Abstract
BackgroundHepatocyte transplantation has been proposed as an effective treatment for patients with acute liver failure (ALF), but its application is limited by a severe shortage of donor livers. Human pluripotent stem cells (hPSCs) have emerged as a potential cell source for regenerative medicine. Human amniotic epithelial stem cells (hAESCs) derived from amniotic membrane have multilineage differentiation potential which makes them suitable for possible application in hepatocyte regeneration and ALF treatment.MethodsThe pluripotent characteristics, immunogenicity, and tumorigenicity of hAESCs were studied by various methods. hAESCs were differentiated to hepatocyte-like cells (HLCs) using a non-transgenic and three-step induction protocol. ALB secretion, urea production, periodic acid-Schiff staining, and ICG uptake were performed to investigate the function of HLCs. The HLCs were transplanted into ALF NOD-SCID (nonobese diabetic severe combined immunodeficient) mouse, and the therapeutic effects were determined via liver function test, histopathology, and survival rate analysis. The ability of HLCs to engraft the damaged liver was evaluated by detecting the presence of GFP-positive cells.ResultshAESCs expressed various markers of embryonic stem cells, epithelial stem cells, and mesenchymal stem cells and have low immunogenicity and no tumorigenicity. hAESC-derived hepatocytes possess the similar functions of human primary hepatocytes (hPH) such as producing urea, secreting ALB, uptaking ICG, storing glycogen, and expressing CYP enzymes. HLC transplantation via the tail vein could engraft in live parenchymal, improve the liver function, and protect hepatic injury from CCl4-induced ALF in mice. More importantly, HLC transplantation was able to significantly prolong the survival of ALF mouse.ConclusionWe have established a rapid and efficient differentiation protocol that is able to successfully generate ample functional HLCs from hAESCs, in which the liver injuries and death rate of CCl4-induced ALF mouse can be significantly rescued by HLC transplantation. Therefore, our results may offer a superior approach for treating ALF.
Highlights
Hepatocyte transplantation has been proposed as an effective treatment for patients with acute liver failure (ALF), but its application is limited by a severe shortage of donor livers
In Hepg2 group, many colonies were visible after 30 days; no colony was observed in the Human amniotic epithelial stem cells (hAESCs) group at the same time (Fig. 2a)
Our results demonstrated that the hepatocyte-like cells (HLCs) exhibited CYP3A4, CYP1A2, CYP7A1, and CYP2B6 activities similar to that found in human primary hepatocytes (hPH) and that the expression level of the enzymes was remarkably higher than hAESCs (Fig. 4b)
Summary
Hepatocyte transplantation has been proposed as an effective treatment for patients with acute liver failure (ALF), but its application is limited by a severe shortage of donor livers. Human amniotic epithelial stem cells (hAESCs) derived from amniotic membrane have multilineage differentiation potential which makes them suitable for possible application in hepatocyte regeneration and ALF treatment. Human pluripotent stem cells (hPSCs) have the capability of self-renewal and differentiating into derivates of the three embryonic layers, which make them suitable candidates for possible application in hepatocyte regeneration and ALF treatment [10, 11]. There are still many questions remain unanswered about the use of stem cells to treat ALF [18], including tumorigenicity, immune incompatibility between donor and receptor, and ethical issues [19, 20] To overcome these shortfalls, the generation of functional hepatocyte from new source of stem cells is necessary. These observations suggest that the hAESCs might be ideal candidates for regenerative medicine and tissue repair
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