Establishment of Novel Detection System for Embryonic Stem Cell-Derived Hepatocyte-Like Cells Based on Nongenetic Manipulation with Indocyanine Green

Abstract

Hepatocytes derived from embryonic stem cells (ESCs) are expected to be useful for basic research and clinical applications. However, in several studies, genetic methods used to detect and obtain them are difficult and pose major safety problems. Therefore, in this study, we established a novel detection system for hepatocytes by using indocyanine green (ICG), which is selectively taken up by hepatocytes, based on nongenetic manipulation. ICG has maximum light absorption near 780 nm, and it fluoresces between 800 and 900 nm. Making use of these properties, we developed flow cytometry equipped with an excitation lazer of 785 nm and specific bandpass filters and successfully detected ESC-derived ICG-positive cells that were periodic acid-Schiff positive and expressed hepatocyte phenotypic mRNAs. These results demonstrate that this detection system based on nongenetic manipulation with ICG will lead to isolate hepatocytes generated from ESCs and provide the appropriate levels of stability, quality, and safety required for cell source for cell-based therapy and pharmaceutical studies such as toxicology.

5-azacytidine promotes terminal differentiation of hepatic progenitor cells

“…A the identification of differentiated hepatocytes in vitro (20,21). The ratios of ICG-positive cells to ICG-negative cells in the non-induced control group and in the group treated with 5-azaC alone, were 3.9±2.8% and 4.8±1.9%, respectively, and no significant difference was detected between the two groups.…”

Hepatocyte growth factor‐induced differentiation of bone mesenchymal stem cells toward hepatocyte‐like cells occurs through nuclear factor‐kappa B signaling in vitro

“…The indocyanine green (ICG) uptake assay determines hepatic function, and we used this assay to establish the successful differentiation of BMSCs into hepatocyte‐like cells as previously described (Yoshie et al, ). Briefly, 5 mg/mL ICG was dissolved in sterile phosphate buffered saline (PBS), which was diluted fivefold with DMEM with low glucose containing 10% FBS.…”