Page 134 - IJB-10-5

International Journal of Bioprinting Liver printing: from structure to application

and acute liver failure. However, there is currently a lack hepatocytes into expandable hepatocyte-derived liver
of robust evidence supporting the long-term therapeutic progenitor-like cells (HepLPCs). This cell line exhibits
effects of hepatocyte transplantation in clinical practice. significantly improved protein synthesis, urea generation,
Additionally, many unresolved issues remain, including ammonia clearance, and hepatocyte growth factor secretion
obtaining quality-assured hepatocytes, improving the compared to traditional artificial hepatocytes, representing
efficiency of transplantation and engraftment, and a new, sustainably expandable, functional human
developing effective immunosuppression protocols. 63 hepatocyte line (Figure 4A). Additionally, overexpression
of FOXA3 in immortalized human HepLPCs (iHepLPCs)
3.2. Bioartificial liver further enhances liver function and results in a more mature
Artificial liver support systems are extracorporeal hepatocyte morphology (Figure 4B). However, its expansion
devices designed to prolong the survival of patients with time is approximately 2 weeks, and it lacks validation in
liver failure by supporting liver regeneration (bridge human experiments, with cell functions inferior to primary
to regeneration) or patients until suitable organs are hepatocytes. Hui et al. achieved the large-scale expansion
72
available for orthotopic liver transplantation (bridge to (×10 ) of functionally proliferative hepatocytes (human-
9
transplantation). Artificial liver support systems include induced hepatocytes, hiHep) by overexpressing liver
non-biological and biological artificial livers. Non- transcription factors (FOXA3, HNF1A, and HNF4A) in
64
biological artificial livers primarily rely on chemical and human fibroblasts and subsequently established a serum-
physical methods, such as filtration or adsorption, to free culture system suitable for differentiating hepatocytes,
remove metabolic toxins. In addition to removing water- thereby stably producing GMP-grade hiHep cells, which
soluble substances, non-biological artificial livers can also have entered the preclinical stage. 73,74 Overall, high-quality
clear lipophilic substances and ALB-bound substances, seed cells are a focal point in the development of biological
such as bilirubin, bile acids, metabolites of aromatic amino artificial livers.
acids, medium-chain fatty acids, and cytokines. 65,66 Besides
detoxification, the liver is also involved in synthesizing 3.3. Liver organoids
proteins such as ALB and clotting factors, storing vitamins Organoids are cell-based in vitro models that mimic
and glycogen, metabolizing fats and hormones, secreting the structure and function of in vivo tissues. They can
bile, and maintaining the balance of the body’s internal be utilized for studying the fundamental mechanisms
environment. The complex mechanisms that uphold of biological development, regeneration, and repair, as
internal homeostasis and ensure bodily equilibrium are well as for applications in diagnostics, disease modeling,
unlikely to be fully replaced by standalone non-biological drug discovery, and personalized medicine. Typically,
detoxification methods. Non-biological artificial livers organoids are derived from embryonic or adult cells,
cannot substitute for the biological functions of the liver. including progenitor or differentiated cells from healthy
Therefore, research has shifted towards biological artificial or diseased tissues. 75–78 In the field of liver organoids,
liver support systems. commonly used hepatocyte types include those derived

Biological artificial livers contain hepatocytes cultured from induced pluripotent stem cells (iPSCs) or embryonic
from artificial sources, which not only possess detoxification stem cells (ESCs), as well as liver progenitor cells from
67
functions but also synthetic and metabolic functions. tissues. Primary hepatocytes can also be used for the
79
Currently, the hepatocytes used in various biological artificial culturing of liver organoids. Huch et al. demonstrated
livers mainly include primary human hepatocytes, porcine that Lgr5+ cells isolated from damaged mouse liver
hepatocytes, and human hepatoblastoma lines. However, bile ducts could be expanded and differentiated into
primary human hepatocytes have limited proliferative liver organoids in a culture medium supplemented with
80
capacity in vitro; porcine hepatocytes are challenged by Rspo1. Similarly, in 2015, they obtained similar results
issues related to xenogenic viruses and immune barriers 68,69 ; using EpCAM+ cholangiocytes from human liver tissue
and human hepatoblastoma lines lack some liver functions, samples. Gene expression profiling demonstrated that liver
such as normal urea cycle enzymes and limited metabolic organoids expressed high levels of hepatocyte markers,
functions, making them less than ideal seed cells. Chen such as ALB, cytokeratin, apolipoprotein B (APOB), and
70
et al. used human induced pluripotent stem cell (hiPSC)- complement factor C3, and they were capable of glycogen
derived hepatocytes, which can be expanded, cryopreserved accumulation and low-density lipoprotein (LDL) uptake.
81
in vitro, and further matured into functional liver organoids Vyas et al. utilized human fetal liver progenitor cells to
at a large scale. However, the induction and differentiation self-organize into liver organoids, which demonstrated
process is complex, requiring a differentiation period the process of liver organogenesis and gradually formed
82
exceeding 38 days. Li et al. used chemical reprogramming differentiated hepatocytes and bile duct structures. Liver
71
technology to transform mouse and human primary organoids derived from human fetal liver progenitor cells
Volume 10 Issue 5 (2024) 126 doi: 10.36922/ijb.3819