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iPS to hepatocyte differentiation system iPS Cell to Hepatocyte Differentiation System
Technical notes Hepatocyte generation from various iPSC sources
Human induced pluripotent stem cell derived hepatocytes Human stem cell-derived hepatocytes
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iPS to hepatocyte differentiation system iPS Cell to Hepatocyte Differentiation System
Technical notes Hepatocyte generation from various iPSC sources
Human induced pluripotent stem cell derived hepatocytes Human stem cell-derived hepatocytes

iPS cell to hepatocyte differentiation overview

Understanding hepatotoxicity of new drug candidates is one of the major challenges in drug discovery. Early detection of toxicity and adverse reactions could decrease the high cost of drug development and significantly improve patient safety. Therefore, the need exists for more predictive and physiologically relevant human cellular models that can improve preclinical toxicity testing and reduce the number of late-stage clinical failures.

To date, primary human hepatocytes have been considered the gold standard for this evaluation as they provide a model to assess key criteria such as drug transport, metabolism, clearance, and in vitro toxicity. However, availability of these cells can be limiting. Moreover, large batches of primary human hepatocytes can be unreliable: they can display variable quality, show large donor variation, and lose functionality very rapidly—even within a few days of in vitro culture.


Stem cell research advancements offer new options

Hepatocytes derived from human induced pluripotent stem (hiPS) cells are a powerful alternative to primary human hepatocytes, as they offer a virtually unlimited source of cells, are easy to culture, and provide reliable and reproducible starting material for drug metabolism studies and toxicity testing.

Cellartis iPS cell-derived hepatocytes show similar functionality to primary human hepatocytes, and their functionality is stable over a prolonged period of time in culture. These hiPS cell-derived hepatocytes express major drug-metabolizing enzymes and transporters and show high specificity to known hepatotoxic compounds.

The Cellartis iPS Cell to Hepatocyte Differentiation System is a reproducible and robust system with a universal protocol to differentiate any hiPS cell line into hepatocytes. Optimized for use with any disease-relevant hiPS cell line, this system allows researchers to generate panels of hiPS cell-derived hepatocytes that capture the heterogeneity of a patient population or to utilize their own patient-derived hiPS cells to generate customized liver disease models.

Cellartis iPS Cell to Hepatocyte Differentiation System

 

Generate consistent, functional hiPS cell-derived hepatocytes

The process of generating hiPS cell-derived hepatocytes begins with the directed differentiation of hiPS cells into definitive endoderm (DE) cells, which are then differentiated further into hepatocytes. The complete system provides media, supplements, and coating reagents for each step of the hiPS-cell-to-hepatocyte differentiation protocol. Starting with approximately 3 x 106 undifferentiated hiPS cells, this system yields 5 x 106 hepatocytes. Importantly, this do-it-yourself system offers a solution for the consistent production of assay-ready cells from patient-derived cells, or from Cellartis-brand iPS cell lines—enabling highly reproducible results.

Successful differentiation depends on the quality of the starting material; a homogeneous, undifferentiated stem cell population is ideal. The iPS Cell to Hepatocyte Differentiation System promotes high-quality starting material by incorporating DEF-CS culture system components, which are designed to support the robust and rapid generation of hiPS cells that maintain pluripotency and exhibit long-term genetic stability in feeder-free, defined conditions.

Using a protocol that combines media, supplements, and coating reagents optimized for 2D monolayer culture, pluripotent cells are differentiated into definitive endoderm using a seven-day protocol. In the final stage, the homogeneous DE cells are further differentiated into hepatocytes, and functional hepatocytes are available for experiments within three weeks.

The standardized protocol of the system maintains high-quality cells from start to finish, generating highly homogeneous hepatocyte cells that initially exhibit high expression levels of early hepatic markers (HNF4α and CK18). As the cells mature in culture, they express the liver-specific markers CYP3A and albumin (Figure 1).

Immunocytochemical analysis of hepatocyte differentiation.

Figure 1. Immunocytochemical analysis of hepatocyte differentiation. hiPS cells were differentiated into functional hepatocytes using the Cellartis iPS Cell to Hepatocyte Differentiation System. Immunostaining of hepatocytes was performed to detect early hepatic markers HNF4α (red, nuclear) and CK18 (green) on Days 14 and 21. As the hepatocytes matured, expression of liver-specific markers CYP3A (red, cytoplasmic) and albumin (green) increased as seen on Day 28, as expected. Cell nuclei were stained with DAPI (blue).

 

hiPS cell-derived hepatocytes exhibit high CYP expression and activity

Hepatocytes derived using the iPS Cell to Hepatocyte Differentiation System express several key cytochrome P450 genes (CYP1A, CYP3A, and CYP2C9) that play an important role in drug metabolism. These cells also express AFP (an endoderm marker), albumin (a hepatocyte maturation marker), and PXR (a nuclear receptor important for CYP induction). The activities of these factors indicate that these hiPS cell-derived hepatocytes function similarly to primary hepatocytes, illustrating their usefulness in drug metabolism studies (Figure 2 and Figure 3).

mRNA expression analysis indicates expression of key cytochrome P450 genes in hepatocytes generated using the iPS Cell to Hepatocyte Differentiation System protocol.

Figure 2. mRNA expression analysis indicates expression of key cytochrome P450 genes in hepatocytes generated using the iPS Cell to Hepatocyte Differentiation System protocol. The graphs illustrate expression levels of several hepatocyte-specific genes (analyzed by RT‑qPCR) over days 11–32 in the differentiation process.

 

CYP enzyme activity of hiPS cell-derived hepatocytes.

Figure 3. CYP enzyme activity of hiPS cell-derived hepatocytes. The graph indicates CYP enzyme activity of hepatocytes from differentiation Days 21–32, as measured by cytochrome P450 activity assay. Formation of the metabolites of acetaminophen (by CYP1A), 1-OH-Midazolam (by CYP3A), and 4-OH-Diclofenac (by CYP2C9) was analyzed.

 

Functional, disease-specific hepatocytes from patient-derived cell lines

The Cellartis iPS Cell to Hepatocyte Differentiation System provides a complete, do-it-yourself solution for directed differentiation of disease-specific or patient-specific hiPS cells into hepatocytes by combining a standardized protocol with optimized, ready-to-use media, supplements, and coating reagents. Takara Bio also offers accessory reagents, cells, and systems—plus expert technical support—designed to meet your research or clinical needs.

This novel differentiation system simplifies the production of large panels of hiPS cell-derived, functional hepatocytes with your desired genotypes/phenotypes for disease modeling, drug discovery, drug metabolism research, and hepatotoxicity studies.

  • Highly reproducible, robust system—the same protocol has been shown to work across 25 different hiPS cell lines. There is no need to optimize for your lines.
  • Ideal for drug metabolism and safety studies—consistently generate panels of functional, hiPS cell-derived hepatocytes with diverse genetic backgrounds.
  • Customized starting materials—start with any disease-relevant hiPS cell lines and create accurate liver disease models.

iPS and ES differentiation to hepatocytes

Directed differentiation to hepatocytes

Make hepatocytes from your own patient- or disease-specific iPS or ES cell lines

Our 15+ years of experience with endodermal lineage differentiation means you can count on us to deliver high-quality hepatocytes.

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