs 时仍然保持着高效性，提示 c-Myc 可以促进直接重编程，并且可能具有广泛的适用性，可用于体外大量生成 HLCs。小鼠移植实验进一步证实了 HUMSC-iHeps 通过恢复肝功能和延长生存时间的治疗潜力。此外，体内安全性评估表明 HUMSC-iHeps 的致瘤潜力风险较低。我们发现 c-Myc 在重编程的早期阶段起主要作用，并且我们进一步揭示了 c-Myc 改变的调控网络。
c-Myc 增强了 HUMSCs 向 HLCs 的重编程效率。更方便地从 HUMSCs 中生成大量功能性 HLCs 可以有益于生物医学研究和肝脏疾病的应用。
