Fig 1.
Conceptual illustration of iPSC-hepatocyte fiber technology.
Human iPSC-hepatocytes are encapsulated in a 3D ECM-rich microenvironment using the cell fiber technology, and the iPSC-hepatocyte fibers are applicable to cell therapy for liver failure and drug development.
Fig 2.
Cell encapsulation and 3D cultivation of human iPSC-hepatocytes in a core-shell hydrogel fiber.
(A) Human iPSC-hepatocytes singly dissociated and diluted into Matrigel were encapsulated into the core region of the fiber at a density of 1 × 108 cells / mL, using the cell fiber technology. (B) The iPSC-hepatocytes were cultured for 7 days in the ECM-rich 3D microenvironment, and compacted cell aggregates were acquired in the fiber.
Fig 3.
Characterization of the hepatic function of the encapsulated hepatocytes.
(A) Immunocytochemistry was performed for hepatic marker proteins 7 days after hepatocyte cultivation. The iPSC-hepatocytes were positive for albumin and ASGPR1. (B) ELISA was performed for quantifying the albumin secreted into the culture medium. Secreted albumin was detected and its contents increased in the fibers during the culturing process (n = 3, P < 0.07). The error bars represent the standard deviation (s.d.) of triplicate samples. (C.) Quantitative RT-PCR was performed for the hepatic marker genes, ALB, and HNF4α. The expression of the marker genes in the hepatocytes from the cell fiber culture was significantly upregulated, compared to those from the conventional spheroid and 2D culture conditions (n = 3, *; P<0.05), **; P<0.01). The error bars represent the s.d. of triplicate samples.
Fig 4.
Evaluation of the hepatic gene expression profile of the iPSC-hepatocytes cultured in the cell fibers.
(A) qRT-PCR was performed for marker genes of the developmental stage of hepatocytes (n = 3, *; P<0.05), **; P<0.01). The error bars represent the s.d. of triplicate samples. (B) qRT-PCR was performed for quantifying the CYP family genes, which encode the CYP enzymes for drug metabolism (n = 3, *; P<0.05), **; P<0.01). The error bars represent the s.d. of triplicate samples.
Fig 5.
Application of the iPSC-hepatocytes cell fibers as transplantable grafts.
(A) The fibers were easily picked up and collected in optimal shapes for transplantation. (B) The fibers were transplanted into the mouse abdominal cavity. (C) Human albumin was detected in the peripheral blood samples from the transplanted mice by ELISA (n = 3). The error bars represent the s.d. of triplicate samples. N.D.; not-detected.